Iron(III) complexes with novel contrast agent properties for magnetic resonance imaging.

Abstract

The present invention provides novel iron(III) complexes having formulas (1) and (2). Additionally, the corresponding ligands having formulas (L1) and (L2) are disclosed for preparing the iron(III) complexes. TIFF2022552705000030.tif104170

Description

[Technical field]

[0001] The present invention relates to a novel iron(III) complex, in particular an iron(III) complex suitable as a contrast agent for magnetic resonance imaging, and further discloses a ligand for preparing said iron(III) complex. [Background technology]

[0002] Low molecular weight gadolinium-based contrast agents (GBCAs) have become the standard for improving the validity of magnetic resonance imaging (MRI) examinations in all areas of medicine. In Germany, more than 10.8 million magnetic resonance imaging (MRI) examinations were performed in 2015 (de.statista.com). In about one third of these MRI examinations, GBCAs were administered intravenously for T1-weighted images, which resulted in improved diagnosis in cases of acute and chronic inflammation, tumors, angiogenesis, etc. In this regard, GBCAs accelerate both T1 relaxation (spin-lattice relaxation) and T2 relaxation (spin-spin relaxation), which leads to an increase in the diagnostically useful MRI signal in the corresponding T1-weighted images. Free gadolinium ions are toxic (see, for example, dos Remedios CG, “Lanthanide ion probes of calcium-binding sites on cellular membranes”, Cell Calcium, 1981;2(1):p.29-51, and Bower DV et al., “Gadolinium-Based MRI Contrast Agents Induce Mitochondrial Toxicity and Cell Death in Human Neurons,and Toxicity Increases With Reduced Kinetic Stability of the Agent”, Investigative Radiology, 2019;54(8):p.453-463). Gadolinium ions, due to their similar chemical behavior to calcium ions, can be taken up, for example, by the liver and bone system, where they can remain for years. Due to the close ionic radius between calcium and gadolinium, gadolinium may act as a calcium antagonist and inhibit, for example, myocardial contractility and the coagulation system (Darrah TH et al., “Incorporation of excess gadolinium into human bone from medical contrast agents”, Metallomics, 2009;1(6):p.479-488). Although gadolinium ions are toxic, GBCAs are approved in approved drugs, provided that they are firmly fixed to complexing agents (e.g. DTPA (diethylenetriaminepentaacetic acid) and DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) and are completely excreted together with the complexing agents in a short time, mainly via the kidneys.

[0003] However, it has become clear that in some circumstances, elimination is not complete. For example, about 12 years ago, a condition known as nephrogenic systemic fibrosis was recognized as a serious side effect of incomplete elimination of gadolinium contrast agents administered to patients with impaired renal function (Grobner T., “Gadolinium-a specific trigger for the development of nephrogenic fibrosing dermopathy and nephrogenic systemic fibrosis?”, Nephrol Dial Transplant. 2006; 21(4): p. 1104-1108, and Idee JM et al., “The role of gadolinium chelates in the mechanism of nephrogenic systemic fibrosis: A critical update”, Crit Rev Toxicol. 2014; 44(10): p. 895-913). Results from various animal studies have shown that gadolinium can still be detected in skin biopsies of rats one year after administration of GBCA (Pietsch H et al., “Long-term retention of gadolinium in the skin of rodents following the administration of gadolinium-based contrast agents”, Eur Radiol 2009;44(4):p.226-233). It has been shown that the degree of gadolinium accumulation depends on the type of complexing agent, especially the stability of the gadolinium complex. In particular, repeated administration of low molecular weight contrast agents, such as those required for breast tumor screening or monitoring the progression of multiple sclerosis, has been shown to deposit in healthy human brain tissue (see, for example, McDonald RJ et al., “Intracranial Gadolinium Deposition after Contrast-enhanced MR Imaging”, Radiology 2015;275(3):p.772-782). Using ICP-MS, it has also been shown that gadolinium deposition of the more stable macrocyclic GBCAs is also observed in samples from deceased patients, particularly in bone and brain (Murata N et al., “Macrocyclic and Other Non-Group 1 Gadolinium Contrast Agents Deposit Low Levels of Gadolinium in Brain and Bone Tissue:Preliminary Results From 9 Patients With Normal Renal Function”, Invest Radiol 2016;51(7):p.447-453).

[0004] In the past, it was also found that low molecular weight iron complexes, especially Fe-DTPA and Fe-tCDTA (tCDTA = trans-cyclohexanediaminetetraacetic acid), can be used as MRI contrast agents. The relaxivity of these iron complexes is lower than that of GBCAs, but this disadvantage can be overcome by using higher doses (see, for example, White DL et al., "IRON(III) COMPLEXES AS MRI CONTRAST AGENTS", Proc. Int. Soc. Magn. Reson. Med. 1985; 1985(S2): p. 906-909). Due to the large amount of endogenous iron (about 4 g in adults) and the systems available for its absorption, storage and transport, it can be assumed that the long-term toxicity of iron complexes is less than that of GBCAs. Furthermore, iron oxide nanoparticles (which can be used as contrast agents or for iron replacement therapy) are generally well tolerated and remain almost entirely in the body after intravenous administration, in contrast to low molecular weight contrast agents, which are almost completely excreted. Iron complexes have also been shown to be suitable for typical current applications, such as dynamic contrast-enhanced MRI (DCE-MRI) used for breast diagnosis (Boehm-Sturm P et al., “Low-Molecular-Weight Iron Chelates May Be an Alternative to Gadolinium-based Contrast Agents for T1-weighted Contrast-enhanced MR Imaging”, Radiology 2017;286(2):p.537-546).

[0005] US Pat. No. 5,362,475 describes, inter alia, the preparation of iron-based contrast agents, specifically the sodium or N-methylglucamine salts of complexes of iron(III) with the tCDTA ligand. Gestin et al. (Nucl. Med. Biol., 1993; 20(6): 755-762) describe, inter alia, the synthesis of N-[methyl(2-aminoethyl)carbamido]-trans-1,2-diaminocyclohexane-N,N',N'-triacetic acid as a novel bifunctional chelator for indium-labeled antibodies. Philipp Boehm-Sturm et al., “Low-Molecular-Weight Iron Chelates May Be An Alternative to Gadolinium-based Contrast Agents for T1-weighted Contrast-enhanced MR Imaging”, RADIOLOGY, Vol. 286, No. 2, February 2018, p. 537-546, discloses Fe-DTPA complexes and their use as contrast agents for magnetic resonance imaging. Mane R. Brandt et al., “ 52g / 55“Mn-Labelled CDTA-based trimeric complexes as novel bimodal PET / MR probes with high relaxivity”, Dalton Trans., 2019, Vol. 48, p. 3003-3008 describes Mn-CDTA complexes that may be used as contrast agents in magnetic resonance imaging. Summary of the Invention [Problem to be solved by the invention]

[0006] Therefore, there remains a need for alternative, gadolinium-free contrast agents. Furthermore, it is desirable for the contrast agents to exhibit pH-dependent resonance behavior so that new diagnostic possibilities arise. [Means for solving the problem]

[0007] The present invention provides novel iron(III) complexes having the formulae (1) and (2) according to claim 1, [ka] During the ceremony, m=1 to 5; Y 1 and Y 2 are independently selected from the group consisting of O, NH and NR, where R represents C1-C15 alkyl or C3-C10 cycloalkyl; Y 3 is selected from the group consisting of O, NH and NR, where R represents C1-C15 alkyl or C3-C10 cycloalkyl; Y 4 are OH, OR, NH2, NHR and NR 5 R 6 wherein R, R 5 and R 6 represents C1-C15 alkyl or C3-C10 cycloalkyl, and R 5 and R 6 are selected independently of each other, Z represents a bridge according to one of the formulas (3) to (10), [ka] During the ceremony, n=0 to 4, o=0 to 4, * and *' are Y in formula (1). 1 and Y 2 Or Y in formula (2) 3 and Y 4 represents the site of attachment of bridge Z to R 1 , R 2 , R 3 and R 4 are independently selected from the group consisting of H, C1-C15 alkyl, and C3-C10 cycloalkyl.

[0008] In a further aspect, the present invention relates to the use of iron(III) complexes having formula (1) or (2) in in vivo diagnostic methods, in particular as contrast agents for magnetic resonance imaging, and in the preparation of magnetic resonance imaging contrast agents containing said iron(III) complexes.

[0009] Furthermore, the present invention provides novel ligands having the formula (L1) or (L2) as intermediates which can be converted into the above complexes with iron(III), [ka] During the ceremony, m=1 to 5; Y 1 and Y 2 are independently selected from the group consisting of O, NH and NR, where R represents C1-C15 alkyl or C3-C10 cycloalkyl; Y 3 is selected from the group consisting of O, NH and NR, where R represents C1-C15 alkyl or C3-C10 cycloalkyl; Y 4 are OH, OR, NH2, NHR and NR 5 R 6wherein R, R 5 and R 6 represents C1 to C15 alkyl or C3 to C10 cycloalkyl, and R 5 and R 6 are selected independently of each other, Z represents a bridge according to one of the formulas (3) to (10), [ka] During the ceremony, n=0 to 4, o=0 to 4, * and *' are Y in formula (1). 1 and Y 2 Or Y in formula (2) 3 and Y 4 represents the site of attachment of bridge Z to R 1 , R 2 , R 3 and R 4 are each independently selected from the group consisting of H, C1-C15 alkyl, and C3-C10 cycloalkyl; However, Z is * -CH2CH2- *’ However, the ligand having the formula (L2) is not limited thereto.

[0010] Further preferred embodiments of the invention are given in the following description. The invention will now be described with the aid of exemplary embodiments and the accompanying drawings. [Brief description of the drawings]

[0011] [Figure 1] 1 shows the pH dependence of the T1 relaxivity and T2 relaxivity of the iron(III) complex (2-1). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0012] The present invention provides novel iron(III) complexes having the formulae (1) and (2) according to claim 1, [ka] During the ceremony, m=1 to 5; Y 1 and Y 2 are independently selected from the group consisting of O, NH and NR, where R represents C1-C15 alkyl or C3-C10 cycloalkyl; Y 3 is selected from the group consisting of O, NH and NR, where R represents C1-C15 alkyl or C3-C10 cycloalkyl; Y 4 are OH, OR, NH2, NHR and NR 5 R 6 wherein R, R 5 and R 6 represents C1-C15 alkyl or C3-C10 cycloalkyl, and R 5 and R 6 are selected independently of each other, Z represents a bridge according to one of the formulas (3) to (10), [ka] During the ceremony, n=0 to 4, o=0 to 4, * and *' are Y in formula (1). 1 and Y 2 Or Y in formula (2) 3 and Y 4 represents the site of attachment of bridge Z to R 1 , R 2 , R 3 and R 4 are independently selected from the group consisting of H, C1-C15 alkyl, and C3-C10 cycloalkyl. The stereochemistry of the two substituents at the asymmetric carbon atom of formula (1) or (2) marked with * can in principle be independent of each other, but preferably, the two substituents at the asymmetric carbon atom of formula (1) or (2) marked with * are in trans position with respect to each other. The bridges in formulae (4) to (10) have as a base structure a ring structure containing 4, 5 or 6 carbon atoms. The stereochemistry of the individual carbon atoms forming the ring structure is of secondary importance in this case and can therefore be of any stereochemistry. This also applies when the carbon atoms of the bridge in formula (3) are asymmetric. For example, when n+o=0 in formula (3), this means that Z is R 2 and an H substituent, meaning that it can be either the (R) or the (S) stereoisomer.

[0013] In a preferred embodiment of the present invention, Z contained in the iron(III) complex having the formula (1) or (2) is selected from the formulae (3) to (6): [ka] In the formula, n, o, R 1 , R 2 , R 3 and R 4 is as defined above. In the above formulas (4) to (10), the binding site of the bridge Z marked with * and *' is Y of formula (1). 1 and Y 2 Or Y in formula (2) 3 and Y 4 The substituents of Y in formula (1) may in principle be cis or trans relative to each other. 1 and Y 2 Group or Y of formula (2) 3 and Y 4 The groups are trans to each other at the attachment sites labeled * and *'. As an example, the group in formula (4) may be a 1,2-trans-cyclohexylene group. Particularly preferably, * and *' in formulae (4) to (10) are therefore in the trans position relative to each other. In a preferred embodiment of the iron(III) complex having formula (1) or (2), Z comprises a group selected from N,N'-1,2-trans-cyclohexylene, N,N'-1,3-trans-cyclohexylene, N,N'-1,4-trans-cyclohexylene, N,N'-1,3-trans-cyclopentylene, N,N'-1,2-trans-cyclobutylene, N,N'-1,3-trans-cyclobutylene and N,N'-1,2-trans-cyclopentylene.

[0014] According to a further embodiment in which the iron(III) complex has formula (1), m=2. More preferably, the iron(III) complex corresponds to formula (1) 1 and Y 2 represents NH. Even more preferably, the iron(III) complex corresponds to formula (2) 3 represents NH, and Y 4 represents OH, NH2 or NHR.

[0015] Particularly preferred exemplary embodiments of iron(III) complexes having formula (1) or formula (2) include the following compounds: Z has the formula (4) to (6), where R 1 =R 2 =R 3 =R 4 =H, or Z has the formula (5), where R 1 =R 2 =R 3 =H and R 4 =CH3, or Z has the formula (5), where R 1 =R 3 =R 4 =H and R 2 =CH3, or Z has the formula (8), where R 1 =R 2 =H and R 3 =CH3, or Z has the formula (8), where R2 =R 3 =H and R 1 =CH3.

[0016] The iron(III) complexes having formulae (1) and (2) provided herein have polydentate ligands derived from cyclohexanediaminetetraacetic acid (CDTA), in particular from trans-cyclohexanediaminetetraacetic acid (tCDTA), and are suitable, for example, as contrast agents for in vivo diagnostic methods, in particular for magnetic resonance imaging (MRI). Compared to Fe-tCDTA, the iron(III) complexes show, inter alia, improved heat and acid resistance. In this regard, preliminary tests have shown that upon heat sterilization in an autoclave (121° C., 15 min; 1 bar; 0.43 mol / L), Fe-tCDTA precipitates, whereas the iron(III) complexes remain in solution under the same conditions.

[0017] The iron(III) complex having formula (1) may have a higher relaxivity than, for example, Fe-tCDTA, and has an overall neutral charge compared to Fe-tCDTA (a complex with a single negative charge). Therefore, a low concentration of counter ion is required at neutral pH (if necessary). Therefore, the osmolality of the iron(III) complex having formula (1) per iron atom is greatly reduced. This is advantageous for acute toxicity, and furthermore, higher doses of the iron(III) complex having formula (1) can be tolerated when used in in vivo diagnostic methods. Although the lower relaxivity of the iron complex compared to the gadolinium complex must be compensated for by higher doses, the lower osmolality is a substantial advantage of the iron(III) complex having formula (1) compared to ionic iron complexes such as Fe-DTPA or Fe-tCDTA. In a preferred embodiment, m=2 in the iron(III) complex having formula (1). The iron(III) complex is therefore a dimer formed from two bridged tCDTA units.

[0018] Particularly preferred are iron(III) complexes having formulae (1-1) and (1-2). [ka]

[0019] The relaxivity of a special iron(III) complex having formula (2) shows a surprisingly strong dependence on pH. This strong pH dependence is observed in the pH range of 5-8, which is particularly attractive for medical imaging. As an example, tumors usually have a slightly more acidic pH than the range above. The pH dependence is due to the terminal Y in the bridge Z. 4 This is assumed to result from the protonation / deprotonation of the group.

[0020] The pH dependence is particularly pronounced for the iron(III) complex having the formula (2-1): [ka] In this case, at low pH, the terminal amino group is protonated, so that the central iron(III) ion is not available for coordination, and instead, coordination occurs with water as a ligand. In contrast, at high pH, ​​the free electron pair on the nitrogen contributes to the coordination of the central ion, so that water is excluded from the coordination site. The presence of water ligands in the complex leads to a substantial increase in T1 and T2 relaxivity, which in turn leads to improved contrast effect. Tumors and inflammation usually have a relatively low extracellular pH, which should lead to increased contrast. This pH dependence can be modeled for the iron(III) complex (2-1) as follows: [ka] This effect can also be exploited, for example, for Chemical Exchange Saturation Transfer (CEST) or Para-CEST imaging, in which coordinated water protons are imaged by CEST. To reach a concentration-independent pH, amide protons may additionally be imaged, for example by CEST, and calculated together with the water protons.

[0021] Preferred embodiments are iron complexes having the formulae (2-1) and (2-2). A particularly preferred embodiment is an iron complex having the formula (2-2): [ka]

[0022] The less flexible terminal amino group cannot form a complex with the central ion, thereby preventing pH-dependent deactivation at that site. 4 Groups, such as the amino groups of the iron(III) complexes having formulae (2-1) and (2-2), can further be used in chemical coupling reactions, for example to label biomolecules or other substances and thereby render them detectable by MRI. In this regard, the amino groups can be converted into functional groups suitable for the envisaged coupling reactions, such as isocyanate groups.

[0023] In the present invention, the term "C1-C15 alkyl" refers to a group represented by the general formula C n H 2n+1where n=1 to 15. As an example, "C1-C5 alkyl" includes methyl, ethyl, n-propyl, 2-propyl, n-butyl, 2-butyl, 2-methylpropyl, 2-methyl-2-propyl, n-pentyl, 2-pentyl, 2-methylbutyl, 3-methylbutyl, 2-methyl-2-butyl, 3-methyl-2-butyl, 2,2-dimethylpropyl and 3-pentyl. Examples of alkyl residues with n≧6 include n-hexyl, n-heptyl, n-octyl, n-nonyl, n-decyl, 2-hexyl, 2-heptyl, 2-octyl, 2-nonyl, 2-decyl, 2-undecyl, 3-hexyl, 3-heptyl, 3-octyl, 3-nonyl, 3-decyl, 3-undecyl, 3-dodecyl, 2-methylpentyl, 2-methylhexyl, 2-methylheptyl, 2-methyloctyl, 2-methylnonyl, 2-methyldecanyl, 2-ethylbutyl, 2-ethylpentyl, 2-ethylhexyl, 2-ethylheptyl, 2-ethyloctyl, 2-ethylnonyl, 2-ethyldecanyl, 2-methyl-2-pentyl, 2-methyl-2-hexyl, 2-methyl-2-heptyl, 2-methyl-2-octyl, 2-methyl 2-methyl-2-nonyl, 2-methyl-2-decyl, 2-methyl-2-undecyl, 3-methyl-2-pentyl, 3-methyl-2-hexyl, 3-methyl-2-heptyl, 3-methyl-2-octyl, 3-methyl-2-nonyl, 3-methyl-2-decyl, 3-methyl-2-undecyl, 3-ethyl-2-pentyl, 3-ethyl-2-hexyl, 3-ethyl-2-heptyl, 3-ethyl-2-octyl, 3-ethyl-2-nonyl, 3-ethyl-2-decyl, 3-ethyl-2-undecyl, 2-methyl-3-pentyl, 4-methyl-3-hexyl, 4-methyl-3-heptyl, 4-methyl-3-octyl, 4-methyl-3-nonyl, 4-methyl-3-decyl, 4-methyl-3-undecyl and 4-methyl-3-dodecyl.

[0024] In the present invention, furthermore, the term "C3-C10 cycloalkyl" should be understood to mean a monocyclic or bicyclic cycloalkyl residue containing 3 to 15 carbon atoms. Examples include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl. In this respect, the cycloalkyl residue may bear one or more substituents with a C1-C5 alkyl residue as defined above. Examples of substituted cycloalkyl residues of this type include 1-methyl-1-cyclopropyl, 1-methyl-1-cyclobutyl, 1-methyl-1-cyclopentyl, 1-methyl-1-cyclohexyl, 1-methyl-1-cycloheptyl, 2-methyl-1-cyclopropyl, 2-methyl-1-cyclobutyl, 2-methyl-1-cyclopentyl, 2-methyl-1-cyclohexyl, 2-methyl-1-cycloheptyl, 3-methyl-1-cyclobutyl, 3-methyl-1-cyclopentyl, 3-methyl-1-cyclohexyl, 3-methyl-1-cycloheptyl, 3-methyl-1-cyclobutyl, 3-methyl-1-cyclohexyl ... -1-cyclopentyl, 3-methyl-1-cyclohexyl, 3-methyl-1-cycloheptyl, 4-methyl-1-cyclohexyl, 4-methyl-1-cycloheptyl, 1,2-dimethyl-1-cyclopropyl, 2,2-dimethyl-1-cyclopropyl, 2,3-dimethyl-1-cyclopropyl, 1,2-dimethyl-1-cyclobutyl, 1,3-dimethyl-1-cyclobutyl, 2,2-dimethyl-1-cyclobutyl, 2,3-dimethyl-1-cyclobutene dimethyl-1-cyclobutyl, 2,4-dimethyl-1-cyclobutyl, 3,3-dimethyl-1-cyclobutyl, 1,2-dimethyl-1-cyclopentyl, 1,3-dimethyl-1-cyclopentyl, 2,2-dimethyl-1-cyclopentyl, 2,3-dimethyl-1-cyclopentyl, 2,4-dimethyl-1-cyclopentyl, 2,5-dimethyl-1-cyclopentyl, 3,3-dimethyl-1-cyclopentyl, 3,4-dimethyl-1-cyclopentyl, 1,2-dimethyl-1 -cyclohexyl, 1,3-dimethyl-1-cyclohexyl, 1,4-dimethyl-1-cyclohexyl, 1,5-dimethyl-1-cyclohexyl, 1,6-dimethyl-1-cyclohexyl, 2,2-dimethyl-1-cyclohexyl, 2,3-dimethyl-1-cyclohexyl, 2,4-dimethyl-1-cyclohexyl, 2,5-dimethyl-1-cyclohexyl, 2,6-dimethyl-1-cyclohexyl, 3,3-dimethyl-1-cyclohexyl, 3,4-Dimethyl-1-cyclohexyl, 3,5-Dimethyl-1-cyclohexyl, 3,6-Dimethyl-1-cyclohexyl, 4,4-Dimethyl-1-cyclohexyl, 1,2,2-Trimethyl-1-cyclopropyl, 1,2,3-Trimethyl-1-cyclopropyl, 1,2,2-Trimethyl-1-cyclobutyl, 1,3,3-Trimethyl-1-cyclobutyl, 1,2,3-Trimethyl-1-cyclobutyl, 2,2,3-Trimethyl-1-cyclobutyl, 2,2,4-Trimethyl-1-cyclobutyl, 1,2,2-Trimethyl-1-cyclopentyl, 1,2, 3-trimethyl-1-cyclopentyl, 1,2,4-trimethyl-1-cyclopentyl, 1,2,5-trimethyl-1-cyclopentyl, 1,3,3-trimethyl-1-cyclopentyl, 1,3,4-trimethyl-1-cyclopentyl, 1,3,5-trimethyl-1-cyclopentyl, 2,2,3-trimethyl-1-cyclopentyl, 2,2,4-trimethyl-1-cyclopentyl, 2,2,5-trimethyl-1-cyclopentyl, 2,3,3-trimethyl-1-cyclopentyl, 2,3,4-trimethyl-1-cyclopentyl, 2,3,5-trimethyl -1-cyclopentyl, 2,3,3-trimethyl-1-cyclopentyl, 2,4,4-trimethyl-1-cyclopentyl, 2,4,5-trimethyl-1-cyclopentyl, 2,5,5-trimethyl-1-cyclopentyl, 3,3,4-trimethyl-1-cyclopentyl, 3,3,5-trimethyl-1-cyclopentyl, 3,4,5-trimethyl-1-cyclopentyl, 3,4,4-trimethyl-1-cyclopentyl, 1,2,2-trimethyl-1-cyclohexyl, 1,2,3-trimethyl-1-cyclohexyl, 1,2,4-trimethyl-1-cyclohexyl xyl, 1,2,5-trimethyl-1-cyclohexyl, 1,2,6-trimethyl-1-cyclohexyl, 1,3,3-trimethyl-1-cyclohexyl, 1,3,4-trimethyl-1-cyclohexyl, 1,3,5-trimethyl-1-cyclohexyl, 1,3,6-trimethyl-1-cyclohexyl, 1,4,4-trimethyl-1-cyclohexyl, 2,2,3-trimethyl-1-cyclohexyl, 2,2,4-trimethyl-1-cyclohexyl, 2,2,5-trimethyl-1-cyclohexyl, 2,2,6-trimethyl-1-cyclohexyl, 2,3,3-trimethyl-1-cyclohexyl, 2,3,4-trimethyl-1-cyclohexyl, 2,3,5-trimethyl-1-cyclohexyl, 2,3,6-trimethyl-1-cyclohexyl, 2,4,4-trimethyl-1-cyclohexyl, 2,4,5-trimethyl-1-cyclohexyl, 2,4.6-trimethyl-1-cyclohexyl, 2,5,5-trimethyl-1-cyclohexyl, 2,5,6-trimethyl-1-cyclohexyl, 2,6,6-trimethyl-1-cyclohexyl cyclohexyl, 3,3,4-trimethyl-1-cyclohexyl, 3,3,5-trimethyl-1-cyclohexyl, 3,3,6-trimethyl-1-cyclohexyl, 3,4,4-trimethyl-1-cyclohexyl, 3,4,5-trimethyl-1-cyclohexyl, 3,4.6-trimethyl-1-cyclohexyl, 3,5,6-trimethyl-1-cyclohexyl, 1,2,3,3-tetramethyl-1-cyclopropyl, 2,2,3,3-tetramethyl-1-cyclopropyl, 1 ,2,2,3-tetramethyl-1-cyclopropyl, 1,2,2,3-tetramethyl-1-cyclobutyl, 1,2,3,3-tetramethyl-1-cyclobutyl, 2,2,3,3-tetramethyl-1-cyclobutyl, 2,3,3,4-tetramethyl-1-cyclobutyl, 1,2,2,3-tetramethyl-1-cyclopentyl, 1,2,2,4-tetramethyl-1-cyclopentyl, 1,2,2,5-tetramethyl-1-cyclopentyl, 1,2,3,3-tetramethyl-1 -cyclopentyl, 1,2,3,4-tetramethyl-1-cyclopentyl, 1,2,3,5-tetramethyl-1-cyclopentyl, 1,2,5,5-tetramethyl-1-cyclopentyl, 2,2,3,3-tetramethyl-1-cyclopentyl, 2,2,3,3-tetramethyl-1-cyclohexyl, 2,2,4,4-tetramethyl-1-cyclohexyl, 2,2,5,5-tetramethyl-1-cyclohexyl, 3,3,4,4-tetramethyl-1-cyclohexyl, 3,3.5.5-Tetramethyl-1-cyclohexyl, 1-ethyl-1-cyclopropyl, 1-ethyl-1-cyclobutyl, 1-ethyl-1-cyclopentyl, 1-ethyl-1-cyclohexyl, 1-ethyl-1-cycloheptyl, 2-ethyl-1-cyclopropyl, 2-ethyl-1-cyclobutyl, 2-ethyl-1-cyclopentyl, 2-ethyl-1-cyclohexyl, 2-ethyl-1-cycloheptyl, 3-ethyl-1-cyclobutyl, 3-ethyl-1-cyclopentyl, 3-ethyl-1-cyclohexyl, 3-ethyl-1-cycloheptyl, 4-ethyl-1-cyclohexyl and 4-ethyl-1-cycloheptyl.

[0025] The iron(III) complexes having formula (1) or (2) may be used in in vivo diagnostic methods, in particular as contrast agents for magnetic resonance imaging. In particular, magnetic resonance imaging contrast agents may contain the iron(III) complexes described above.

[0026] Ligands having the following formula (L1) or (L2) are required for the preparation of iron(III) complexes: [ka] During the ceremony, m=1 to 5; Y 1 and Y 2 are independently selected from the group consisting of O, NH and NR, where R represents C1-C15 alkyl or C3-C10 cycloalkyl; Y 3 is selected from the group consisting of O, NH and NR, where R represents C1-C15 alkyl or C3-C10 cycloalkyl; Y 4 are OH, OR, NH2, NHR and NR 5 R 6 wherein R, R 5 and R 6 represents C1-C15 alkyl or C3-C10 cycloalkyl, and R 5 and R 6 are selected independently of each other, Z represents a bridge according to one of the formulas (3) to (10), [ka] During the ceremony, n=0 to 4, o=0 to 4, * and *' are Y in formula (1). 1 and Y 2 Or Y in formula (2) 3 and Y 4 represents the site of attachment of bridge Z to R 1 , R 2 , R 3 and R 4 are each independently selected from the group consisting of H, C1-C15 alkyl, and C3-C10 cycloalkyl; However, Z is * -CH2CH2- *’ However, the ligand having the formula (L2) is not limited thereto. The two substituents at the asymmetric carbon atom marked with * in formula (L1) or (L2) are preferably in the trans position. When Z corresponds to formulas (4) to (10), Y in formula (L1) 1 and Y 2 Group or Y of formula (L2) 3 and Y 4 The groups are preferably in the trans position at the binding sites labeled * and *'.

[0027] More preferably, the ligand corresponds to formula (L1) 1 and Y 2 respectively represent NH. When the ligand corresponds to formula (L2), preferably Y 3 represents NH, and Y 4 represents OH, NH2 or NHR. Ligands having the formulae (L1-1), (L1-2) and (L2-2) are particularly preferred. [ka]

[0028] The above statements regarding the iron(III) complexes should be mentioned with regard to further preferred embodiments of the ligands having the formulae (L1) and (L2).

[0029] The preparation of all iron(III) complexes having formula (2) can be carried out according to the following general reaction scheme: [ka]

[0030] Y 3 , Y 4 and Z are as defined above for the iron(III) complexes with formula (2) or the ligands with formula (L-2). The synthesis starts from anhydrous CDTAMA, preferably anhydrous tCDTAMA, an approach known from the literature. See Gestin JF, Faivre-Chauvet A, Mease RC, et al., Nucl Med Biol, 1993; 20(6), p. 755-762. Furthermore, this lithotripsy literature also describes the reaction of anhydrous tCDTAMA with, for example, ethane-1,2-diamine (synonym: ethylenediamine) to give N-[methyl(2-aminoethyl)carbamido]-trans-1,2,diaminocyclohexane-N,N',N'-triacetic acid, where a large excess of the diamine in DMSO (dimethylsulfoxide) is added to the anhydride. An aqueous solution of an excess of Fe(III) salt is added to the ligand (L2) and the precipitated complex (2) is isolated. As already mentioned above, Z is a group having the formula (3)-(10).

[0031] The preparation of iron(III) complexes having formula (1) can be carried out in a similar manner according to the following general reaction scheme: [ka]

[0032] This synthesis also starts from the known anhydride CDTAMA, in particular tCDTAMA. However, here the anhydride is placed in solution and, with vigorous stirring, a coupling agent (e.g., a diamine) is added dropwise. Under these conditions, the diamine acts, for example, as a coupling agent for the two anhydrides. An excess of an aqueous solution of an Fe(III) salt is added to the ligand (L1) and the precipitated complex (1) is isolated. As above, Z is a group having the formula (3)-(10).

[0033] According to an alternative method for preparing the iron(III) complexes having formula (1) or (2), the iron loading of the ligand is carried out with iron(III) hydroxide, as described in patent application PCT / EP2019 / 064750. In this method, iron(III) hydroxide is first produced using iron(III) chloride and sodium hydroxide, which is then washed with water. The iron(III) hydroxide is then directly used to load the ligand with iron(III). Furthermore, according to a further alternative variant, iron loading may be performed by iron(III) chloride (Boehm-Sturm P. et al., “Low-Molecular-Weight Iron Chelates May Be an Alternative to Gadolinium-based Contrast Agents for T1-weighted Contrast-enhanced MR Imaging”, Radiology 2017;286(2):p.537-546), but this suffers from the disadvantage of high NaCl concentrations. According to a further variant, iron loading is carried out with iron(III) nitrate.

[0034] Exemplary embodiments Synthesis of iron(III) complex (2-1) [ka] 6.53 g (17.9 mmol) of trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid (CDTA) was added to a solution of 12.87 mL (136.2 mmol) of acetic anhydride and 2.75 mL of pyridine. After stirring at room temperature under argon for 24 h, the reaction mixture was filtered and washed with acetic anhydride followed by excess ethyl acetate. The solid was collected and dried under vacuum to give 5.30 g of a white powder (CDTA monoanhydride). CDTA monohydrate (CDTAMA) (14.4 mmol, 5 g) was added portionwise as a solid to a solution of 19.31 mL of ethylenediamine (289 mmol) and 23.75 mL of DMSO under argon over 6 hours. After the addition, the reaction mixture was stirred at room temperature overnight. The mixture was then concentrated using a rotary evaporator to a thick orange oil that solidified on standing. The residue was dissolved in methanol and a precipitate formed at room temperature. This was then washed with methanol. 3.98 g of 2,2'-({(1R,2R)-2-[{2-[(2-aminoethyl)amino]-2-oxoethyl}(carboxymethyl)amino]cyclohexyl}-azanediyl)diacetic acid (L2-1) was obtained as a white solid. 2 g of iron(III) nitrate nonahydrate was dissolved in 10 mL of water and filtered. A mixture of 5 mL of ammonium hydroxide solution (28-30% aqueous solution) and 5 mL of water was added dropwise to the iron(III) nitrate solution. The resulting insoluble iron(III) hydroxide was collected by filtration (Buchner funnel) and washed several times with water. 1.9 g of L2-1 was dissolved in 4 mL of water. Iron(III) hydroxide was added and the volume of the solution was made up to 10 mL with water. The solution was heated at 95° C. with stirring for about 2 hours. After this, the solution was cooled and filtered (0.2 μm) and then the volume was reduced to 2 mL by heating. The iron complex was precipitated by adding acetone (about 30 mL), centrifuged and dried. Iron(III) complex 2-1 was dissolved in water and the pH was adjusted to 7.3-7.4 with meglumine.

[0035] Synthesis of Ligand (L1-1) [ka] 6.42 g (17.6 mmol) of trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid (tCDTA) was added to a solution of 12.66 mL (134.0 mmol) of acetic anhydride and 2.71 mL of pyridine. After stirring at room temperature under argon for 24 h, the reaction mixture was filtered and washed with acetic anhydride followed by excess ethyl acetate. The solid was collected and dried under vacuum to give 5.16 g of a white powder. N-[methyl-(2-aminoethyl)carbamido]-trans-1,2-diaminocyclohexane-N,N',N'-triacetic acid CDTAMA (12.5 mmol, 4.33 g) was added in portions as a solid to a solution of 0.25 mL ethylenediamine (3.75 mmol), 36.99 mL DMSO, and 4.04 mL (50 mmol) pyridine under argon over 6 hours. After addition, the reaction mixture was stirred overnight at room temperature. After drying on a rotary evaporator, the residue was washed with copious amounts of ether and ethanol, and then purified by column chromatography on Sephadex G-10 gel. The product fractions were analyzed using HPLC, collected, and then lyophilized.

[0036] Comparative Example [Fe(III)tCDTA] - Synthesis of [ka] 20 g of iron(III) nitrate nonahydrate was dissolved in 100 mL of water and filtered. A mixture of 50 mL of ammonium hydroxide solution (28-30% aqueous solution) and 50 mL of water was added dropwise to the iron(III) nitrate solution. The resulting insoluble iron(III) hydroxide was collected by filtration (Buchner funnel) and washed several times with water. 17.1 g of tCDTA was dissolved in 40 mL of water. Iron(III) hydroxide was added and the volume of the solution was made up to 100 mL with water. The solution was heated at 95°C with stirring for approximately 2 hours. After this, the solution was cooled and filtered (0.2 μm) and then the volume was reduced to 20 mL by heating. The iron complex was precipitated by adding acetone (approximately 300 mL), followed by centrifugation and drying. Fe-tCDTA was dissolved in water and the pH was adjusted to 7.3-7.4 with meglumine (iron concentration 38 mg / mL; 0.68 M).

[0037] Synthesis of the ligand tCDTA-mono-trans-1,4-diaminocyclohexane (L2-2) [ka] 6.53 g (17.9 mmol) of trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid (CDTA) was added to a solution of 12.87 mL (136.2 mmol) of acetic anhydride and 2.75 mL of pyridine. After stirring at room temperature under argon for 24 h, the reaction mixture was filtered and washed with acetic anhydride followed by ethyl acetate. The solid was collected and dried under vacuum to yield 5.3 g of N-[methyl-(2-aminoethyl)carbamido]-trans-1,2-diaminocyclohexane-N,N',N'-triacetic acid (81.2%, CDTAMA) as a white powder. 2.4 g of N-[methyl-(2-aminoethyl)carbamido]-trans-1,2-diaminocyclohexane-N,N',N'-triacetic acid (6.93 mmol, CDTAMA) was added to a solution of 3.17 g of trans-1,4-diaminocyclohexane (27.7 mmol) in 18.24 mL of DMSO at 110° C. under argon for 6 h to dissolve. After the addition was complete, the mixture was stirred at room temperature overnight. The mixture was concentrated under reduced pressure, taken up in anhydrous methanol, and crystallized at room temperature for 2 days. The solid was washed with methanol. 1.89 g of L2-2 (79%) was obtained as a white solid.

[0038] Synthesis of the ligand trans-1,4-diaminocyclohexane-tCDTA dimer (L1-2) [ka] 6.42 g (17.6 mmol) of trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid (CDTA) was added to a solution of 12.66 mL (134.0 mmol) of acetic anhydride and 2.71 mL of pyridine. After stirring at room temperature under argon for 24 h, the reaction mixture was filtered and washed with acetic anhydride followed by ethyl acetate. The solid was dried under vacuum (CDTAMA; white powder). CDTAMA (17.9 mmol, 5.16 g) was added to a solution of trans-1,4-diaminocyclohexane (7.4 mmol, 0.85 g), 4.82 mL (59.6 mmol) of pyridine, and 58.80 mL of DMSO, and stirred overnight at room temperature under argon. The mixture was dried on a rotary evaporator. The white residue was dissolved in absolute ethanol, and the supernatant was then removed from the residue and dried again on a rotary evaporator. The resulting crystals gave one product peak by HPLC.

[0039] Determination of T1 and T2 relaxivity Figure 1 shows the pH dependence of the T1 and T2 relaxivities in water for the iron(III) complex having formula (2-1) (0.94 T NMR relaxometer, Bruker Minispec mq 40, Karlsruhe, Germany). It shows that both the T1 relaxivities (Figure 1A) and T2 relaxivities (Figure 1B) for the iron(III) complex having formula (2-1) follow a sigmoidal relaxivity increase with decreasing pH, with the change being maximum at pH 6–7. At pH < 6, the maximum T1 relaxivities are 1.7 L mmol / L. -1 ·s -1 , or T2 relaxivity 1.9 L·mmol -1 ·s -1 reached. Subsequent T1 relaxivity measurements used ultrapure water and fetal bovine serum (FBS, pH 7.7) as solvents for the metal complexes. Relaxivity measurements at different magnetic field strengths were performed with the following instruments: 0.94T: NMR relaxometer (Bruker Minispec mq 40 (Germany)), 1.5T: SIEMENS MAGNETOM Sonata MRI, 3T: SIEMENS MAGNETOM Lumina MRI, and 7T: Bruker Pharmascan Kleintier-MRI. All other parameters are listed in Table 1. [Table 1]

[0040] The results of the T1 relaxivity measurements (see Table 1) show that the exemplary embodiments according to the present invention (2-2) and (1-2) have significantly higher relaxivities compared to [Fe(III)tCDTA], especially at the particularly clinically relevant magnetic field strengths of 1.5T and 3T. In the case of the dimer (1-2), the relaxivities of the whole complex molecule, which may be related to biodistribution, are particularly high. With respect to use as a contrast agent for magnetic resonance imaging in in vivo diagnostics, this may lead to improved imaging. Surprisingly, the iron complex (1-2) exhibits hepatic excretion.

Claims

1. An iron(III) complex having the formula (1) or (2): 【Chemistry 1】 【change】 During the ceremony, m=1, Y 1 and Y 2 are selected independently from the group comprising O, NH and NR, where R represents C1-C15 alkyl or C3-C10 cycloalkyl; Y 3 is selected from the group comprising O, NH and NR, where R represents C1-C15 alkyl or C3-C10 cycloalkyl; Y 4 OH, OR, NH 2 , N.H.R. and N.R. 5 R 6 wherein R, R 5 and R 6 represents C1-C15 alkyl or C3-C10 cycloalkyl, and R 5 and R 6 are selected independently of each other, Z represents a bridge according to one of the formulas (3) to (10), 【Chemistry 2】 【change】 During the ceremony, n=0 to 4; o=0 to 4; * and *' are Y in formula (1). 1 and Y 2 Or Y in formula (2) 3 and Y 4 represents the site of attachment of bridge Z to R 1 , R 2 , R 3 and R 4 are each independently selected from the group consisting of H, C1-C15 alkyl and C3-C10 cycloalkyl; Iron(III) complexes.

2. 2. The iron(III) complex of claim 1, wherein the two substituents at the asymmetric carbon atom of formula (1) or (2) marked with * are in the trans position relative to each other.

3. Z has the formula (4) to (10), and at the binding sites of the bridges Z marked with * and *', Y of formula (1) 1 and Y 2 Group or Y of formula (2) 3 and Y 4 3. The iron(III) complex of claim 1, wherein the groups are in trans position relative to each other.

4. The iron(III) complex has the formula (1), 1 and Y 2 The iron(III) complex according to any one of claims 1 to 3, wherein represents NH.

5. The iron(III) complex has the formula (2), 3 represents NH, and Y 4 OH, NH 2 or NHR.

6. 2. The iron(III) complex of claim 1 having the formula (1-1), (1-2), (2-1) or (2-2). 【Chemistry 3】 【change】 【change】 【change】

7. Use of an iron(III) complex according to any one of claims 1 to 6 as a contrast agent for magnetic resonance imaging.

8. A contrast agent for magnetic resonance imaging, comprising the iron(III) complex according to any one of claims 1 to 6.

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