NO174048B - MACROCYCLIC COMPOUNDS AND DIAGNOSTIC AGENTS CONTAINING THESE - Google Patents

Abstract

Compounds of the general formula I <IMAGE> in which Y is a nitrogen or phosphorus atom, A<1> and A<2> are identical or different and are each a straight-chain or branched alkylene group having 2 to 6 carbon atoms, U<1>, U<2>, U<3>, U<4> are identical or different and are each a direct bond or a straight-chain or branched alkylene group having 1 to 6 carbon atoms, D<1>, D<2>, D<3>, D<4> are identical or different and are each an oxygen or sulphur atom, an alkylene group having 1 to 6 carbon atoms or a group N-R<7> where R<7> has the meaning of a hydrogen atom, a straight-chain or branched alkylene chain having 1 to 4 carbon atoms, which carries a COOR<1> group at the end, in which R<1> represents a hydrogen atom or a metal ion equivalent, D<5> has the meaning given for D<1>, D<2>, D<3> and D<4> and the group <IMAGE> where R<5> has the meaning of a hydrogen atom or of a straight-chain or branched, saturated or unsaturated C1-C20-alkylene group which may contain imino, phenyleneoxy, phenyleneimino, amide, ester groups, oxygen, sulphur and/or nitrogen atoms and may be substituted by hydroxyl, mercapto, imino and/or amino groups and has either a functional group or bound via this group a macromolecule B at the end, s and t are integers from 0 to 5, R<2> is hydrogen, a linear or branched, saturated or unsaturated, alkyl, acyl or acylalkyl group of 1 to 16 carbon atoms which is unsubstituted or substitute by one or more hydroxyl or lower alkoxy groups, or is -CH2-X-V where X has the meaning of carbonyl, of a straight-chain or branched alkylene group having 0 to 10 carbon atoms, which is unsubstituted or substituted by one or more hydroxyl or lower alkoxy groups, or of a straight &tilde& chain or branched alkylene group having 2 to 23 carbon atoms which is interrupted by oxygen atoms, V has the meaning <IMAGE> or -COOR<6> in which R<3> and R<4>, independently of one another, are hydrogen, a linear o branched alkyl group having 1 to 16 carbon atoms which is unsubstituted or substituted by one or more hydroxyl or lower alkoxy groups, or R<3> and R<4> together with the nitrogen atom represent a saturated 5- or 6-membered ring which may contain a further hetero atom and R<6> represents hydrogen or a saturated, unsaturated, straight-chain or branched or cyclic hydrocarbon radical having up to 16 carbon atoms or represents an aryl or aralkyl group, or R<2> or R<3> denotes a second macrocycle of the formula I' <IMAGE> which is bound via an alkylene chain (K) containing 2 to 20 carbon atoms, which, if desired carries carbonyl groups at the ends and, if desired, is interrupted by one or more oxygen atoms or R<1>-carboxymethylimino groups or substituted by one or more hydroxyl, lower alkoxy or carboxy-lower alkyl groups, which macrocycle can have a structure which is different from that of the parent structure, or R<2> denotes B or CH2-COB, with the proviso that if Rz represents B or CH2-COB, R<5> denotes a hydrogen atom, that at least two COORi groups are present in the molecule and that two hetero atoms of the macrocycle are each bound to at least two carbon atoms via an alkylene group and functional groups present in the molecule are, if desired conjugated with macromolecules and, if desired free carboxyl groups have been converted with organic or inorganic bases or amino acids and basic groups with inorganic or organic acids to the salts.

Description

The invention relates to the object characterized in the patent claims, i.e. macrocyclic complex formers, complexes and complex salts, agents containing these compounds and their use as diagnostics.

Already in the early 1950s, metal complexes were taken into account as contrast agents for radiology. However, the compounds used at the time were toxic in such a way that they were not considered for use in humans. It was therefore quite surprising that certain complex salts have proved to be sufficiently tolerable, so that they could be considered for routine use in humans for diagnostic purposes. As the first representative of this substance class, the dimeglumine salt of Gd DTPA (gadolinium-III complex of diethylene-triaminepentaacetic acid) which is described in the European patent application with publication number 71564, has so far in the clinical trial on over 1000 patients proven very useful as contrast agent for nuclear magnetic resonance imaging. The center of gravity for the application lies in diseases of the central nervous system.

An important reason for the good tolerability of Gd DTPA in the clinical application lies in the high activity in the nuclear spin tomography, especially in many brain tumours. Due to its good activity, Gd DTPA at 0.1 mmol/kg body weight can be dosed much, much lower than, for example, x-ray contrast agents in many x-ray examinations.

As a further representative of the komleksalten have

the meglumine salt of Gd DOTA (gadolinium-III complex of 1,4,7,10-tetraazacyclododecane-tetraacetic acid) described in the German patent application 34 01 052, proved useful for diagnostic purposes.

However, it is also desirable to be able to use higher dosed chelates. This is particularly the case for the detection of certain diseases outside the central nervous system by means of nuclear spin tomography (NMR diagnostics), but particularly when using the chelates as X-ray contrast agents.

Compared to iodinated X-ray contrast agents, chelates can offer a number of advantages: a) Radiation absorption in the high-energy area and thus reduction of the radiation exposure for the patients and improvement of the prerequisites for the energy subtraction method. b) Avoidance of the known, unpredictable, partly even life-threatening or fatal, so-called "allergenic" or

the cardiovascular side effects of the currently used iodinated X-ray contrast agents, which are known as "contrast agent reactions".

The prerequisite for this is:

high concentration of radiation-absorbing elements in the solution (X-ray) or strong influence on the NMR signals a pharmacokinetics that is suitable for diagnostics - very firm binding of the metal ions in separable complexes also under in-vivo conditions - good tolerability of the highly concentrated, high-dose complex solution low allergenic potential for all the components in. the contrast agent - high stability and shelf life for the chemical components of the contrast solution.

These requirements apply to varying degrees and in different ways, but in principle to all applications of the mentioned complexes in in-vivo diagnostics.

The substances according to the invention and the solutions produced from them meet the aforementioned requirements in a surprising way. They have a strong effect that is adaptable to the current principles of the diagnostic method (X-ray, NMR, ultrasound, nuclear diagnostics).

The substances according to the invention are used: 1. For NMR diagnostics in the form of their complexes with the ions of the transition metals with the serial numbers 21 to 29, 42 and 44. 2. For NMR and X-ray diagnostics in the form of their complexes with the ions of the lanthanide elements with the serial numbers 57 to 70. 3. The substances intended for use in NMR diagnostics as well as those intended for use in X-ray diagnostics are suitable for ultrasound diagnostics. 4. For the radiodiagnosis in the form of its complexes with the radioisotopes of the elements with the ordinal numbers 27, 29, 31, 32, 38, 39, 43, 49, 62, 64, 70 or 77.

Even without special precautions, their pharmacokinetics allow improving the diagnosis of many diseases. The complexes are for the most part excreted unchanged and quickly again, so that especially in the case of the use of relatively toxic metal ions as active principle on

despite the high dosage, no harmful effects can be observed that can be traced back to the metal.

The practical application of the new complexes and complex formers is also facilitated by their sufficient, often even very good, chemical stability.

A further significant advantage of the described complexes and complex formers is their extraordinary chemical versatility. Next to the central atom, the properties can be adapted to the requirements for tolerability, handleability, etc. by choosing many substituents and/or salt formers. Thus, a much-desired specificity of the compounds for structures in the organism, for certain bio-chemical substances, for metabolic processes, for conditions in tissues or body fluids can be achieved in diagnostics, especially when linked to biological substances or to substances that exhibit a reaction with biological systems . Such substances which are suitable for the connection can be low-molecular (e.g. glucose, amino acids, fatty acids, bile acids, porphyrins) or high-molecular (poly-saccharides, proteins, antibodies, etc.) or also constitute structures foreign to the body, which, however, are distributed on specific way in the body or reacts with the constituents in the body. The utilization of such principles becomes possible the faster the more sensitive the detection method for a diagnosticum is.

The macrocyclic compounds according to the invention, i.e. the 1,4,7,10-tetraazacyclododecane derivatives, are characterized by the general formula I:

where

each R<1>, independently of one another, is hydrogen or a metal ion equivalent;

R<2> is a linear or branched, saturated or unsaturated alkyl group of up to 16 carbon atoms, substituted with 1 to 5 hydroxy or C₁-C₁ ₂-alkoxy groups,

-CH2-X-V where X has the meaning of carbonyl, a linear or branched alkylene group with 1-10 carbon atoms which is optionally substituted with 1 to 5 hydroxy or C 1 -C 4 -alkylene groups or a linear or branched alkylene group

which are interrupted by oxygen atoms and which have 5-23 carbon atoms,

where V has the meaning

and R<4> independently of each other is hydrogen, a linear or branched alkyl group with 1-16 carbon atoms, which is optionally substituted with 1 to 5 hydroxy or C 1 -C 4 -alkyl groups, or R<3> or R 4 together with the nitrogen atom is a pyrrolidine, piperidine, morpholine or piperazine ring, or R2 or R<3> is an additional macroring of formula I'

which is bound via an alkylene chain containing from 2 to 20 carbon atoms, and which optionally has carbonyl groups in the end positions and is optionally interrupted by from 1 to 4 oxygen atoms or is substituted with from 1 to 5 hydroxy-, C^-C^- Alkoxy or carboxy C 1 -C 3 -alkylene groups, or

R2 denotes B or CH2-COB, where B denotes a macromolecule or biomolecule that accumulates to a particularly large extent in the organ or organ part being examined, or in the tumour,

and, if desired, salts formed with free carboxyl groups and organic or inorganic bases, or with amino acids and basic groups with inorganic or organic acids.

Compounds of the general formula I with R<1> in the sense of hydrogen are termed complex formers and with at least two of the substituents R<1> in the sense of a metal ion equivalent are termed metal complexes.

When not all acidic hydrogen atoms are substituted with the central ion, one, several or all remaining hydrogen atoms) can be replaced by cations of inorganic and/or organic bases or amino acids. Suitable inorganic cations are, for example, the lithium ion, the potassium ion, the calcium ion and especially the sodium ion. Suitable cations of organic bases are, among others, those of primary, secondary or tertiary amines, such as e.g. ethanolamine, diethanolamine, morpholine, glucamine, N,N-dimethylglucamine and especially N-methylglucamine. Suitable cations of amino acids are, for example, cations from lysine, arginine and ornithine.

The complex compounds can also be linked to macromolecules, which are known to be particularly enriched in the organ or organ part to be examined. Such macromolecules are, for example, hormones, dextrans, polysaccharides, polychelons, hydroxyethyl starch, polyethylene glycol, desferrioxamine, bleomycin, insulin, prostaglandin, steroid hormones, amino sugars, amino acids, peptides, such as polylysine, proteins (such as immunoglobulins and monoclonal antibodies ) or lipids (also in the form of liposomes). Conjugates with albumins, such as human serum albumin, antibodies, such as e.g. monoclonal antibodies specific for tumor-associated antigens or antimyosin. Instead of the biomolecules, suitable synthetic polymers such as e.g. polyethyleneimines are attached. The resulting diagnostic agents are, for example, suitable for use in tumor and infarction diagnostics. As monoclonal antibodies for the conjugation, those that are directed against mainly cell membrane antigens are particularly suitable. As such, e.g. monoclonal antibodies or their fragments (F(ab)2) are suitable for tumor preparation, such as e.g. are directed against the carcinoembryonic antigen (CEA), human chorionic gonadotropin (J3-hCG) or other tumor antigens, such as glycoproteins. Antimyosin, antiinsulin and antifibrin antibodies are also suitable.

For liver examinations or for tumor diagnostics, for example, conjugates or inclusion compounds with liposomes (which can for example be used as unilamellar or multilamellar phosphatidylcholine-cholesterol vesicles) are suitable.

The preparation of the macrocyclic compounds with the general formula I takes place by using, in a manner known per se, in compounds with the general formula I,

which instead of the COOR^ group contains the COOZ group with Z in the sense of a carboxyl protecting group, however without exhibiting a macromolecule B, or the group CH2-COB,

cleave off the protecting groups Z and the thus obtained acids (R^ in the general formula I stands for hydrogen) if desired

a) in a manner known per se is reacted with at least one metal oxide or metal salt of an element with the ordinal numbers

21-29, 31, 32, 38, 39, 42-44, 49, 57-70 or 77 and then, if desired, acidic hydrogen atoms present are transferred to physiologically tolerable salts with inorganic and/or organic bases or amino acids and basic groups present are transferred to physiologically tolerable salts with inorganic or organic acids or

b) in a manner known per se is reacted with at least one metal oxide or metal salt of an element with the ordinal numbers

21-29, 31, 32, 38, 39, 42-44, 49, 57-70 or 77 and then the thus obtained metal complexes are bound in a manner known per se via functional groups contained in the molecule or to R<2> respectively to the CO group contained in R<2> and, if desired, present acidic hydrogen atoms are transferred with inorganic and/or organic bases or amino acids and present basic groups with inorganic or organic acids to physiologically tolerable salts, or c) are attached to i and in a manner known per se a macromolecule via the functional groups contained in the molecule

or to R<2> respectively. to the CO group contained in R<2> and then in a manner known per se is reacted with at least one metal oxide or metal salt of an element with the ordinal numbers 21-29, 31, 32, 38, 39, 42-44, 49 , 57-70 or 77 and then, if desired, present acidic hydrogen atoms are transferred with inorganic and/or organic bases or amino acids and present basic groups with inorganic or organic acids to physiologically tolerable salts.

Examples of carboxyl protecting groups Z include lower alkyl, aryl and aralkyl groups, for example methyl, ethyl, propyl, butyl, phenyl, benzyl, diphenylmethyl, triphenylmethyl, bis(p-nitrophenyl)methyl as well as the trialkylsilyl group.

The removal of the protective groups Z takes place in a manner known per se, for example by hydrolysis, alkaline saponification of the ester, preferably with alkali in an aqueous-alcoholic solution at temperatures from 0 to 50°C or when it concerns e.g. tert-butyl esters using trifluoroacetic acid.

The production of the educts takes place by cyclization of two reactants. The thus obtained cyclic compounds are then reacted, possibly after removal of the protective groups, to introduce the substituent R<2>.

The circularization is carried out using methods that are known from the literature [e.g. Org. Synth. 58, 86 (1978), Macrocyclic Polyether Syntheses, Springer Verlag Berlin, Heidelberg, New York 1982, Coord. Chem. Fox. 3, 3 (1968), Ann. Chem. 1976, 916]: One of the two reactants has two leaving groups at the chain end, the other two nucleophiles which displace these leaving groups. As an example, mention should be made of the reaction of terminal, possibly heteroatom-containing dibromo, dimesyloxy, ditosyloxy or dialkoxycarbonyl alkylene compounds with terminal diazaalkylene compounds which optionally additionally contain heteroatoms in the alkylene chain, of which one of the two reactants is R<5>"- substituted.

Nitrogen atoms present are optionally protected, e.g. as tosylates, and are released before the subsequent alkylation reaction by means of methods known from the literature.

If diesters are used in the cyclization reaction, the diketo compounds thus obtained must be reduced by means of methods known to those skilled in the art, e.g. with diborane.

The subsequent alkylation takes place with halogen ketones, esters, acids or alkanes, which may be substituted with one or more hydroxy or lower alkoxy groups and optionally contain oxygen atom(s) in the chain. Furthermore, the alkyl residue can also contain a terminal amino group.

When the alkylation takes place with a dihalogenated alkane, compounds of the general formula I occur with two macrocyclic rings connected by a carbon bridge.

Other methods known from the literature for the synthesis of compounds with more than one ring are e.g. between an amine and a carbonyl compound (e.g. acid chloride, mixed anhydride, activated ester, aldehyde), two amine substituted rings and a dicarbonyl compound (e.g. oxalyl chloride, glutardialdehyde), two rings, each exhibiting a nucleophile group, and an alkylene compound having two escape groups, in the case of terminal acetyls an oxidative linkage (Cadiot, Chodkiewicz in Viehe "Acetylenes", 597-647, Marcel Dekker, New York, 1969).

The chain connecting the rings can then be modified by subsequent reactions (e.g. hydration).

In the alkylation with haloacetic acid, an intermediate with R<2> = CH2X-COOH is obtained, which is transferred to the mono-amide over the mixed anhydride with chloroformic acid ester or by means of dicyclohexylcarbodiimide and reaction with a primary or secondary amine of the general formula

Examples of suitable amines include: dimethylamine, diethylamine, di-n-propylamine, diisopropylamine, di-n-butylamine, diisobutylamine, di-sec.butylamine, N-methyl-n-propylamine, dioctylamine, dicyclohexylamine, N-ethylcyclohexylamine , diisopropenylamine, benzylamine, aniline, 4-methoxyaniline, 4-dimethylaminoaniline, 3,5-di-methoxyaniline, morpholine, pyrrolidine, piperidine, N-methyl-piperazine, N-ethyl-piperazine, N-(2-hydroxyethyl)-piperazine , N-(hydroxymethyl)-piperazine, piperazineacetic acid isopropyl-amide, N-(piperazino-methylcarbonyl)-morpholine, N-(piperazino-methylcarbonyl)-pyrrolidine, 2-(2-hydroxymethyl)piperidine, 4-(2-hydroxyethyl)- piperidine, 2-hydroxymethylpiperidine, 4-hydroxymethylpiperidine, 2-hydroxymethyl-pyrrolidine, 3-hydroxy-piperidine, 4-hydroxypiperidine, 3-hydroxy-pyrrolidine, 4-piperidone, 3-pyrroline, piperidine-3-carboxylic acid amide, piperidine-4- carboxylic acid amide, piperidine-3-carboxylic acid diethylamide, piperidine-4-carboxylic acid dimethylamide, 2,6-dimethylpiperidine, 2,6-dimethylmorpholine, N-acetyl-p iperazine, N-(2-hydroxy-propionyl)-piperazine, N-(3-hydroxypropionyl)-piperazine, N-(methoxyacetyl)-piperazine, 4-(N-acetyl,N-methylamino)-piperidine, piperidine-4- carboxylic acid-(3-oxa-pentamethylene)-amide, piperidine-3-carboxylic acid-(3-oxapentamethylene)-amide, N-(N',N'-dimethyl-carbamoyl)piperazine, pyrazoline, pyrazolidine, imidazoline, oxazolidine , thiazolidine, 2,3-dihydroxypropylamine, N-methyl-2,3-dihydroxypropylamine, 2-hydroxy-l-(hydroxymethyl)-ethylamine, N,N-bis-(2-hydroxy-ethyl)-amine, N-methyl -2/3,4,5,6-pentahydroxyhexylamine, 6-amino-2,2-dimethyl-1,3-dioxepin-5-ol, 2-hydroxyethylamine, 2-amino-1,3-propanediol, diethanolamine, ethanolamine . The polyhydroxyalkylamines can advantageously also be used in protected form for reaction, e.g. as O-acyl derivatives or as ketals. This applies in particular when these derivatives are more convenient and cheaper to produce than the polyhydroxyalkylamines themselves. A typical example is 2-amino-1-(2,2-dimethyl-1,3-dioxolan-4-yl)-ethanol, the acetonide of 1-amino-2,3,4-trihydroxybutane, prepared according to DE-OS 31 50,917.

The subsequent removal of the protecting groups is problem-free and can e.g. take place by treatment with an acidic ion exchanger in an aqueous-ethanolic solution.

The acylation takes place with a corresponding acyl derivative, especially with an acyl halide or anhydride. If anhydrides or halides of di- or poly-carboxylic acids are used, compounds of the general formula I are obtained, in which the two macrocyclic rings are linked amide-like via a carbon bridge.

Examples of the conversion of hydroxy or amino groups which are bound to aromatic or aliphatic residues are the reactions carried out in anhydrous, aprotic solvents such as tetrahydrofuran, dimethoxyethane or dimethylsulfoxide in the presence of an acid scavenger such as e.g. sodium hydroxide, sodium hydride or alkali or alkaline earth carbonates, such as sodium, magnesium, potassium-calcium carbonate, at temperatures between 0°C and the boiling point of the solvent in question, preferably however between 20 and 60°C, with a substrate of the general formula

IV

where W stands for a nucleofuge such as Cl, Br, J, CH3C5H4-SO3, L for an aliphatic, aromatic, arylaliphatic, branched, linear or cyclic hydrocarbon residue of up to 20 carbon atoms and Fu for the desired terminal functional group. As examples of compounds with the general formula IV should be mentioned

Conversions of carboxy groups can e.g. is carried out by means of the carbodiimide method (Fieser, Reagents for Organic Syntheses 10, 142) over a mixed anhydride [Org. Prep. Pro. Int. 7, 215 (1975)] or over an activated ester

(Adv. Org. Chem., Part B, 472).

The compounds of the general formula I with R-*- in the sense of a hydrogen atom constitute complex formers. They can be isolated and purified or, without isolation, transferred to metal complexes of the general formula I with at least two of the substituents R^ in the sense of a metal ion equivalent.

The production of the metal complexes according to the invention takes place in the manner described in DE-OS 34 01 052 and EP 71564, whereby the metal oxide or a metal salt (for example the nitrate, acetate, carbonate, chloride or sulphate) of the elements with ordinal numbers 21-29, 31, 32, 38, 39, 42-44, 49, 57-70 or 77 are dissolved or suspended in water and/or a lower alcohol (such as methanol, ethanol or isopropanol) and reacted with the solution or suspension of the equivalent amount of the complexing acid with the general formula I with R<1> in the sense of a hydrogen atom and then, if desired, acidic hydrogen atoms present in acid groups are substituted with cations of inorganic and/or organic bases or amino acids.

The neutralization thereby takes place with the help of inorganic bases (e.g. hydroxides, carbonates or bicarbonates) of e.g. sodium, potassium or lithium and/or organic bases such as primary, secondary and tertiary amines, such as e.g. ethanolamine, morpholine, glucamine, N-methyl- and N,N-dimethylglucamine, as well as basic amino acids, such as lysine, arginine and ornithine.

For the production of the neutral complex compounds, enough of the desired bases can be added to the acidic complex salts in aqueous solution or suspension, for example, so that the neutral point is reached. The solution obtained can then be evaporated to dryness in a vacuum. It is often an advantage to precipitate the formed neutral salts by adding solvents that are miscible with water, such as e.g. lower alcohols (methanol, ethanol, isopropanol and others), lower ketones (acetone and others), polar ethers (tetrahydrofuran, dioxane, 1,2-dimethoxyethane and others) and thus obtain crystals that are easy to isolate and easy to clean . It has been shown to be particularly advantageous to add the desired base to the reaction mixture already during complex formation and thereby save a process step.

If the acidic complex compounds contain several acidic groups, it is often appropriate to prepare neutral mixed salts, which contain inorganic as well as organic cations such as counterions.

This can take place, for example, by reacting the complex-forming acid in aqueous suspension or solution with the oxide or salt of the element that supplies the central ion and half of the required amount of an organic base needed for neutralization, the formed complex salt is isolated, purified if desired and then the necessary amount of inorganic base is added for complete neutralization. The addition of bases can also take place in the reverse order.

Basic groups can be transferred to pharmaceutically acceptable salts with inorganic and/or organic acids.

Examples of inorganic bases used are lithium, sodium and potassium hydroxide. As organic bases are, among others, primary, secondary and tertiary amines, such as e.g. ethanolamine, morpholine, glucamine, N-methyl- and N,N-dimethyl-glucamine, as well as basic amino acids, such as lysine, arginine and ornithine, suitable.

For salt formation with basic groups, inorganic acids, such as hydrochloric acid, and organic acids, such as citric acid, speaking.

The conjugate formation can e.g. take place via a carboxyl group in the complex compound or via the functional group which is located at the end of the C^-C20-a-alkylene group in the substituent R^, as defined further above. During the conjugate formation of the acids with macromolecules, several acid residues can be bound to this. In this case, several central ions can be attached to a macromolecule.

The connection to the desired macromolecules likewise takes place using methods known per se, such as those e.g. described in Rev. room. Morphol. Embryol. Physiol., Physiologie 1981, 18, 241 and J. Pharm. Pollock. 68, 79 (1979), for example by reaction between the nucleophilic group in a macromolecule, which e.g. the amino, phenol, sulfhydryl, aldehyde or imidazole group and an activated derivative of the complexing agent. Examples of activated derivatives include monoanhydrides, acid chlorides, acid hydrazides, mixed anhydrides (see, for example, G.E. Krejcarek and K.L. Tucker, Biochem. Biophys. Res. Res. Commun. 1977, 581), activated esters, nitrenes or isothiocyanates. Conversely, it is also possible to react an activated macromolecule with the complexing acid. For conjugation with proteins, substituents with e.g. the structure CgH4N2, C6H4NHCOCH2, C6H4NHCS or C6H4OCH2CO.

The conjugation of the complex-forming acid with dextrans and dextrins likewise takes place using methods known per se, for example by activating the polysaccharides with cyanogen bromide and subsequent reaction with amino groups in the complex-forming acid.

In the case of the use of complex compounds containing radioisotopes, their preparation can be carried out by means of the methods described in "Radiotracers for Medical Applications", Volume 1, CRC-Press, Boca Raton, Florida.

The agents according to the invention fulfill all requirements for being suitable as a contrast agent for nuclear spin tomography. Thus, they are eminently suitable for that, after oral or parenteral application by increasing the signal intensity and improving the expressive power of the image obtained by means of the nuclear spin tomograph. Furthermore, they exhibit the high efficiency necessary to burden the body with the smallest possible amounts of foreign substances and the good compatibility necessary to maintain the non-invasive nature of the examinations.

The good water solubility of the agents according to the invention makes it possible to prepare highly concentrated solutions, so that the volume load for the circuit can be kept within reasonable limits and to compensate for the dilution through the body fluid. Furthermore, the agents according to the invention not only exhibit a high stability in vitro, but also a surprisingly high stability in vivo, so that only an extremely slow release or exchange of the inherently toxic ions that are not covalently bound in the complexes in during the time in which the new

the contrast agents are again completely excreted.

Usually, the agents according to the invention for use as NMR diagnostics are dosed in amounts of 0.001-5 mmol/kg, preferably 0.005-0.5 mmol/kg. Details of the application are discussed, for example, in H.J. Weinmann et al., Am. J. of Roentgenology 142, 619 (1984).

Particularly low doses (below 1 mg/kg) of organ-specific NMR diagnostics are e.g. applicable for the detection of tumors and of heart attacks. Furthermore, the complex compounds according to the invention can advantageously be used as shift reagents.

The agents according to the invention are, due to their favorable radioactive properties and the good stability of the complex compounds contained in them, also suitable as radiodiagnostics. Details for application and dosage are described e.g. in "Radiotracers for Medical Applications", CRC-Press, Boca Raton, Florida.

A further imaging method with radioisotopes

is the positron emission tomography, which uses positron-emitting isotopes such as ^Sc, ^^ sc, ^<2>Fe, 55Co Qg <68>Ga. (Heiss, W.D., Phelps, M.E., Position Emission Tomography of Brain, Springer Verlag Berlin, Heidelberg, New York 1983.)

In the following table, the closest compound to the state of the art is Gd-DOTA (formula I, where R<2> = CH2COOH, 3 x R<1> = Gd3+; counterclaims W 086/02352; EP 124766;

FR 2539996; US-4647447; ABOUT. Vol. 97 (1982), 206960Z) compared to representative compounds according to the invention.

It is clear that the loss of an acetic acid group at a nitrogen atom of the macrocycle surprisingly does not lead to a deterioration, but to a clear improvement of the properties of the new complexes. Thus, the T^relaxivity value which is a measure of the signal amplification of the contrast agent used, for example in the case of the compound according to example 5, is clearly higher than the corresponding value for the nearest prior art compound (DOTA).

With regard to side effects such as pain, damage to blood vessels and cardiovascular disturbances due to osmotic pressure, these are greatly reduced for the examined new complexes in comparison with the blood-clearly hypertonic Gd-DOTA.

The agents according to the invention are eminently suitable as X-ray contrast agents, whereby it must be particularly emphasized that with these no signs of the anaphylaxis-like reactions can be recognized in biochemical-pharmacological examinations which are known from the iodine-containing contrast agents. They are particularly valuable because of the favorable absorption properties in areas of higher tube voltages for digital subtraction techniques.

In general, the agents according to the invention are dosed for use as X-ray contrast agents analogously to e.g. meglumine diatrizoate in amounts of 0.1-5 mmol/kg, preferably 0.25-1 mmol/kg.

Details of the use of X-ray contrast agents are discussed e.g. in Barke, Rontgenkontrastmittel, G. Thieme, Leipzig (1970) and P. Thurn, E. Biicheler, "Einfuhrung in die Rontgendiagnostik", G. Thieme, Stuttgart, New York (1977).

The agents according to the invention are, as their acoustic impedance is higher than this impedance of body fluids and tissues, also suitable as a contrast agent for ultrasound diagnostics, particularly in the form of suspensions. They are usually dosed in amounts of 0.1-5 mmol/kg, preferably 0.25-1 mmol/kg.

Details of the application of ultrasound diagnostics are described, e.g. in T.B. Tyler et al., Ultrasonic Imaging 3, 323 (1981), J.I. Haft, "Clinical Echocardiography", Futura, Mount Kisco, New York 1978 and G. Stefan "Echokardiographie", G. Thieme Stuttgart/New York 1981.

In all, it has been successful in synthesizing new complex formers, metal contplexes and metal complex salts, which open up new possibilities in diagnostic and therapeutic medicine. Above all, the development of new imaging procedures in medical diagnostics is highly desirable.

The following examples serve to further explain the object of the invention.

In the following, the synthesis of starting material is described in the form of examples, starting from a ring closure reaction: a) l-benzyl-4,7,10-tris-(p-tolylsulfonyl)-1,4,7,10-tetraazacyclododecane

To a solution of 164.6 g of N,N',N"-tris-(p-tolylsulfon-yl)-diethylenetriamine-N,N"-disodium salt in 2.16 1 of dimethylformamide is added dropwise at 100°C with stirring 145 g of N ,N-bis-[2,2'-(p-tolylsulfonyloxy)]-ethane-benzylamine dissolved in 900 ml of dimethylformamide during 3 hours. Then, while stirring at 80°C, 1 1 water is added dropwise, and it is stirred for a further 18 hours at room temperature, then cooled to 0°C, and the precipitate is suctioned off, washed with a little ice-cold ethanol and dried at 15 torr and 60°C. 175 g of the title compound are obtained.

Alternative procedure 1

An analogous method for the production of tetraazacyclododecane derivatives can be found in M. Hediger and T.A. Kaden, Heiv. Chim. Acta 66, 861 (1983).

30.94 g of N,N',N"-tris-(p-tolylsulfonyl)-diethylenetri-amine-N,N"-disodium salt and 28.12 g of N-bis-(2-methane-sulfon-yloxy-ethyl) -triphenylmethylamine is stirred with 530 ml of dimethylformamide for 20 hours at 80-85°C, then cooled and stirred into a solution of 30 g of potassium carbonate in 5 1 of ice water. The precipitate is suctioned off, the filter cake is washed with 0.5 1 and dried at 20°C in a vacuum at 150 torr. For purification, dissolve in 230 ml of chloroform and 5 ml of triethylamine, filter, evaporate in vacuo to 200 ml, and add 250 ml of ethyl acetate to the solution at boiling temperature. The mixture is allowed to stand overnight to cool, and the precipitated crystals are suctioned off. 22.18 g of 1,4,7-tris-(p-tolylsulfonyl)-10-trisphenylmethyl-1,4,7,10-tetraazacyclododecane are obtained, mp: 185-188°C (decomposition).

To remove the trityl protecting group, 31.4 g of the tritosyl-trityl derivative prepared in this way is stirred in a mixture of 100 ml of glacial acetic acid, 75 ml of water and 300 ml of dioxane for one hour at 80°C. Then evaporate in vacuum at 60°C to a large extent, dilute with 300 ml of ice water and add 40 ml of 11 N caustic soda (pH above 12). This mixture is shaken with 300 ml of chloroform, the phases are separated, the water phase is extracted twice with 100 ml of chloroform each time, and the combined chloroform phases are dried over sodium sulphate and evaporated in vacuo. The foamy residue is treated with 300 ml of diethyl ether, whereby crystallization occurs. It is filtered off, the crystals are dried in a vacuum at 60°C and 150 torr, and 21 g of 1,4,7-tris-(p-tolylsulfonyl)-1,4,7,10-tetraazacyclododecane are obtained, m.p.: 202 -203°C.

The thus obtained tritosyl compound (21 g) is dissolved in 200 ml of dimethylformamide, 13.71 g of anhydrous potassium carbonate, 4.95 g of sodium iodide and 7.92 g of benzyl bromide are successively added to the solution, and it is stirred for 5 hours at 100°C. It is then cooled to 20°C, the mixture is stirred into 4 1 of ice water, filtered off with suction, and the residue is dissolved in 2 1 of dichloromethane. The solution is extracted with 100 ml of water, dried over sodium sulphate and evaporated in vacuo. The residue is dissolved at boiling temperature in 500 ml of acetonitrile and crystallizes overnight, filtered off, and the crystals are dried at 50°C and 150 torr. 16.20 g of 1-benzyl-4,7,10-tris-(p-tolylsulfonyl)-1,4,7,10-tetraazacyclododecane are obtained, m.p.: 217-219°C.

b) N-benzyl-1,4,7,10-tetraazacyclododecane

150 g of 1-benzyl-4,7,10-tris-(p-tolylsulfonyl)-1,4,7,10-tetraazacyclododecane are heated with 900 ml of HBr/acetic acid (40% strength) and 125 g of phenol for 16 hours at 50°C. After cooling to 20°C, dilute with 1 1 ether, cool to -s-5°C, and the precipitated crystals are suctioned off. To isolate the free base, the product is dissolved in 500 ml of 4 N caustic soda, saturated with potassium carbonate and extracted several times with chloroform, dried over magnesium sulfate and evaporated in vacuo. 39 g of the title compound are obtained as a light yellow, viscous oil.

A sample is characterized as tri-hydrochloride: m.p. 210°C

(under cleavage).

Alternative procedure 2

To a solution of 11.2 g of 1,4,7,10-tetraazacyclododecane in 900 ml of tetrahydrofuran at -s-20°C, 58 ml of triethylamine is added and, with stirring, a solution of 16.2 ml of benzoyl chloride in 280 ml of tetrahydrofuran, whereby the room temperature rises slightly above -5-10°C. It is then stirred for 16 hours at 0-10°C, the precipitate is filtered off, and the solution is evaporated in vacuo. The residue is chromatographed on 1 kg of silica gel and eluted with dioxane-water-ammonia solution (8:1:1). The uniform fractions after DSC are combined, evaporated, dissolved in dichloromethane and filtered to remove a small haze, and the solution is evaporated. 19.60 g of 1,4,7-tribenzoyl-1,4,7,10-tetraazacyclododecane are obtained, m.p.: 120-125°C.

11.5 g of the thus obtained tribenzoate is stirred in 150 ml of dimethylformamide with 8.3 g of anhydrous potassium carbonate, 3.0 g of sodium iodide and 7.2 ml of benzyl bromide for 18 hours at 100°C. It is filtered, the solution is evaporated in vacuo, the residue is stirred twice with 50 ml of hexane each time and decanted. The hexanes are discarded. For purification, dissolve in dichloromethane and chromatograph with dichloromethane-methanol (37:3) over 0.5 g of silica gel. 10.2 g of 1,4,7-tribenzoyl-10-benzyl-1,4,7,10-tetraazacyclododecane are obtained, m.p.: 105-109°C.

To remove the benzoyl group, this product (2.87 g) is dissolved in 290 ml of tetrahydrofuran, 11.2 g of potassium t-butylate is added and heated for 48 hours under reflux. It is filtered, evaporated in vacuo, the residue is added under ice-cooling to 100 ml of water and extracted three times with 50 ml of dichloromethane each time. The combined dichloromethane phases are shaken with 10 ml of water, dried over sodium sulphate and evaporated in vacuo. The initially oily residue crystallizes very slowly and is triturated with 20 ml of hexane. After suction and drying, 1.15 g of N-benzyl-1,4,7,10-tetraazacyclododecane are obtained, m.p. 75-78°C.

c) 1-benzyl-4,7,10-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane

To a solution of 131.8 g of N-benzyl-1,4,7,10-tetraazacyclododecane in 1.5 1 of dichloromethane, 200 g of triethylamine and 260 g of bromoacetic acid ethyl ester are successively added at 0°C over the course of 2 hours. It is stirred for a further 16 hours at room temperature, shaken with a 5% sodium carbonate solution and salt water, dried over magnesium sulphate and evaporated in vacuo. The residue is dissolved in 200 ml of chloroform and filtered over 2 kg of silica gel, whereby it is eluted with 1 1 chloroform-methanol (95:5). 210 g of the title compound are obtained as a viscous oil.

d) N,N',N"-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane

Dissolve 100 g of 1-benzyl-4,7,10-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane in 0.5 1 acetic acid and 0.5 1 ethyl acetate, add 5 g of palladium on charcoal (10% ) and shaken for 5 hours under hydrogen. The catalyst is filtered off, the solution is evaporated in vacuo, the residue is dissolved in chloroform, shaken with 100 ml of saturated soda solution and 100 ml of salt water, dried and evaporated in vacuum. The residue is purified by ball-tube distillation at 10~<3> torr and 120°C. N,N',N"-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane is obtained as a viscous, pale yellow oil.

Yield: 65 g.

IR (film): 3400, 2935, 2878, 1738 /cm.

Example l

Gadolinium- III- complex of N-[ 2, 3- dihydroxyv- N- methyl- propyl-carbamoylmethyl3- 1, 4, 7, 10- tetraazacyclododecane- N', N'', N' 1' - triacetic acid

Dissolve 55 g of N-[2,3-dihydroxy-N-methyl-propylcarbamoylmethyl]-N',N<1>',N<1>''-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane in 0.5 1 ethanol, add 96 ml of 3 N sodium hydroxide solution and stir for 3 hours at 20°C, then evaporate in vacuo, add 300 ml of water and adjust to pH 6 with 2 N hydrochloric acid. To this solution, 31.94 g of gadolinium acetate is added and stirred for 18 hours at 50°C, then applied to a

anion exchanger Araber1it# IRA 410, and then the aqueous eluate is applied to a cation exchanger Araberlite IRC 50. The eluate is evaporated in vacuo and dried. 47.14 g (73% of theory) of the title compound are obtained as a colorless powder.

The starting material for the preparation of the title compound according to example 2 is obtained in the following way: a) N,N',N"-tris-(ethoxycarbonylraethyl)-1,4,7,10-tetraazacyclododecane-N'''-acetic acid 20 g N,N ',N"-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane is dissolved in 300 ml of dichloromethane, 10.40 g of triethylamine is added and then a solution of 4.40 g of chloroacetic acid in 100 ml is added dropwise at 0°C dichloromethane and stirred for 20 hours at room temperature. For processing, it is distributed between dichloromethane and phosphate buffer pH 6, dried over magnesium sulfate and evaporated in a vacuum. 23 g of the desired compound is obtained as a viscous oil.

b) N-[ 2, 3- dihydroxy- N- methyl- propylcarbamoylmethyl]-N', N'', N'''- tris-( ethoxycarbonvlraethyl)- 1, 4, 7, 10- tetraazacyclododecane

To a solution of 48.86 g (100 ramol) N,N',N"-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane-N'''-acetic acid in 500 ml dichloromethane is added dropwise at 0° C 10.5 g of triethylamine and then a solution of 14 g of chloroformate isobutyl ester. It is stirred for one hour at 0° C. and then a solution of 10.52 g of N-methylamino-2,3-propanediol in 100 ml of chloroform is added dropwise and stirred in 2 hours at room temperature.Then shake with sodium bicarbonate solution and salt water,

dried over magnesium sulfate and evaporated in vacuo. For purification, the chloroform solution of the residue is filtered through 500 g of silica gel, and 55 g of the title compound is obtained.

Example 2

Gadolinium- III- complex of N-[ N- ethylcarbamoylmethyl3-1, 4, 7, 10- tetraazacyclododecane- N', N'', N'''- triacetic acid

Dissolve 24 g of N-[N-ethylcarbamoylmethyl3-N',N'',N''-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane in 250 ml of ethanol, add 47 ml of 3 N caustic soda and stirred for 3 hours at 20°C, evaporated in vacuo, 150 ml of water added and adjusted to pH 6 with 2 N hydrochloric acid. 15.55 g of gadolinium acetate are added and stirred for 5 hours at 60°C. The solution is then purified as described in the preceding examples using ion exchangers. 20.39 g of the title compound are obtained as a colorless powder.

Preparation of the starting material:

Dissolve 25 g (51.2 mmol) of N,N',N"-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane-N'''-acetic acid (see example la) in 200 ml of dichloromethane, add 5.40 g of triethylamine at 0° C. and then 7 g of chloroformate isobutyl ester. It is stirred for one hour at 0° C. and then a solution of 2.31 g of ethylamine in 20 ml of dichloromethane is added dropwise, stirred for 2 hours at room temperature, shaken with saturated sodium bicarbonate solution and brine, dried over magnesium sulfate and evaporated to dryness in vacuo. For purification, the dichloromethane solution of the residue is filtered over 200 g of silica gel, and 24 g of N-[N-ethylcarbamoylmethyl3-N',N'',N' is obtained ''-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane.

Example 3

Gadolinium- III- complex of N-(2, 3- dihydroxy-l- propyl)-1, 4, 7, 10- tetraazacvklododecane- N', N'', N'''- triacetic acid

Dissolve 4.30 g of N,N',N''-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane in 100 ml of dichloromethane, add 4.20 g of triethylamine and 2.21 g of 3-chloro-l ,2-propanediol. It is stirred for 16 hours at room temperature, shaken with water and evaporated in vacuo. The residue is stirred for 6 hours with 100 ml of 1 N

caustic soda, adjusted to pH 6 with 2 N hydrochloric acid and stirred for 16 hours with 3.34 g of gadolinium acetate at 50°C. The solution thus obtained is subjected to an ion exchanger purification, and 3.62 g of the title compound is obtained as a colorless powder.

Example 4

Gadolinium- III- complex of N-(2- aminoethyl)- 1, 4, 7, 10- tetraazacyclododecane- N', N'', N'''- triacetic acid

2.10 g of triethylamine and 1.22 g of N -(2-chloro-ethyl)-acetamide. It is stirred for 18 hours at room temperature, shaken with water and evaporated in vacuo. The residue is stirred for 8 hours with 50 ml of 1 N caustic soda at 60°C, adjusted to pH 6 with 2 N hydrochloric acid and stirred for 16 hours with 3.34 g of gadolinium acetate at 50°C. The solution thus obtained is subjected to an ion exchanger purification, and 3.21 g of the title compound is obtained as a colorless powder.

Example 5

Bis- gadolinium- III- complex of 1, 1'-( 1, 3- propylene)- bis-( 1, 4, 7, 10- tetraazacyclododecane- 4, 7, 10- tri-triacetic acid)

A solution of 8.60 g of N,N',N''-tris-(ethoxycarbonyl-methyl)-1,4,7,10-tetraazacyclododecane and 4.2 g of triethylamine in 200 ml of dichloromethane is added to 2.01 g of 1, 3-dibromopropane and stirred for 20 hours at room temperature, shaken with water and salt water and evaporated in vacuo. The residue is filtered with dichloromethane over 150 g of silica gel and evaporated. A tough oil is obtained which is stirred in 60 ml of 1 N caustic soda for 16 hours, then diluted with 100 ml of water and adjusted to pH 6 with 2 N hydrochloric acid. After adding 6.68 g of gadolinium acetate, the mixture is stirred for 16 hours at 50°C, and the solution is purified over an anion and cation exchanger. 6.56 g of the title compound is obtained as a colorless powder.

Example 6

Bis- gadolinium III- complex of succinyl- bis-(1, 4, 7, 10- tetraazacyclododecane- 4, 7, 10- triacetic acid)

A solution of 4.30 g of N,N',N''-tris-(ethoxycarbonyl-methyl)-1,4,7,10-tetraazacyclododecane and 4.20 g of triethylamine in 100 ml of dichloromethane is added dropwise at 0°C 1.705 g of succinic acid dichloride, dissolved in 20 ml of dichloromethane, and then stirred for one hour at room temperature. It is shaken out with sodium bicarbonate solution and salt water and evaporated in a vacuum. The residue is chromatographed on 100 g of silica gel with dichloromethane/ethyl acetate (0-30%).

A tough oil is obtained which is stirred with 40 ml of 1 N caustic soda for 4 hours, then diluted with water (100 ml) and adjusted to pH 6 with 2 N hydrochloric acid. After adding 3.34 g of gadolinium acetate, the mixture is stirred for 16 hours at 50°C, and the solution is purified over ion exchangers. 3.90 g of the title compound are obtained as white powder.

Example 7

N-( 2- hvdroxvethyl)- 1. 4. 7, 10- tetraazacvklododecane-N', N'', N'''- triacetic acid

A solution of 6.46 g of N,N',N''-tris-(ethoxycarbonyl-methyl)-1,4,7,10-tetraazacyclododecane in 150 ml of dichloromethane is added to 3.2 g of triethylamine and 1.45 g of 2- chloroethanol. It is stirred for 4 hours at room temperature, then shaken with sodium bicarbonate solution and salt water, dried over magnesium sulphate and evaporated in vacuo. The residue is stirred for 16 hours with 60 ml of 1 N caustic soda. By adding 5 N hydrochloric acid, the pH is adjusted to 2.5. The obtained suspension is applied to an ion exchanger (DOWEX® 50W-X4 in the H+<->form), eluted with water and

then with 0.5 M NH3 solution. It is evaporated in vacuo, and the title compound is isolated by adding ethanol and suctioning off the precipitate. 4.24 g of the title compound are obtained, the purity of which is verified by pH titration and elemental analysis.

Example 8

N-[ N-(2- hydroxyethyl)- carbamoylmethyl3- 1, 4, 7, 10- tetraazacyclododecane- N', N'', N'' 1- triacetic acid

A solution of 12.50 g of N,N',N''-tris-(ethoxycarbonyl-methyl)-1,4,7,10-tetraazacyclododecane-N'''-acetic acid in 250 ml of dichloromethane is added at 0°C 5 .22 g of triethylamine and then 3.50 g of chloroformate isobutyl ester. After one hour, 1.60 g of ethanolamine, dissolved in 50 ml of dichloromethane, are added dropwise, stirred for 2 hours at room temperature, shaken with soda solution and salt water, dried over magnesium sulfate and evaporated in vacuo. The residue is chromatographed with chloroform/acetone (10:1) on 200 g of silica gel, and 11 g of N-(2-hydroxyethyl)-carbamoylmethyl-N',N'',N'''-tris-(ethoxycarbonylmethyl)-1 are obtained ,4,7,10-tetraazacyclododecane.

This product is stirred with 100 ml of 1 N caustic soda

5 hours at room temperature, then acidify with dilute hydrochloric acid

to pH 2.5, and the suspension is purified on a cation exchanger (DOWEX<®>50W-X4), eluting with water and finally with 0.5 M NH 3 solution. The eluate is evaporated extensively, and after addition of ethanol the title compound crystallizes out, which is isolated by filtration. 7.2 g of the title compound are obtained, the purity of which is checked by titration and elemental analysis.

Example 9

Gadolinium complex of N-(morpholinocarbonylmethyl)- 1, 4, 7, 10-tetraazacyclododecane- N', N'', N'''- triacetic acid

55.77 g of N-(morpholinocarbonylmethyl)-N',N'',N'''-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane are hydrolyzed analogously to example 2 with caustic soda and complexed with gadolinium acetate. 45.19 g (72% of theory) of the title compound is obtained as a white powder.

The starting material for the preparation of the title compound is obtained by working analogously to example 2b, but morpholine is used instead of N-methyl-amino-2,3-propane-diol.

Example 10

Gadolinium complex of N-(morpholinocarbonylmethyl)- 1, 4, 7, 10-tetraazacyclododecane- N', N' 1, N'''- tris-( 2- methyl- acetic acid)

53.06 g of N-(morpholinocarbonylmethyl)-N',N'',N'''-tris-(1-ethoxycarbonyl-1-ethyl)-1,4,7,10-tetraazacyclododecane is hydrolyzed with caustic soda analogously to example 2 and complexed with gadolinium acetate. 52.20 g (78% of theory) of the title compound are obtained as a white powder.

Example 11

Bis- gadolinium complex of l, l'-(2-hydroxyv- l, 3- propylene)- bis-1, 4, 7, 10- tetraazacvklododecane- 4. 7, 10- triedacetic acid

4.63 g of epichlorohydride is added to a solution of 43.05 g of N,N',N''-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane in 450 ml of dimethylformamide. After one hour, 7.5 g of sodium iodide are added, and it is heated for 24 hours at 80°C. It is evaporated in vacuo, the residue is distributed between water and chloroform, the chloroform phase is dried over sodium sulphate and evaporated in vacuo. The residue is chromatographed on 1 kg of silica gel with dichloromethane-10% acetone, and 27.5 g of 1,1'-(2-hydroxy-1,3-propylene)-bis-1,4,7,10-tetraazacyclododecane triacetic acid ethyl ester is obtained ) as viscous oil.

9.17 g of the thus produced ester are dissolved in

200 ml of ethanol and stirred with 30 ml of 3 N caustic soda for 20 hours at room temperature, then adjusted to pH 6 with hydrochloric acid and stirred for 16 hours at 60°C with 6.68 g of gadolinium acetate, and the solution is purified over an anion and cation exchanger, and 10.05 g of the title compound are obtained as white powder.

Example 12

N- methylglucamine salt of the manganese II- complex of N-[ N-(2-hydroxyethyl)- carbamoylmethyl]- 1, 4, 7, 10- tetraazacyclododecane- N', N'', N'''- triacetic acid

8.95 g (20 mmol) of N-[N-(2-hydroxyethyl)-carbamoyl-methyl]-1,4,7,10-tetraazacyclododecane-N',N'',N'''-triacetic acid are suspended in 30 ml of water and heated with 1.40 g (20 mmol) manganese II oxide for 3 hours at 100°C. 3.90 g (20 mmol) of N-methylglucamine are then added, heated for a further 12 hours at 100°C, and the solution evaporated in vacuo to dryness. 13.8 g of the title compound are obtained as a pink powder, m.p. 140-143°C.

Example 13

Dysprosium- III- complex of N-(morpholinocarbonylmethyl)-1, 4, 7, 10- tetraazacyclododecane- N', N'', N'''- triacetic acid

20 g of N-(morpholinocarbonylmethyl)-N',N'',N'''-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane are hydrolyzed with caustic soda analogously to example 2 and complexed with dysprosium acetate. 16.3 g of the title compound are obtained as white powder.

Example 14

Preparation of liposomes loaded with gadolinium-N-(morpholinocarbonylmethyl)-1,4,7,10-tetraazacyclododecane-N',N'',N'''-triacetic acid

According to the procedure described in Proe. Nati. Acad. Pollock. USA, 75, 4194, a lipid mixture of 75 mol% egg phosphatidylcholine and 25 mol% cholesterol as dry substance is prepared. 500 mg of this is dissolved in 30 ml of diethyl ether and 3 ml of an aqueous 0.1 Is solution of the gadolinium complex of N-(morpholinocarbonyl-methyl)-1,4,7,10-tetraaza-N',N is added dropwise in an ultrasonic bath '',N'''-triacetic acid. The ultra-sonic treatment is continued for a further 10 min., and it is evaporated in vacuo. The gel-like residue is suspended in 0.125 M sodium chloride solution and centrifuged repeatedly at 0°C and 20,000 g to separate unencapsulated gadolinium complex. The suspension is then subjected to freeze-drying in a multi-purpose glass. The application takes place as a colloidal dispersion in a 0.9% sodium chloride solution.

Example 15

Preparation of a solution of the yttrium-90 complex of the conjugate of 1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid with monoclonal antibody

For a suspension of 4 mg of 1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid in 1 ml of water, 2 mg of N-(3-dimethylamino-propyl)-N'-ethyl-carbodiimide hydrochloride is added and then 1 ml of a solution of 0.6 mg of monoclonal antibody (with specificity against melanoma antigen) dissolved in 0.05 molar sodium hydrogen carbonate buffer (pH 7.8). It is stirred for 2 hours at room temperature and dialyzed against a 0.3 M sodium phosphate buffer. Then 1 ml of a yttrium-90 solution in acetate buffer pH 6 [prepared according to Int. J. Appl. Radiate. Isot., vol. 36

(1985), p. 803] and incubated for 24 hours at room temperature.

The solution is applied to a Sephadex5 G 25 column, and the radioactive protein fraction is sterilized and filled onto multi-disinfector tubes. A storable dry preparation is obtained by lyophilization.

Example 16

The indium III complex of N-(morpholinocarbonylmethyl)-1, 4, 7, 10- tetraazacyclododecane- N', N'', N'''- triacetic acid

Work is carried out analogously to example 14, and it is complexed with radioactive <111->indium chloride. To investigate whether the metal ions are completely bound as a chelate, the solution of the title compound is examined thin-layer chromatographically on silica gel plates in the system methanol-water (2:1). Non-chelated metal ions are thereby recognized as a radioactive zone at the starting spot. If necessary, the chelation is completed by further addition of N-(morpholino-carbonylmethyl)-N',N'',N'''-tris-(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane and subsequent ester cleavage.

Similarly, the gadolinium-153 complex of N-(morpholinocarbonylmethyl)-1,4,7,10-tetraazacyclododecane-N',N'',N'''-triacetic acid is obtained.

Claims (9)

1. 1,4,7,10-tetraazacyclododecane derivatives, characterized in that they have the general formula I: where each R<1>, independently of one another, is hydrogen or a metal ion equivalent; R2 is a linear or branched, saturated or unsaturated alkyl group of up to 16 carbon atoms, substituted with 1 to 5 hydroxy or C₁-C₁-alkoxy groups, -CH2-X-V where X has the meaning of carbonyl, a linear or branched alkylene group of 1- 10 carbon atoms which are optionally substituted with 1 to 5 hydroxy or C 1 -C 4 -alkoxy groups or a linear or branched alkylene group which is interrupted by oxygen atoms and which is 5 to 23 carbon atoms, and R<4> independently of each other is hydrogen, a linear or branched alkyl group with 1-16 carbon atoms, which is optionally substituted with 1 to 5 hydroxy or C₁-C₄-alkoxy groups, or R<3> or R<4 > together with the nitrogen atom is a pyrrolidine, piperidine, morpholine or piperazine ring, or R2 or R<3> is a further macroring of formula I' which is bound via an alkylene chain containing from 2 to 20 carbon atoms and which optionally has carbonyl groups in the end positions and is optionally interrupted by from 1 to 4 oxygen atoms or is substituted with from 1 to 5 hydroxy-, C-L-C 4 -alkyloxy or carboxy-C 3 -C 4 -alkylene groups, or R<2> denotes B or CH2-COB, where B denotes a macromolecule or biomolecule that accumulates to a particularly large extent in the organ or part of the organ being examined, or in the tumour, and, if desired, salts formed with free carboxyl groups and organic or inorganic bases, or with amino acids and basic groups with inorganic or organic acids. 2. Compounds according to claim 1, characterized in that R<1> stands for a hydrogen atom. 3. Compounds according to claim 1, characterized in that at least two of the substituents R<1> are metal ion equivalents of at least one metal with ordinal numbers 21-29, 42, 44 or 57-70. 4. Compounds according to claim 1, characterized in that at least two of the substituents R<1> are metal ion equivalents of at least one radionuclide of an element with the ordinal numbers 27, 29, 31, 32, 38, 39, 43, 49 , 64, 70 or 77. 5. Compounds according to claim 1, characterized in that these are N-[2,3-dihydroxy-N-methyl-propylcarbamoylmethyl]-1,4,7,10-tetraazacyclododecane-N',N'',N'''-triacetic acid, N-[N-ethylcarbamoylmethyl]-1,4,7,10-tetraazacyclododecane-N<*>,N'',N'''-triacetic acid, N-(2,3-dihydroxy-1-propyl)-1,4,7,10-tetraazacyclododecane-N',N'',N'''-triacetic acid, N-(2-aminoethyl)-1,4,7,10-tetraazacyclododecane-N',N'',N'''-triacetic acid, N-(2-hydroxyethyl)- 1, 4, 1,10-tetraazacyclododecane-N',N1',N''<1->triacetic acid, N-[N-(2-hydroxyethyl)-carbamoylmethyl]-1,4,7,10-tetraazacyclododecane-N<*>,N'',N'''-triacetic acid, N-(morpholinocarbonylmethyl)-1,4,7,10-tetraazacyclododecane-N',N'',N'''-triacetic acid, N-(morpholinocarbonylmethyl)-1,4,7,10-tetraazacyclododecane-N',N'',N''<*->tris-(2-methyl-acetic acid), and the complexes of these compounds with paramagnetic metals or radioisotopes. 6. Compounds according to claim 1, characterized in that these are gadolinium, manganese and dysprosium complexes of 1,1'-(1,3-propylene)-bis-(1,4,7,10-tetraazacyclododecane-4,7,10-triacetic acid), succinyl-bis-(1,4,7,10-tetraazacyclododecane-4,7,10-triacetic acid) and 1,1'-(2-hydroxy-1,3-propylene)-bis(1,4,7,10-tetraazacyclododecane-4,7,10-triacetic acid). 7. Diagnostic means, characterized in that it contains at least one metal complex with the general formula I according to claim 1 with at least two of the substituents R<1> in the sense of a metal ion equivalent, possibly with the additions that are common in galenics. 8. Use of the compounds according to claim 2 as complex formers. 9. Use of at least one metal complex of the general formula I with at least two of the substituents R<1> in the sense of a metal ion equivalent for the production of agents for NMR, X-ray, ultrasound or radio diagnostics.