IE872038L - 1,4,7,10-tetraazacyclodecane triacrtic acid complexes - 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

6 0 6 7 7 - i - The invention relates to the subject-matter characterised in the patent claims, that is to say, to macrocyclic complexing agentsE, complexes and complex salts,, to agents containing those compoundsf to their use as diagnostic 5 agents and therapeutic agent's and also to a process for the preparation of those compounds and agents. * Metal complexes were already being considered for use as contrast media for radiology at th© beginning of the 1950'"s. The compounds used at that time were f however, 10 so toxic that there was no question of using them in humans. It was therefore extremely surprising that certain complex salts proved to be sufficiently tolerable to be considered for routine use in humans for diagnostic purposes. As the first representative of this class of j 5 compound, the dimeglumine salt of Gd DTPA (gadolinium- (III) complex of diethylenetriarainepentaacetic acid) described in EP-A-71564 has hitherto proved to be very suitable as a contrast medium for nuclear spin tomography in clinical tests on over 1000 patients. The compound is 2o primarily used in connection with diseases of the central nervous system.

An important reason for the good tolerance of Gd DTPA in clinical use is its high degree of efficacy in nuclear spi n tomography, especially in the case of many brain 25 tumours. Owing to its high degree of efficacy, Gd DTPA can, at 0.1 mmol/kg body weight, be administered at very much lower doses than can, for example, X-ray contrast media in many X-ray examinations, Macrocyclic complexing agents andr especially, i 4,7.. iO-tatraaza-30 cyclododecane-M*' ,N" ,* ^""-tetraacetic acids which are the most closely related to the subject-matter off the invention, are known from FR-A-2539906, WO-A-8602352, C.A. 97 (1982)„ 2059502 and EP-A-124766.

Similar complexing agents are also described in the earlier publications EP-A-232751 and EP-A-238 196.

As a further representative of the complex salts, the meglumine salt of Gd OOTA (gadoliniutn( III) complex of 1,41, 7„ lo-tetraasacyclododecanetetraacetic acid) described in DE-A-34 01 052 has been found to be suitable for diagnostic purposes.

There is now, howeverf a desire to use chelates also at higher doses. This is the case especially for the purpose of identifying certain diseases outside the central nervous system by means of nuclear spin tomography (NMJR diagnostics), but very especially when using chelates as X-ray contrast media.

Chelates can offer a number of advantages over iodated X-ray contrast media: a) Radiation absorption in the higher energy range, and thus reduction of radiation stress on the patient and improvement of prerequisites for the energy subtraction method. b) avoidance of the unforeseeable, and in some cases even life-threatening or fatal, so-called "allergv-type" or cardiovascular side effects, known as "contrast media reactions", of present-day iodated X-ray contrast media.

The prerequisites are: - high concentration of radiation-absorbing elements in the solution (X-ray) or strong influence on the MMR signals pharmacokinetics suitable for diagnostics, very secure binding of the metal ions in excratahle complexes, even under i_n vivo conditions - good tolerance of the highly concentrated, high-dosage complex solution - low allergic potential of all constituents of the contrast medium - high degree of stability and long shelf-life of the chemical constituents of the contrast solution.

Those requirements apply to varying degrees and in different waysbut basically hold true for all applications of the mentioned complexes in in vivo diagnostics and, in some cases, in therapy.

The compounds according to the invention and the solutions prepared therefrom fulfil the mentioned requirements in a surprising manner. They have a strong activity that can be adapted to the particular principles of the diagnostic or therapeutic method (X-ray, NMR, ultrasound, nuclear medicine) by choosing suitable metal atoms.

The compounds according to the invention are used! 1. For NMR diagnostics in the form of their complexes with the ions of the transition metals of atomic numbers 21 to 29 and 4 2 and 44. 2 „ For NMR and X-ray diagnostics in the forra of their complexes with the ions of the lanthanide elements of atomic numbers 57 to 70 3. Suitable for ultrasound diagnostics are both those compounds that are intended for use in NMR diagnostics and those intended for use in X-ray diagnostics .

«... For radiodiagnostics and radiotherapy in the form o their complexes «5.th the radioisotopes of the elements of atomic number 27, 29, 31 s 32,, 38# 39, 43, 49, 62, 64, 70 or 11, Evan without specific measures, their pharmacokinetics permit an improvement; in the diagnosis of a large number of diseases. The complexes are for the most part excrete again rapidly and unaltered so that, especially when relatively toxic metal ions are used as active ingredient, no harmful effects that can be attributed to the metal are observed in spite of a high dosage.

The practical application of the novel complexes and complexing agents is also facilitated by their adequate, and often even very good, chemical stability.

Another important advantage of the complexes and complex ing agents described is their extraordinary chemical versatility. Apart from by the choice o£ the central atom, the properties can be adapted by the choice of a wide range of substituents and/or salt-forming agents to the requirements of efficacy, pharmacokinetics, tolerance, ease of use, etc. For example, it is possible to attain a specificity of the compounds, very desirable in diagnostics and therapy„ for structures in the organism,, for certain biochemical substances, for metabolic processes,, for conditions prevailing in tissue or body fluids, especially by coupling to biological substances or to substances having an interaction with biological systems. Such substances suitable for coupling may be of low molecular weight (for example glucose, amino acids,, fatty acids,, bile acids., porphyrins) or high molecular weight (polysaccharides, proteins,, antibodies, etc.) or may also be structures that are foreign to the body but that are distributed in the body in a specific manner or that react with constituents of the body. The use of such substances % The compounds according to the invention can also be used in radiotherapy in the form of their complexes »?ith radioisotopes, such as, for example, '-92rr„ Furthermore, the cotnplexing agents according to the invention are suitable as such or in the form of ^eak complexes with preferably body-specific ions (Ca2~, Mg2", Zn2~„ Fe2*/34" ) for the treatment of heavy metal poisoning or certain thesaurismoses™ The macrocyclic compounds according to the invention are characterised by the general formula I for 1,4,7,10-tetraazacyclododecane derivatives: wherein each R1, independently of the others, represents hydrogen or a metal ion equivalent, R5 represents hydrogen or a wethy1 or ethyl group, R2 represents a linear or branched, saturated or unsaturated alkyl group that; has up to 16 carbon s (I) atoms arid that is substituted by from 1 to b hydroxy or C^-C^-alkoxv croups, -CH2=X-V in «hich X represents carbonyl, a linear or branched alkylene group having from 1 to 10 carbon atoms that is optionally substituted by from 1 to 5 hydroxy or C-^-C^-alkoxy groups, or a linear or branched alkylene group that is interrupted by oxygen atoms and has from 2 to 23 carbon atoms,, /s3 V represents ~N , in which each of R3 and R4, XR4 independently of the other, represents hydrogen, a linear or branched alkyl group that has up to 16 carbon atoms and that is optionally substituted by from 1 to 5 hydroxy or C^-C^-alkoxv groups, or R3 and R4 together with the nitrogen atom represent a pyrrolidine, piperidine, morpholine or piperazine ring, or R3 represents a second macrocycie of the formula I' that is linked via an alkylene chain that has from 2 to 20 carbon atoms and optionally carries carbonyl groups at its ends and is optionally interrupted by from 1 to 4 oxygen atoms or substituted by from 1 to 5 hydroxy,, C^-C^alkoxy or carboxv-Ci -C^-alkvl groups, or represents B or CH2-COB, B representing a macro-molecule or bioraolecule that accumulates to an especially great extent in the organ or organ part to foe investigated or in the tumour, (!' ) - 7 - and functional groups present in the molecule are, if desired, conjugated with Eoioraoiecules, and the salts thereof with organic or inorganic bases or amino acids or with, inorganic or organic acids. 5 Compounds of the general formula I in which R1 represents hydrogen are referred to as cbmplexing agents and compounds of the general formula I in which at. least two qf the substituerkts represent a metal ion equivalent are referred to as metal complexes. 10 Suitable alkyl svabstituents E2, R3 and are saturated, unsaturated,, linear or branched hydrocarbons having up to 16 carbon atoms,, preferably saturated hydrocarbons which are substituted, optionally in the case of R3 and and by definition in the case of R2t, by frosa 1 to 5 hydroxy 15 or lower alkoxy groups.

Alkoxy groups having from 1 to 4 carbon atoms are especially methoxy and ethoxy groups.

There may be mentioned as optionally substituted alkyl groups,, for example, the methyl, ethyl, 2-hvdroxyethyl, 20 2-hydroxy-1~(hydroxyiaethyl)-ethyl, l-(hydroxymethvl )- ethyl„ propyl, isopropenvl, 2- and 3-hydroxvpropyl, 2,3-dihydroxypropyl,, butyls isobutenyla 2-, 3- and 4-hydroxy-butyl „ 2-, 3- and 4-hydroxy-2-i»ethylbutyl, 2- and 3-hvdroxyisobutyl-2, 3 , 4-trihydroxybutv 1 and 2-methoxy-25 ethyl groups.

Preferred are unsubstituted alkyl groups having from 1 to 7 carbon atoms, such as,, for example, the methyl,, ethyl, n-propyl, isopropyl, n-butyl, isobutyl,, pentyl and hexyl groups. Also preferred are mono- and poly-hydroxy-sub- 30 stituted alkyl groups having £row. 2 to 7 carbon atoms and from 1 to 5, preferably from 1 to 4, hydroxy groups,, such - 8 ~ 10 1 5 ast3 for example, 2- and 3-hydroxypropyl f 1,3-dihydroxy-isopropyl, I-(hvdroxymethyl)-ethyl, bis- and tris-(hydr-oxymethyl )methyl, 2, 3-dihydroxy-l-hydroxymethyIpropyl, 21,3,4 ,5 £, 6-pentahydroxyhexy 1 and preferably 2-hydroxy-ethyl,, 2-hydroxy-l-(hvdroxymethyl)-ethyl, 2,3-dihydroxy-propyl and 2,, 3,4-trihydroxybutyl.

When R3 and R4 together with the nitrogen atom represent a saturated five™ or siK-saembered ring optionally containing a further hetero atom,, represents pyrrolidine, piperidine, morpholine or piperazine.

The alkylene chain to which the second macrocvcle I1 is linked optionally carries carbonyl groups at its ends, and contains from 2 to 20 carbon atoms. It may be interrupted by frost l to 4 oxygen atom(s) or may be substituted by from 1 to 5 hydroxy, Cj_-4-alkoxy or carhoxy-C1-4-alkyl group(s), Alkoxy and alkyl groups having from 1 to 4 carbon atoms are especially methoxy, ethoxy, methyl and ethyl groups.

Examples are: ICH,J -ch,-d-ch2-„ - (chg!. - . -(CH2-CH2-0-CH2-CH2j-a - (CHjO-ch? 5 . JCKj-O-CHgJj-„ -CH2.CH,-SO-CH2-CH2J3-. -CH2-CM2-(0-CH2-CH23 4-„ -cmj-ch oh OH -CH-CH- 0M OH -Ciyc-.

CH,-C00H - cm-CM,, - . 4 1 I 4 oh oh When X represents carbon atoms, the preferred. an alkylene group having from 1 methylene group (i carbon atom) to 10 is If not all of the acidic hydrogen atoms are substituted by the central ion„ one, several or all of the remaining hydrogen, atom(s) may be replaced by cations of inorganic and/or organic bases or amino acids. Suitable inorganic cations are, for example, the lithium ion, the potassium ions the calcium ion and,, especially,, the sodium ion. Suitable cations of organic bases are, inter alia, those of primary* secondary or tertiary amines, such as, for example,, ethanolamine, diethanolamine, morpholine, glucamine,, N,N-dimethylglucamine and, especially, N-methylglucamine. Suitable cations of amino acids are, for example,, those of lysine,, arginine and ornithine.

The complex compounds can also be linked to macro-molecules that are known to accumulate to an especially great extent in the organ or organ part to be investigated., Such macromolecules are, for example, hormones, dextrans,, polysaccharides, polychelones, hydroxvethyl starch,, polyethylene glycol,, desferrioxamines, bleomycins,, insulin, prostaglandins, steroid hormones, amino sugars, amino acids4, peptides,, such as polvlysine, proteins (such as,s for example,, immunoglobulins and monoclonal antibodies) or lipids (also in the form of liposomes). Snecial orominence should be given to conjugates with albumins,, such as human serum albumin, antibodies,, such as, for example, monoclonal antibodies specific to tumour-associated antigens,, or antirayosin. Instead of the biomolecules it is also possible to link suitable synthetic polymers, such as polyethyleneimines. The diagnostic agents formed, therefrom are suitable,, for example,, for use in tumour and infarction diagnostics.

Suitable monoclonal antibodies for conjugation are especially those that are directed against antigens located predominantly at the cell membrane. Suitable as such are, for example, for tumour-imaging, monoclonal antibodies or their fragments (F(ab)2) that are directed, for example, against the carcinoemforyonal antigen (CE&)* human chorionic gonadotropin (fl-hCG) or other antigens located on the tumour, such as glycoproteins. Also suitable are, inter alia, anti-myosin, anti-insulin and anti-fibrin antibodies.

Suitable for liver investigations or for tumour diagnostics are, for example, conjugates or clathrates with liposomes (which are used, for example, as unilamellar or multi-lamellar phosphatidylcholine-cholesterol vesicles).

The macrocvclic compounds of the general formula I are prepared as follows:; in a manner known per se, in compounds of the general formula II wherein R5 is as defined above, " has the meaning given for R^, but is not to represent a macromolecule or biomolecule B or the group CH2-COB, and Z represents carboxy-protecting groups, the protecting groups Z are split off and the resulting acids (R1 in the general formula I represents hydrogen) are, if desired, a) reacted in a manner known aer se with at least one -11- or metal oxide or metal salt of an element of atomic number 21 to 29, 31, 32, 38, 39, 42 to 44, 49, 57 to 70 or 77 and then, if desired, acidic hydrogen atoms present are converted with inorganic and/or organic bases or amino acids and basic groups present are converted with inorganic or organic acids into physiologically tolerable salts, b) reacted in a manner known per se with at least one 10 metal oxide or metal salt of an element of atomic number 21 to 29, 31, 32, 38, 39, 42 to 44, 49, 57 to 70 or 77 and then the resulting metal complexes are linked in a manner known per se via functional groups contained in the molecule or to R2 or to the 15 CO group contained in R2 a macromolecule and, if desired„ acidic hydrogen atoms present are converted with inorganic and/or organic bases or amino acids and basic groups present are converted with inorganic or organic acids into physiologically toler-20 able salts,, or c) linked in a manner known per se via the functional groups contained in the molecule or to R2 or to the CO group contained in R2 a macromolecule and then 25 reacted, in a manner known per se, with at least one metal oxide or metal salt of ©n element of atomic number 21 to 29, 31, 32, 38, 39, 42 to 44, 49, 57 to 70 or 77 and then, if desired,, acidic hydrogen atoms present are converted with inorganic and/or 30 organic bases or amino acids and basic groups present are converted with inorganic or organic acids into physiologically tolerable salts.

Suitable carboxy-protecting groups Z are lower alkyl, aryl, and aralkyl groups, for example the methyl, ethyl,, 35 propyl, butyl, phenylbenzyl, diphenyliaethy 1, triphenyl- -12- methyl and bi s(p-n i trophenyI ) methyI groups and also trialkylsilyl groups.

The protecting groups Z are split off in a manner known per se„ for example by hydrolysis,, alkaline saponifies-5 tion of the esters., preferably with an alkali in aqueous- alcoholic solution at temperatures of from 0 to 50 "C or in the case of, for example, tert.-butyl esters, by means of trifluoroacetic acid.

The educts are prepared by cyclising two reactants; the '0 resulting cyclic compounds are thent, optionally after splitting off protecting groups., reacted with halogen alkanes,, halogen esters or halogen acids in order to introduce the substituent R^.

The cyclisation is effected in accordance with methods 15 known from the literature (for example Org. Synth. 58, 86 (1978), Hacrocyclic Polvether Syntheses, Springer Verlag Berlin, Heidelberg, Hew York, 1982, Coord. Cheta, Rev. 3, 3 (1968), Ann. Chem, 1976. 916) : one of the two reactants carries two leaving groups at the end of the chain and 2o the other carries two nucleophiles which displace those leaving groups. There may be mentioned by way of example the reaction of terminal dibromo-, dirnesyloxy-, ditosyloxv- or dialkoxvcarbonv1-aIkylene compounds containing a nitrogen atom with terminal diazaalkylene 25 compounds containing an additional nitrogen atom in the alkylene chain.

Hitrogen atoms present are optionally protected, for example in the form of tosvlatesand are freed,, before the subsequent alkylation reaction, in accordance with 30 processes known from the literature.

If diesters are used in the cyclisation reaction„ then the resulting diketo compounds must be reduced in accordance with processes known to the person skilled in the art, for example using diborane.

The subsequent alkylation is carried out using halogen esters, halogen acids or halogen alkanes which may be substituted by one or more hydroxy or lower alkoxy groups and optionally contain oxygen atom(s) in the chain. The alkyl radical may also contain a terminal amino group.

If the alkylation is carried out with a dihalogenated alkane, compounds of the general formula II are £ortaed having two macrocyclic rings connected via a carbon bridge.

Other processes known from the literature for the synthesis of compounds having more than one ring are, for example, reactions of an amine with a carbonyl compound (for example acid chloride, mixed anhydride, activated ester? aldehyde); of two amine-substituted rings with a dicarbonvl compound (for example oxalvl chloride, glutaric dialdehyde) ,• of two rings each having a nucleo-philic group with an alkylene compound carrying two leaving groups; in the case of terminal acetyls, oxidative coupling (Cadiot, Chodkiewics in Viehe "Acetylenes", 597-647, Marcel Dekker, Mew York, 1969). The chain linking the rings can subsequently be .modified by secondary reactions (for example hydrogenation).

The hydrolysis, which may be necessary,, of the ester groups formed during the alkylation with halogen esters is carried out in accordance with methods known to the person skilled in the art (for example using basic catalysts, such as alkali metal or alkaline earth metal carbonates or hydroxides).

In the alkylation with haloacetic acid, an intermediate product is obtained in which = CH2X-COOH and which is converted into the raonoamide by way of the mixed anhydride using chloroforinic acid ester or by means of dicyclohexylcarbodiimide and reaction with a primary or secondary amine of the general formula R3 HN Examples of suitable amines are: dimethyl amine, diethylaraine, di-n-propvl amine, diiso-propylamine„ di-n-butylamina, diisobutvlamine, di-sec.-butylamine, H-methyl=n-propylarnine, dioctylamine, morpho-line,, pyrrolidine,, piperidine, pyrazoline, 2,3-dihydroxv-propylamine,, W-methyl-2,3-dihydroxvpropylamine, 2-hydroxy-1™ (hydroxvmethy1) -ethylamine , N , M-bis ( 2-hydroxv-ethyl)amine, N-methyl-2,3,4,5,6-pentahydroxvhexylamine, 2-hydroxyethvlamine, 2-amino-l,3-propanediol, diethanol-araine and ethanolamine.

The polyhydroxyalkylaraines can advantageously also be used for reaction in protected form, for example in the form of O-acvl derivatives or in the form of ketals. This applies especially when those derivatives are easier and cheaper to prepare than are the polyhydroxyalkylamines themselves , A typical example is the 2-amino-l-(2,2-dimethyl-1g 3-dioxolan-4-yl) -ethanol e the acetonide of 1-araino-2„3,4-trihydroxybutane, prepared in accordance with DE-A— 31 50 917.

The subsequent removal of the protecting groups does not present problems and can be effected, for example, by treatment with an acidic ion exchanger in aqueous-ethanolic solution. - ! 5 - The compounds of the general formula I in which R~ represents a hydrogen atom are complexing agents. They can be isolated and purified, or they can, without being isolated, be converted into metal complexes of the 5 general formula I i« which at least two of the sub- stituents R1 represent a metal ion equivalent.

The snetal complexes according to the invention are prepared in the manner disclosed in OE-A-34 01 052 and EP-A-71564 by dissolving or suspending the metal oxide 10 or a metal salt (for example the nitrate, acetate, carbonate, chloride or sulphate) of the element of atomic number 21 to 29, 31, 32, 38, 39* 42 to 44, 49, 57 to 70 or 77 in water and/or a lower alcohol (such as methanol, ethanol or isopropanol) and reacting ?•/! th a 15 solution or suspension of the equivalent amount of the complexing acid of the general forrcula I in which R1 represents a hydrogen acom and then, if desired, replacing acidic hydrogen atoms of acid groups present by cations of inorganic and/or organic bases or amino acids. 20 Neutralisation is carried out in this process with the aid of inorganic bases (for example hydroxides, carbonates or hydrogen carbonates) of, fcr example, sodium, potassium or lithium, and/or with the aid of organic bases, such as, inter alia, primary, secondary and 25 tertiary amines, such as, for example, ethanolamine, morpholine, glucamine, N-methyl- and N,N-dimethyl-glucamine, and also basic amino acids, such as, for example, lysine, arginine and ornithine.

In order to prepare the neutral corrplex compounds, it is 30 possible, for example, to add to the acidic complex salts in aqueous solution or suspension an amount of the desired bases that is sufficient to reach neutralisation. The solution obtained can then be concentrated to dryness ir> vacuo. It is often advantageous to precipitate the resulting neutral salts by the addition of water-miscible solvents, such as, for example, lower alcohols (methanol, ethanol, isopropanol and others), lower ketones (acetone and others), polar ethers (tetrahvdro-furan, dioxane, 1,2-dimethoxyethane and others) in order thus to obtain crystal).isates that are readily isolated and purified* It has proved especially advantageous to add the desired base to the reaction mixture as early as the coioplexing stage and thereby to eliminate one process step.

If the acidic complex compounds contain several free acidic groups it is often advantageous to produce neutral mixed salts which contain both inorganic and organic cations as counter-ions.

That can be effected, for example, by reacting the complexing acid in aqueous suspension or solution with the oxide or salt of the element yielding the central ion and half the amount of an organic base necessary for neutralisation, isolating the complex salt formed, purifying it, if desired, and then adding the required amount of inorganic base for complete neutralisation. The sequence in which the bases are added can also be reversed.

Basic groups can be converted into pharmaceutical^ acceptable salts using inorganic and/or organic acids.

There are used as inorganic bases, for example, lithium, sodium and potassium hydroxide. Suitable organic bases are, inter alia, primary, secondary and tertiary amines, such as, for example, ethanolamine, morpholine,, glue™ amine, N-methyl- and N,N-dimethyl-glucaraine, and also basic amino acids, such as, for example, lysine, arginine and ornithine. -17- Inorganic acids, such as, for example, hydrochloric acid, and organic acids, such as, for example, citric acid, are suitable for salt formation with basic groups, Conjugate formation can be carried out, for examples by 5 way of a carboxy group of the complex compound or by way of a functional group- When forming the conjugate of acids with »acromo1 ecu1as, several acid radicals can be bound to the macromolacule» In that case several central ions may be bound to one macromolecule* 10 Coupling to the desired macromolecules is likewise carried out in accordance with methods known per se, such as described,, for example, in Rev. Houra. Morphol„ Embryo1„ Physiol„3 Physiologie 1981, 18, 241 and J.

Pharm. Sci„ 68, 79 (1979), for example by reacting the 15 nucleophilic group of a macromolecule, such as the amino, phenol , sulphhydryl, aldehyde or imidazole group, with an activated derivative of the complexing agent. Suitable activated derivatives are, for example, mono-anhydrides „ acid chlorides, acid hydrazides, mixed 20 anhydrides (see, for example# G.E. Krejcarek and K.L. Tucker, Biochem. Biophvs. Res. Commun. 1977, 581), activated esters, nicrenes or isothiocvanates. Conversely s it is also possible to react an activated macro-molecule with the complexing acid. Substituents having, 25 for example, the structure CgH4HHCOCH2/ C^H^NHCS or CgH^OCf^CO are also suitable for conjugation with proteins.

The conjugation of the complexing acid with dextrans and dextrins is likewise effected in accordance with methods 30 known oer se„ for example by activating the polysaccharides with cyanogen bromide and then reacting with amino groups of the complexing acid. - 18 - When complex compounds containing radioisotopes are used, their preparation can be effected in accordance with the methods described in "Radiotracers for Medical Applications", Volume CRC-Press, Boca Raton, Florida. 5 The pharmaceutical agents according to the invention are likewise prepared in a manner known per se by suspending or dissolving the complex compounds according to the invention„ optionally with the addition of adjuvants customarily used in galenical pharmacy, in an aqueous 10 medium and then optionally sterilising the suspension or solution. Suitable adjuvants are,, for examplef physiologically acceptable buffers (such as, for example, tromethamine), small amounts of complexing agents (such as, for example, diethylenetriaminepentaacetic acid), or, 15 if necessary, electrolytes, such as, for example, sodium chloride, or, if necessary, anti-oxidants, such as, for example, ascorbic acid.

If suspensions or solutions of the agents according to the invention in water or physiological saline solution 20 are desired for enteral administration or other purposes, they are mixed with one or more adjuvant(s) customarily used in galenical pharmacy (for example methylcellulose, lactose, mannitol) and/or surfactant(s), (for example lecithins, Tween(R), Myrj(R)) and/or flavourings to 25 improve the taste (for example essential oils).

In principle it is also possible to prepare the pharmaceutical agents according to the invention without isolating the complex salts. Special care must in any case be taJcen to effect chelate formation in such a 30 manner that the salts and salt solutions according to the invention are practically free of uncomplexed toxically active metal ions.

That can be ensured, for example, by means of colour indicators, such as xylenol orange, by control titrations during the preparation process. The invention accordingly also relates to processes for the preparation of the complex compounds and their salts. Purification of the isolated complex salt can be used as a final safety measure.

The pharmaceutical agents according to the invention preferably contain from 1 pmol to 1 mol/1 of the complex salt and are generally administered in doses of from 0.001 to 5 mmol/kg„ They are intended for enteral and parenteral administration.

The agents according to the invention meet the many and various prerequisites for suitability as contrast media for nuclear spin tomography. For example, they are excellently suited, after oral or parenteral administration, for improving the information content of the image obtained by means of the nuclear spin tomograph, by increasing the signal intensity. They also exhibit the high degree of efficacy that is necessary in order to minimise the burden of foreign substances on the body, and the good tolerance that is necessary to maintain the non-invasive character of the investigations.

The good water-solubility of the agents according to the invention enables highly concentrated solutions to be produced so that the volume-load on the circulation can be kept within tolerable limits and dilution by body fluids can be compensated for„ In addition, the agents according to the invention not only have a high degree of stability 1b vitro but they also have a surprisingly high degree of stability in vivo, so that a release or an exchange of the ions that are not covalently bonded in the complexes, and which are toxic oer se„ takes place - 20 - only extremely slowly during the time in which the novel contrast media are completely excreted again.

Generally,, the agents according to the invention are, for use as NMR diagnostic agents, administered in doses of 5 from 0.001 to 5 mraol/kg„ preferably from 0.005 to 0.5 mmol/kg„ Details of usage are discussed, for example, in H.J. Meinmann et al,. P Am. J. of Roentgenology 142, 519 ( 1984) .

Especially low doses (less than 1 rag/kg) of organ-10 specific NMR diagnostic agents can be used, for example, for the detection of tumours and cardiac infarctions. The complex compounds according to the invention can also advantageously be used as shift reagents.

Owing to their favourable radioactive properties and the I5 good stability of the complex compounds contained in them, the agents according to the invention are also suitable as radiodiagnostic agents. Details of their use and dosage are described, for example, in "Radiotracers for Medical Applications", CRC-Press, Boca Raton, 20 Florida.

Another imaging method with radioisotopes is positron emission tomography which uses positron-emitting isotopes, such as, for example, ^3Sc, 4^Sc, ^2Fe, 5^Co and 68Ga. (Heiss, W.D., Phelps, M.E., Positron Emission 25 Tomography of Brain„ Springer Verlag Berlin, Heidelberg, New York 198 3.) The compounds according to the invention can also be used in radioimmunotherapy. This procedure differs from the corresponding diagnostics only by the amount and type of 30 radioactive isotope used. The aim is to destroy tumour cells by high-energy short-wave radiation with as short a range as possible. The specificity of the antibody used is of crucial importance since non-specifically localised antibody conjugates lead to the destruction of healthy tissue.

The antibody or the antibody fragment of the antibody-metal complex according to the invention serves to transport the complexin an imraunospecific manner irs relation to the antigen concerned, to the target organ where the metal ion selected for its cytocidal properties can emit radiation that causes lethal damage to the cells. Suitable 6-emitting ions are, for example,, 4SSc, 47Sc, 48Sc„ 72Ga and 73Ga. Suitable ©-emitting ions having low half-life values are, for example* 21iBi„ 212Bi9 213Bi and 214Bif, 2i2Bi being preferred.

In the case of in vivo administration of the therapeutic agents according to the invention, the agents can be administered together with a suitable carrier, such as, for example, serum or physiological saline solution,, and together with another protein,, such asP for exanple, human serum albumin. The dosage depends on the type of cellular disorder, the metal ion used and the type of imaging method.

The therapeutic agents according to the invention are administered parenterally„ and preferably intravenously. Details of the use of radiotherapeutic agents are discussed, for example„ in R.w. Kozak et al„ TIBTEC, October 1986,, 262.

The agents according to the invention are outstandingly suitable as X-ray contrast media and special attention should be drawn to the fact that they do not give rise to any signs of the anaphylaxis-type reactions known for iodine-containing contrast media in biochemical-pharsna- cological studies. They are especially valuable for digital subtraction techniques owing to their favourable absorption properties in areas of relatively high tube voltages.

In general „ the agents according to the invention are administered, when used as X-ray contrast media,, analogously to, for example, meglumine-diatri 2oate, in doses of from 0-1 to 5 nns,oI/kg? preferably from 0.25 to 1 anool/kg.

Details of the use of X-ray contrast media are discussed, for example, in Barke, Rdntgenkontrastaittel (X-ray Contrast Media),, G. Thieme, Leipzig (1970) and P. Biurn, E. BGcheler - "Sinfiihrung in die 35ntgendiagnostik" (Introduction to x~Ray Diagnostics), G. Thieme, Stuttgart,. Mew fork (197?).

Because their acoustic impedance is greater than that of body fluids and tissues, the agents according to the invention are also suitable as contrast Media for ultrasound diagnostics, especially in the form of suspensions. They are generally adninistered in doses of from 0-1 to 5 unol/kg, preferably from 0.25 to 1 aasaol/kg.

Details of the use of ultrasound diagnostic agents are described, for example, in T.B. Tyler gt , Ultrasonic Xaaging 3,323 (1981), J.I. Haft, "Clinical Echocardiography" , Futura, Mount Kisco„, New York 1978 and G. Stefan "Bchokardiographie" G. Thieve Stuttgart/New York 1981.

In summary, it has been possible to synthesis© novel cosaplexing agents, metal complexes and metal complex salts that open up new possibilities in diagnostic and therapeutic medicine. This development would appear to be - is - desirable above all in connection with the development of new types of imaging processes in medical diagnostics.

The following Examples serve to illustrate the subject-matter of the invention in more detail. 5 In the following«, the synthesis of starting material,, starting with a cyclisation reaction, is described by way of examples a) l-benzvl-4.7 ,10-tris(o-tolvlsulfonvl)-1,4,7.10-tefcra-azacvclododscane 145 g of M,M-his[2,2*-(g-tolylsulphonyloxy)Jethane-benzylaxnine, dissolved in 900 ml of dimethy 1 forrnamide, are added dropwise, with stirring, at 100*C over a period of 3 hours to a solution of 164.6 g of the N,N"-disodiura salt of M,M'' ,E^,a-tris(Q-tolylsulphonyl)diethv1-enetriamine in 2.16 litres of dimethylformamide. 1 litre of water is then added dropwise„ with stirring, at 80"C and the whole is stirred for a further 18 hours at room temperature, then cooled to 0*C and the precipitate is filtered off with suction, washed with a small amount of ice-cold ethanol and dried at 15 torr and 60"C. 175 g of the title compound are obtained- Alternative Route 1 An analogous method for the preparation of tetraazacvclo-dodecane derivatives can be found in M. Hediger and T.A. 2 5 Kaden, Helv„ Chim. Acta 56, 861 (1983). 30-94 g of the W,N"-disodium salt of N,N",N"-tris(g-tolylsulphonyl Jdiethylenetriaraine and 2 8.12 g of N-bis(2-methanesu1phonyloxvethyl)triphenyl ine t h y 1 a in ine are stirred with 530 ml of dimethylf ormamide for 20 hours at 30 80-85*C„ then cooled and stirred into a solution of 30 g of potassium carbonate in 5 litres of ice-water. The 1 5 24 - precipitate is filtered off with suction and the filter cake is washed with 0.5 litre of water and dried at 20"C in vacuo at 150 torr. For pur if icationl7 the product is dissolved in 230 sal of chloroform and 5 ml of triethyl-5 amine* The solution is filteredf concentrated .in vacuo to 200 rel and 250 ml of ethyl acetate are added to the solution at boiling point - The whole is allowed to cool overnight and the precipitated crystals are filtered off with suction. 22.13 g of 1,4,7-tris(B-tolylsulphonyl)-10-10 triphenylmethyl-1,4„7,10-tetraazacyclododecane are obtained, m.p.: 185~188"C (decomposition).

In order to split off the trityl protecting group, 31.4 g of the tritosvltrityl derivative obtained in this manner 15 are stirred in a mixture of 100 ml of glacial acetic acid, 75 ml of water and 300 ml of dioxane for 1 hour at 80*C. The whole is then extensively concentrated in vacuo at s 0*C and diluted with 300 ral of ice-water„ and 40 ml of UN sodium hydroxide solution (pH above 12) are added. 2Q The mixture is shaken with 300 nil of chloroform., the phases are separated, the aqueous phase is extracted twice with 100 ral of chloroform each time and the combined chloroform phases are dried over sodium sulphate and concentrated by evaporation in vacuo. The foamy 25 residue is treated with 300 ml of diethyl ethere, where upon crystallisation occurs. The crystals are filtered off with suction and dried In vacuo at 60*0 and 150 torr to yield 21 g of 1,4,7-tris(p-tolylsulphonyl)-1,4,7,10-fcetraazacyclododecane, 2q 135®p. " 202-203 O.

The tritosyl compound so obtained (21 g) is dissolved in 200 ml of dimethylformamide,, and 13.71 g of anhydrous potassium carbonate, 4.95 g of sodium iodide and 7.92 g of benzyl bromide are added in succession to the solution - 25 - and the whole is stirred for 5 hours at 100*0, The mixture is then cooled to 20'Cg stirred into 4 litres of ice-water and filtered with suction and the residue is dissolved in 2 litres of dichlororaethana» The solution is 5 extracted with 100 ml of water, dried over sodiuw sulphate and concentrated by evaporation jjQ v$cuo. The residue is dissolved at boiling point in 500 sal of acetonitrile and is left to crystallise out overnight. The crystals are filtered off with suction and dried at 10 50 "C and 150 torr to yield 16.20 g of l-benzyl-4„7„10-tris(a-tolylsulphonyl)-l ,4,7,10-tetraazacyclododecane, n.p.: 217-219*C, lb) M-benzvl-1 . 4 7 ,10-tetraazacvclododecane 150 g of l-benzyl-4 7,, 10-tris (o-tolylsulphonyl) -I, f 7-15 10-tetraazacyclododecane are heated for 16 hours at 50*C with 900 ral of HBr/acetic acid (40 % strength) and 125 g of phenol. After cooling to 20 C, the whole is diluted ferith 1 litre of ether,, cooled to ~5 C and the precipitated crystals are filtered off with suction. In order to 20 isolate the free bases the product is dissolved in 500 ml of 4N sodium hydroxide solution, saturated wi th potassium carbonate and extracted several times with chloroform, dried over magnesium sulphate and concentrated by evaporation irj vacuo. 39 g of the title compound are 25 obtained in the form of a pale ye 1 low viscous oil. A sample was characterised as the trihvdrochlorids: melting point 210 C (with decomposition) „ Alternative Route 2 58 rel of triethylawine are added at -20"C to a solution 30 of 11.2 g of 1,4,7,10-tetraazacyclododecane in 900 ml of itetrahydrofur&n and a solution of 16.2 sal of benzoyl chloride in 280 snl of tetrahydrofuran is added dropwise with stirring over 3 hours, the room temperature rising to slightly above -10C. Tha whole is then stirred for 16 hours at 0-10'Ce tha precipitate is filtered off and the solution is concentrated by evaporation jja vacuo. The residue is chromatographed on 1 kg of silica gel and eluted with dioxane/water/arnmon1a solution (8:1:1). The fractions that are uniform according to thin layer chromatography are combined, concentrated by evaporation, dissolved in dichloromethane and filtered to remove a small degree of turbidity and the solution is concentrated by evaporation to yield 19.60 g of 1,4,7-triben~ zoyl-l,4 /?, 10-tetraazacyclododecane, M.p.s 1.20—125*C- 11.5 g off the resulting tribenzoste are stirred in 150 ml of dimethylforraamide 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» The whole is filtered, the solution is concentrated by evaporation in vacuo, the residue is stirred twice v;ith 50 ml of hexane each time and decanted., The hexane phases are discarded. For pur if ication, the product is dissolved in dichloromethane and chromatographed with dichlorornethane/methanol f 37 : 3 ) on 0.5 kg of silica gel10.2 g of 1,4 ,7-triben~ zoyl-10-benzyl-l,4,7,10-tetraazacyclododecane are obtained,, m.p.: 105-109 "C.

In order to split off the benzoyl groups, the product (2.87 g) is dissolved in 290 ml of tetrahydrofuran. 11.2 g of potassium terfc.-butoxide are added and the whole is heated under reflux for 4 8 hours. It is then filtered and concentrated by evaporation in vacuo and 3.00 ml of water are added to the residue while cooling with ice and the whole is extracted three times with 5© ml of dichloromethane each time. The combined dichlor- - 27 - omethane phases are shaken with 10 ml of water, dried over sodium sulphate and concentrated by evaporation i_Q vacuo. The residue,, which is at first oily,, crystallises very slowly and is triturated with 20 ml of hexane. After filtering off with suction and dryings 1.15 g of N~ benzyl-1,4,7,10-tetraazacyclododecane are obtained, in.p.: 75-7S"C. c) 1 tetraazacvcloclodecane 1q 200 g of triethylamine and., over 2 hours, 260 g of bromoacetic acid ethyl ester are added in succession at 0 *'C to a solution of 131.8 g of M-benzyl-l „ 4 „ 7,10-tetraazacyclododecane irs 1.5 litres of dichloromethane. The who Is is stirred for a further 16 hours at roora 15 temperature,, is shaken with 5 % sodium carbonate solution and brine, dried over raagnesiura sulphate and concentrated by evaporation in vacuo. The residue is dissolved in 200 ml of chloroform and filtered over 2 kg of silica gel f, elution being effected with 1 litre of chloroform/-20 methanol (95 5) . 210 g of the title compound are obtained in the form of a viscous oil. d) N . N" , N68-trisf ethoxvcarbonvIroethvl)-1, 4 . 7,10-tetraaza-cvclododecane 100 g of l-benzyl-4 171 10-tris(ethoxycarbonylme thyl)-2,4#7„10-tetraazacyclododecane are dissolved in 0.5 litre of acetic acid and 0-5 litre of ethyl acetate, and. 5 g of palladium-on-carbon (10 %) are added, and the whole is shaken for 5 hours under hydrogen. The whole is filtered off from the catalyst and concentrated in vacuo. The residue is dissolved in 1 litre of chloroform,, shaken with 100 ml of saturated soda solution and 100 ral of brine,, dried and concentrated by evaporation jjj vacuo. - 28 - The residue is purified by bulb tube distillation at !0"J torr and 12 0" C. N,N',N"~tris(ethoxycarbonylmethyl) ~ 3,, 4 , 7 „ 10-tetraazacyclododecane is obtained in the form of a pals yellow viscous oil.

Yield- 65 g IR (film): 3400, 2935, 2B7B, 1738/cm.

Example 1.

Gadolinium(III) complex of N-C2„ 3-dihydroxy-N-methyl-propylcarbamoylmathyl ]-1,4,7,10-tetraa zacyclododecane- N' „ N", M" ?l-triacetic acid 55 g of N-[ 2, 3-dihydroxy-M-raethylpropylcarbanoyliaethyl |-N19 ,NM,N"'-tris(ethoxycarbonylraethyl)-l,4e?( 10-tetraazacyclododecane are dissolved in 0.5 litre of ethanolt and 95 stl of 3N sodium hydroxide solution are added to the solution and the whole is stirred for 3 hours at 20"C and then concentrated in vacuo,. 300 sal of water are added and the pH is adjusted to 6 with 2N hydrochloric acid. 31,94 g of gadolinium acetate are added to the solution and the whole is stirred for 18 hours at 50*0, then passed over an anion exchanger Amber 1 ire IRA 410 and then the aqueous eluate is passed over a cation exchanger Amber1 ire IRC 50- The eluate is concentrated by evaporation in vacuo and dried.., 47.14 g (73 % of the theoretical yield) of the title compound are obtained in the forxa of a colourless powder.

Analysis: Cgg H34 Gd N5 O© (545.77) C 37-20 H 5.31 Gd 24.35 N 10.85 (calc„) 37-52 5-19 24.09 10.87 (found) The starting material for the preparation of the title - 2V - compound according to Example 2 is obtained in the following manners a) N,N', N"*-tris (ethoxycar bony line thy 1) -1, , 7, 10-tetraaza-cyclododecane-N' "-acetic acid 5 20 g of #?' (ethoxycarbonylmethyl)-1,4,7, 10- tetraazacyclododecane are dissolved in 300 sal of di-chloromethane. 10.40 g of triethylastine are added and then a solution of 4.40 g of chloroacetic acid in 100 wl of dichlorowetinairae is added dropwise at o"C arsd the whole 10 is stirred for 20 hours at room temperature. For working-up, the mixture is partitioned between dichloromethane and phosphate buffer pH 6, dried over magnesium sulphate and concentrated by evaporation la vacuo. 23 g of the desired compound are obtained in the form of a viscous 15 oil. to) M- £ 23-dihydroxy-M-methylpropylcarbamoylmethyl )-N * , NM , N" ?-tris(ethoxycarbonylmethyl )-l ,4,7,10-tetraazacyclododecane 10.5 g df triethylamine and then a solution of 14 g of 20 chloroforinic acid isobutyl ester are added dropwise at 0"C to a solution of 48.86 g (100 mmol) of H,h'''..Mca-tris (ethoxycarbonylmethyl) -i4 ,7, lO-tetraazacyclo-dodecane-Nr"-acatic acid in 500 ml of dichloromethane. The ^hole is stirred for 1 hour at 0"C and then a 25 solution of 10.52 g of N-Mtethylamino-2,3-propanediol in 10*0 ml of chloroform is added dropwise thereto and stirred for 2 hours at room temperature. After shaking with sodium hydrogen carbonate solution and brine and drying over wagnesium sulphate, the product is concen-30 trated by evaporation in vacuo. For purification, the chloroform solution of the residue is filtered over 500 g of silica gel to yield 55 g of the title compound.

JO - Example 2 Gadoli niua(III) complex of N-[N-ethylcarbamoylmethyl]-1,4,7,10-tetraazacyclododecane-N',N",M"'-triacetic acid 24 g ot N-£iSf-efchylcarbamoyliaethyl J-K* '-trisfethoxy- 5 carbonylsaethyl)-lJ,4i?lilO-tetraazacyclodod«cane are . , dissolved in 250 sal of ethanol- 4? ml of 3N sodium hydroxide solution are added to the solution and the whole is stirred for 3 hours at 20"C and concentrated Jjj vacuo- 150 wl of water are added and the pH is adjusted 10 to 6 with 2N hydrochloric acid. 15.55 g of gadolinium acetate are added and the whole is stirred for 5 hours at 60 "C- The solution is then purified by means of ion exchangers as described in the preceding Examples. 20.39 g of the title compound are obtained in the form of 15 a colourless powder.

Analysis: Ciq H3Q Gd N5 O7 (585»72) C 36.91 H 5.16 Gd 26.85 N 5.16 (calc,) 36.98 5.28 26.58 5.07 (found) Preparation of the starting material: 20 25 g (51.2 mmol) of N „ N NM-tris (ethoxycarbonylmethyl) -1, 4 , 7s3.0-tetraazacyclododecane-N"*-acetic acid (see Example 2a) are dissolved in 200 ml of dichloromethane, and then 5.40 g of triethy.lamine followed by 7 g of chloroformic acid isobutyl ester are added at 0 * C. The 2 5 whole is stirred for 1 hour at 0™C and then a solution of 2 - 31 g of ethylamine in 20 ml of dichloromethane is added dropwise. The mixture is stirred for 2 hours at roos temperature and, after shaking with saturated sodium hydrogen carbonate solution and Ibrine and drying over 30 magnesium sulphate, the product is evaporated to dryness in. vacuo. For purification,, the dichloromethane solution -31- of the residue is filtered over 200 g of silica gel to yield 24 A viscous oil is obtained which is stirred for 4 hours with 40 ml of 1M sodium hydroxide solution,, then diluted with water (100 ml) and adjusted to pH 6 with 2M hydro-20 chloric acid. After the addition of 3.34 g of gadolinium acetate,, the whole is stirred for 16 hours at 50"c and the solution is purified by means of ion exchangers to yield 3.90 g of the title compound in the form of a white powder. 25 Analysis: C32 Gd2 Ng Oj (1083-28) C 35.48 H 4.47 Gd 29.03 N 10.34 (calc.) 35.31 4.18 28.81 10.57 (found) Example 7 N-(2-hydroxysthy1)-1,4 p7,10-tetraazacyclododecane-N ' , N" , N" '-triacetic acid 3.2 q of triethylamine and 1.45 g of 2-chloroethanol are added to a solution of 6.46 g of KfN' sH,,™tris(ethoxy-carbonylmethyl )~J., 4, 7f 10-tetraazacyclododecane in 150 ral of dichloromethane. The whole is stirred! for 4 hours at room temperature andt. after shaking ~»ith sodium hydrogen carbonate solution and brine„ the product is dried over magnesium sulphate and concentrated by evaporation in, vacuo. The residue is stirred for 16 hours with 60 ml of IN sodium hydroxide solution- The pH is adjusted to 2.5 by the addition of 5N hydrochloric acid. The resulting suspension is poured onto an ion exchanger (DOWEX 50W-X4 in the H4" form) and then eluted with water followed by 0.5M HH3 solution. The whole is concentrated in vacuo and the title compound is isolated by adding ethanol and filtering off the precipitate with suction. 4.24 g of the title compound are obtained, the purity of which is checked by pH titration and elemental analysis.

Analysis^ Ci6 H30 N4 07 (390.44) C 49.22 H 7.75 N 14.35 (calc.) 49.48 7.83 14.09 (found) Example 8 N-(N-(2-hydroxyethyl)-carbamoylmethylJ-l,4„7,10-tetraazacyclododecane -i"? ' sN"„M"'-triacetic acid 5.22 g of triethylamine and then 3.50 g of chloroformic acid isobutyl ester are added at 0"C to a solution of 12.50 g of N„N',H"-tris(ethoxycarbonylmethyl)-ls4#7,10- - 35 - tetraazacyclododecane-H"' -acetic acid in 250 ml of dichloromethane. After 1 hour, 1.60 g of ethanolamine dissolved in 50 ml of dichloromethane are added dropwise, the whole is stirred for 2 hours at room temperature and, 5 after shaking with soda solution and brine,, the product is dried over magnesium sulphate and concentrated by evaporation in vacuo. The residue Is chromatographed with chloroform/acetone (10si) on 200 g of silica gel to yield 11 g of N- (2-hvdroxyethyl) carbamoylmethyl-M®, N"', NM * -1 o tris(ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane .

The product is stirred for 5 hours at room temperature with 100 ml of IN sodium hydroxide solution and then acidified to pH 2.5 with dilute hydrochloric acid and the 15 suspension is purified by means of a cation exchanger (DOWEX 50W-X4)„ elution being effected with water and then with 0.5M NH3 solution. The eluate is extensively concentrated and, after the addition of ethanol, the title compound crystallises out and is isolated by 20 filtration, 7.2 g of the title compound are obtained, the purity of which is examined by titration and elemental analysis.

Analysis: C^g H33 N5 Oq (447.49) C 48.31 H 7.43 N 15.65 (calc.) 25 48.20 7.48 15.49 (found) Example 9 Gadolinium complex of N-(raorpholinocarbonylmethyl)-1,4,7,10-tetraazacyclododecane-H ' „ Ns", N"' -triacetic acid 30 55.77 g of N-(morpholinocarbonylmethyl)-N"fNw,NM tris(ethoxycarbonylmethyl)-1,4,7,lO-tetraazacyclododecan® are hydrolvsed analogously to Example 1 with sodium hydroxide solution and complexed with gadolinium acetate 45.19 g (72 % of the theoretical yield) of the title compound are obtained in the form of a white powder.

The starting material for the preparation of the title compound is obtained by proceeding as in Example 2b but using morpholine instead of N-methylamino-2,3-proparie-diol.

Analysis; C20 H32 Gd N5 0@ (527.75) C 3 8.27 H 5.14 Gd 25.04 N 11.16 (calc.) 38.02 5.09 24.83 11.35 (found) Example 10 Gadolinium complex of M-( morphol inocarhorty lmethyl)-1,4,7, lO-tetraasacyclododecane-N* ,N" , Mn # -tr is( 2-methyl-acetic acid) 53.06 g of N-(morpholinocarbonylmethyl)-n' ,N" ,NIB ''-tris(1 ethoxycarbony1-1-ethyl) ~ 1, 4,7,10-tetraazacyclododecane are hvdrolysed analogously to Example 2 with sodium hydroxide solution and complexed with gadolinium acetate 52.20 g (78 % of the theoretical yield) of the title compound are obtained in the form of a white powder.

Analysis: C23 H3q Gd N5 08 (669.83) C 41.24 H 5.72 Gd 23.48 N 10.46 (calc.) 41.35 5.65 23.33 10.62 (found) Example 11 Bis-gadolinium complex of !,!'-(2-hydroxy-ls3-propylene) bis(1,4,7,10-tetraazacyclododecane-4,7,10-triacetic acid) - 37 - 4.63 g ot epichlorohydrin are added to a solution of 43.05 g of H , NN,d-tris( ethoxycarbonylmethyl )-l, 4 ,7 10-tetraazacyclododecane in 450 ml of diraethylformamide. After 1 hour, 7.5 g of sodium iodide are added and tha 5 whole is heated for 24 hours at 80"c and then concen trated In vacuo. The residue is partitioned between water and chloroform,, the chloroform phase is dried over sodium sulphate and concentrated by evaporation In vacuo. The residue is chromatographed on I kg of silica gel with 10 dichlororaethana/10% acetone to yield 27„5 g of hydroxy-1, 3-propylene) -bis (1,4,7,lO-tetraasacyclo-dodecanetriacetic acid ethyl ester) in the foria of a viscous oil. 9.17 g of tha resulting ester are dissolved in 200 ml of ] 5 ethanol and stirred for 20 hours at roora temperature with 30 ml of 3M sodium hydroxide solution, then adjusted to pH 6 with hydrochloric acid and stirred for 16 hours at SO"C with 6.68 g of gadolinium acetate and the solution is purified by weans of anion and cation exchangers to 2o yield 10.05 g of the title compound in the form of a tJhite powder.

Analysis s ^50 ^^2 ^8 ®13 (1057.28) C 35.22 H 4.77 Gd 29.75 N 10.60 (calc.) 35.03 4.89 29.49 10 „ <51 (found) 2 5 Example 12 N-rnethylglucamine salt of the manganese(II) complex of N-[N-(2-hydroxyethyl)-carbamoylmethyl J-1,4,7,10-tetraaza-cyclododecane-N', N ",N"'-triacetic acid 30 8.95 g (20 mmol) of N-[N-(2-hydroxyethyl )-carbamoyl~ methyl j-x,4,7 „ 10-tetraazacyclododecane-N' ,NW,NM '-triacetic acid are suspended in 30 ml of water and heated - 38 - with 1.40 g (20 mmol) of manganese(11,) oxide for 3 hours at 100*C. 3.90 g (20 mmol) of N-raethylglucarnine are then added and, after heating for a further 12 hours at 100"C, the solution is evaporated to dryness in vacuo. 13.8 g of 5 the title compound are obtained in the form of a pink powder, in. p. 140-14 3 "C.

Analysis^ C25 H^g Mn O13 (695.64) C 4 3.17 H 6.96 Mn 7.90 N 12.08 (calc.) 10 43.44 7.16 7.69 12.01 (found) Example 13 Dysprosium(III) complex of M-(morpholinocarbonylmethyl)-1,4,7,10-tetraazacvclododecane-N'eN",N"'-triacetic acid 20 g of N-(morpholinocarbonylmethy1)-Nf,N",N"f-tris-15 (ethoxycarbonylmethyl)-1,4,7,10-tetraazacyclododecane are hydrolvsed analogously to Example 2 with sodium hydroxide solution and complexed with dysprosium acetate. 16.3 g of the title compound are obtained in the form of a white powder. 20 Analysis; C20 ^32 °8 (633.01) C 37.95 H 5.10 Dy 25.67 N 11.06 (calc.) 37.71 4.92 25.81 11.32 (found) Example 14 Preparation of liposomes loaded with gadolinium-M-25 (morpholinocarbonylmethy1)-l,4,7,10-tetraazacyclo- dodecane-N ' , Na",, N" ' -triacetic acid A lipid mixture of 75 mol % egg phosphatidylcholine and 25 mol % cholesterol are prepared in the form of a dry - 39 - substance in accordance with tha procedure described in Proc. Natl. Acad. Sci. U.S.A. 7b, 4194. 500 ma thereof are dissolved in 30 ml of diethyl ether, and 3 ml of an aqueous 0.1M solution of the gadolinium complex of M-5 (morphol inocarbony lmethyl) -1a 4,7, 10-tetraaza-N*, Nsa, M'8 *- triacetic acid ara added dropwise thereto in an ultrasound bath- The ultrasound treatment, is continued for another 10 minutes and then concentration jjj vacuo is carried out. The gal-like residue is suspended in 0„125M 10 sodium chloride solution and repeatedly centrifuged at O'C and 20„000 g in order to separate non-encapsulated gadolinium complex. The suspension is then subjected to freeze-drying in multivials. Administration is affected in the form of a colloidal dispersion in 0.9 % sodium 15 chloride solution.

Example 15 Preparation of a solution of the yttrium-90 complex of the conjugate of 1,4,7,lO-tetraasacyclododecane-N,N#,N"„-N"'-tetraacetic acid with monoclonal antibodies 20 2 rag of N-( 3-dimethylaminopropyl) -H' -ethylcarbodiimide hydrochloride and then 1 ml of a solution of 0.6 mg of monoclonal antibodies (with specificity against melanoma antigen) dissolved in 0.05 molar sodium hydrogen carbonate buffer (pH 7.8) are added to a suspension of 4 mg of 2 5 1,4,7,10-tetraazacyclododecane-N,N',N",M"'-tetraacetic acid in 1 ml of water. The whole is stirred for 2 hours at room temperature and dialvsed against a 0.3M sodium phosphate buffer. 1 ml of an vttrium-90 solution in acetate buffer pH 6 (prepared in accordance with Int. J. 30 Appl. Radiat. Xsot., Vol 36 id="p-1985" id="p-1985" id="p-1985"

[1985], p. 803) is then added and the whole is incubatad for 24 hours at room temperature. The solution is passed over a Sephadex G 25 column and the radioactive protein fraction is sterile-filtered and introduced into multivialsA storable dry prspara- -.4 0 - t ion is obtained by lyophi1isation.

Example 16 Indium(IXI) complex of N~(morpholinocarbonylmethyl)-1,4,7,10-tetraazacyclododecane-M', N"' ,N"' -triacetic acid 5 The procedure is analogous to Example 9 and complexing is effected with radioactive 1 indiurn chloride. In order to check whether the metal ions have been completely bound in the form of a chelate, a solution of the title compound is investigated by thin-layer chromatography on 10 silica gel plates in the system methanol/water (2:1).

Non-chelated metal ions are recognised as a radioactive zone at the starting spot. If necessary, the chelation is completed by adding more N-(morpholinocarbonylmethyl)-N' ,N" , N" f'-tris (ethoxycarbonylmethyl) -1, 4, 7,10-tetraaza-15 cyclododecane and subsequent ester cleavage.

The gadolinium-153 complex of N-(morpholinocarbonylmethy 1 )-1,4,7,10-tetraazacyclododecane-N',H",N"'-triacetic acid is obtained in the same manner.

Claims (5)

1. -41™ Patent Claims 1. 1,4,7, I0-tetraa2acyclododecane derivatives of the general formula I j^ooc (I) cooa1 5 wherein each R1, independently of the others, represents hydrogen or a metal ion equivalent, represents hydrogen or a methyl or ethyl group, represents a linear or branched, saturated or 10 unsaturated alkyl group that has up to 16 carbon atoms and that is substituted by from 1 to 5 hydroxy or C^-C^-alkoxy groups, -CH2-X-V in which X represents carbonyl, a linear or branched alkylene group having from 1 to 10 15 carbon atoms that is optionally substituted by from 1 to 5 hydroxy or Ci-C^-alkoxy groups, or a linear or branched alkylene group that is interrupted by oxygen atoms and has from 2 to 23 carbon atoms, V represents -N „ in which each of R3 and R4, 2 0 independently of the other, represents hydrogen, a linear or branched alkyl group that has up to 16 carbon atoms and that is optionally substituted by from 1 to 5 hydroxy or Ci-C4-alkoxy groups, or R3 and R* together with the nitrogen atom represent a 25 pyrrolidine, piperidinemorpholine or piperazine ring, or or JJ3 represents a second macrocycle of the formula I' - 42 - that is linked via an alkylene chain that has from 2 to 20 carbon atoms and optionally carries carbonyl groups at its ends and is optionally interrupted by 5 from 1 to 4 oxygen atoms or substituted by frora 1 to 5 hydroxy . C^-C^-alkoxy or carboxy-Cj -C^,-a 1 ky 1 groups f or represents b or ch^-cob*, b representing a macromolecule or bioraolecule that accumulates to an 10 especially great extent in the organ or organ part to be investigated or in the tumour, and functional groups present in the molecule are, if desired, conjugated with biomoleculest, and the salts thereof with organic or inorganic bases or amino acids or 15 with inorganic or organic acids.

2. Compounds according to claim 1f characterised in that each R1 represents a hydrogen atom.

3. Compounds according to claim 1,, characterised in that at least two of the substituents r1 are metal ion 20 equivalents of at least one metal of atomic number 21 to 29, 4 2P 44 or 57 to 70. 4s Compounds according to claim 1, characterised in that at least two of the substituents R~ are metal ion equivalents of at least one radionuclide of an element of 25 atomic number 21, 29, 31,, 32, 38,, 3943,, 49„ 64, 70 or 77. 5. N-[ 2,3-dihydroxy-M-methylpropylcarbaraoylmethylJ - - 43 - 1,4,7,10-tetraazacyclododecane-N',N",N"'-triacetic acid, N-[N-ethylcarbamoylmethyl]-l,4,7,10-tetraazacyclo-dodecane-N' ,N'6 ,N" '-triacetic acid, N-(2,3 -d i hydroxy-1-propyl)-1,4,7,10-tetraazacyclo-5 dodecane-N' -triacetic acid, N-( 2-anainoethyl )-i,4,7 , lO-tetraazacyclododecane-M",M8fl, -N""-triacetic acid, N-(2-hydroxyethyl)-1,4,7,lo-tetraazacyclododecane- N£? , N15 , N18'-triacetic acid„ 10 N-[N-(2-hydroxyethyl)-carbamoylmethyl]-1,4,7,10-tetraa2a- cyclododecane-M*,NH,N"'-triacetic acid, N-( snorpholinocarbonyInethy 1) -1,4 , 7,10-tetraa2acyclo-dodecane-M' ,N" ,NM •' -triacetic acid, N- (morphol inocarbonylraethyl) -1, 4 ,7,10-tetraazacyclo-15 dodecane-M' ,H" , H" '-tris-( 2-methylacetic acid), and the complexes of those compounds with paramagnetic metals according to claim 2 or radioisotopes according to claim

4. 6. 1,1'-(1,3-propylene)-bis(1,4,7,10-tetraazacvclo-20 dodecane-4,7,10-triacetic acid), <*> succinyl-bis(1,4 s 7,10-tetraazacyclododecane-4,7,10-triacetic acid)e « 1 , 3.- (2-hydroxy-l, 3-propy lene) -bis (1, 4 „ 7 , lO-tetraasa-cyclododecane-4 ,7,10-triacetic acid) t, and the complexes of those compounds with paramagnetic metals according to claim 3 or radioisotopes according to claim 4. 7. Gadolinium, manganese and dysprosium complexes of the coraplexing agents mentioned in claims 5 and 6. 8. Pharmaceutical agents containing at least one metal complex of the general formula I in which at least two of the substituents R1 represent a metal ion equivalent,, optionally in the form of liposomes and optionally with the adjuvants customarily used in galenical pharmacy. S „ Use of the compounds according to claim 2 as complexing agents. 10. Use of at least one metal complex of the general formula I in which at least two of the substituents R~ represent a metal ion equivalent for the preparation of media for NMR diagnostics X-ray diagnostics e ultrasound diagnostics or radiodiagnostics and radiotherapy., 11- Process for the preparation of 1,4,7,10-tetraaza-cyclododecane derivatives according to claims 1 to 3 f characterised in that, in a manner known per se, in compounds of the general formula XI (II) wherein is as defined above, R2has the meaning given for R2f tout is not to repres- % »— 4 j — ent a rnacromolecule or bionolecule B or the group CH2~C0.B< and Z represents carboxy-protecting groups,, tha protecting groups Z are split off and the resulting 5 acids (!R^ in the general formula I represents hydrogen) are,, if desired,, a). reacted in a manner known oer with at least one metal oxide or metal salt of an elenent of atomic number 21 to 29, 31, 32, 38, 39,, 42 to 44e 49, 57 10 to 70 or 77 and then, if desired„ acidic hydrogen atoms present are converted with inorganic and/or organic bases or amino acids and basic groups present are converted with inorganic or organic acids into physiologically tolerable salts, 15 or b) reacted in. a wanner known per se with at least one metal oxide or metal salt of an element of atomic number 21 to 29, 31, 32, 38„ 39,. 42 to 44, 49, 57 to 70 or 77 and then the resulting laetal complexes 20 are linked in a manner known oer se via functional groups contained in the molecule or to R2 or to the CO group contained in R2 a rnacromolecule and, if desired, acidic hydrogen atoms present are converted with inorganic and/or organic bases or amino acids 2 5 and basic groups present are converted with inor ganic or organic acids into physiologically tolerable salts, or c) linked 1 :n a manner known per se via the functional 30 groups contained in the raolecule or to R2 or to the CO group contained in R2 a snacroiaolecule and then reacted,, in a manner known oer se.. with at least one metal oxide or metal salt of an elenent of atomic number 21 to 29, 31, 32, 38, 39,, 42 to 44, 49, 57 to 35 70 or 77 and then, if dasired,, acidic hydrogen - 46 - atoms present are converted with inorganic and/or organic bases or amino acids and basic groups present are converted with inorganic or organic acids into physiologically tolerable salts. 5 12. Process for the preparation of the pharmaceutical agents according to claim 8, characterised in that the coasplex compound dissolved or suspended in water or physiological saline solution is made iff*to a for® suitable for enteral or parenteral administration, 10 optionally together with adjuvants customarily used in galenical pharmacy. 13- A compound substantially as hereinbefore described with reference to the Examples. 14, A pharmaceutical agent substantially as hereinbefore 15 described with reference to the Examples. 1

5. a use substantially as hereinbefore described with reference to the Examples. 18. A process substantially as hereinbefore described %» it h reference to the Examples. Dated this 28th day of July 1987 CRUICKSHANK & CO Agents for the Applicants 1 , Holies Street Dublin 2.