HK1050898B

HK1050898B - Intermediate used for manufacturing cascade polymer complexes

Info

Publication number: HK1050898B
Application number: HK03103098.2A
Authority: HK
Inventors: 黑里贝特‧施米特-维利希; 约翰尼斯‧普拉茨克; 贝恩德‧拉迪谢尔; 安德烈亚斯‧米勒; 托马斯‧弗伦策尔
Original assignee: 舍林公开股份有限公司
Priority date: 1995-07-04
Filing date: 2003-04-30
Publication date: 2005-03-04

Description

Intermediate product for preparing cascade polymerization complex used as contrast agent

Technical Field

The invention relates to intermediates (I' A) for the preparation of cascade polymeric complexes for use as contrast agents.

Background

Contrast agents used in modern clinical imaging techniques nuclear spin tomography (MRI) and Computed Tomography (CT) [ Magnevist^(R)、Pro Hance^(R)、Ulrtavist^(R)And Omniscan^(R)]Dispersed throughout the extracellular space of the body (intravascular space and interstitial space). The dispersion space comprises about 20% of the body volume.

Because of the very specific location with respect to the locally dispersed space, clinically extracellular MRI contrast agents were first successfully used in the diagnosis of brain and spinal cord diseases. In the brain and spinal cord, extracellular contrast agents in normal tissues cannot leave the intravascular space due to the blood-brain barrier. In the pathological process of Blood-Brain Barrier rupture (e.g., malignant tumors, inflammation, demyelinating diseases, etc.), areas of increased vascular Permeability for these extracellular Contrast agents are formed in the Brain (Schmiedl et al, MRI of Blood-Brain Barrier Permeability in astrocytomas: Application of Small and Large Molecular Weight Contrast agents (MRI of Blood-Brain Barrier Permeability in biochemical: Application of Small and Large Molecular Weight Contrast media), Magn.Reson.Med.22: 288, 1991). Using vascular permeability abnormalities, tissue with high contrast intensity relative to normal tissue is identified as diseased tissue.

The effect on Radiographic imaging to Estimate transit time and Blood Volume (First-Pass Entry of non-ionic Contrast Agent into the myocardial Extravascular space), Circulation 84: 2071, 1991. The concentration of the contrast agent is therefore no longer dependent on the vascular permeability but only on the size of the extracellular space in the respective tissue. With such contrast agents it is not possible to distinguish the blood vessel from the interstitial space around it.

Particularly for angiography, a contrast agent dispersed only in blood vessels is required. With the aid of nuclear spin tomography, the use of such contrast agents which remain in the blood makes it possible to divide tissue with sufficient blood flow and tissue with insufficient blood flow, and can thus be used for the diagnosis of ischemia. If an angiographic agent is used, the infarcted tissue can also be demarcated from the surrounding normal or ischemic tissue based on its ischemia. This is important if, for example, it can be used to distinguish between myocardial infarction and ischemia.

Most patients suspected of having cardiovascular disease today (the most mortality disease in the western developed countries) have to undergo invasive diagnostic tests. In contemporary angiography, diagnostic radiology with iodine-containing contrast agents is frequently used. These checks have various drawbacks: there is a risk of exposure to radioactivity, application difficulties and strain, and these disadvantages are further compounded by the much higher concentrations of iodine-containing contrast agents used in NMR contrast agents.

Thus, there is a need for NMR contrast agents (contrast agents retained in blood) that can label vascular spaces. These compounds are distinguished by good compatibility and high efficacy (significantly increased MRI signal intensity).

Complexes bonded to macromolecules or biomolecules have been used to date to address at least some of this problem, but their success has been limited to a limited extent.

As in european patent applications No. 0088695 and No. 0150844, the number of paramagnetic centers in the complexes described is not sufficient to achieve satisfactory contrast results.

If the number of metal ions required is increased by the repeated incorporation of complex units into the biomacromolecule, unacceptable damage can occur to the affinity and/or specificity of the biomolecule [ J.Nucl. Med.24, 1158(1983) ].

Generally macromolecules may be suitable as contrast agents for angiography. However, 24 hours after intravenous injection of mice, albumin-GdDTPA (Radiology 1987; 162: 205) was present at approximately 30% of the dose, as in lung tissue. In addition, only 20% of the dose was eliminated in 24 hours.

The macromolecule polylysine-GdDTPA (European patent application publication No. 0233619) has also proven to be suitable as a contrast agent for retention in blood. But due to limitations in the method of manufacture, the compound comprises a mixture of molecules of different sizes. Glomerular filters were found to be excreted unchanged in the mouse excretion test. Due to limitations of the synthetic method, polylysine-GdDTPA may also contain macromolecules that are too large to pass through renal capillaries when glomeruli are filtered, and thus remain in the body.

In addition, there have been reports of contrast agents based on macromolecules of carbohydrates such as dextran (european patent application publication No. 0326226). A disadvantage of this class of compounds is that they can only carry about 5% of the signal enhancing paramagnetic cations.

The polymers described in European patent application No. 0430863 and the polymers described in German patent application laid-open specification DE 4344460 have a step towards contrast agents which are retained in the blood, since they do not have the multiplicity of sizes and molecular weights of the above-mentioned polymers. Very similar to the compounds disclosed in "Magnetic Resonance in medicine" (by Wiener et al, 1994), but still further improvements are needed in view of complete elimination, compatibility, and/or efficacy.

A class of complexes which is well suited for use as NMR and X-ray diagnostic reagents without the above-mentioned disadvantages has been found to comprise a nitrogen-containing cascade polymer having ligands, at least 16 ions of an element having an atomic number of 20 to 29, 39, 42, 44 or 57 to 83, and optionally cations of inorganic and/or organic bases, amino acids or amino acid amides, and optionally acylated amino groups.

Disclosure of Invention

It is an object of the present invention to provide intermediates (I' A) for the preparation of compounds of the general formula (I):

wherein

R¹' independently of one another, represents a hydrogen atom, a metal ion of an element having an atomic number of 20 to 29, 39, 42 to 44 or 57 to 83 or an acid-protecting group,

R²represents a hydrogen atom, and is represented by,

R³' represents a group

R⁴Represents a linear or branched C₁-C₂₁An alkyl chain optionally interrupted by 1 to 7 oxygen atoms and/or optionally substituted by hydroxyl or benzyloxy,

U⁶represents a methylene group or an ethylene group,

t' represents a-C^*O, -COOH, -N ═ C ═ O or-N ═ C ═ S group, and

C^*o represents an activated carboxyl group.

Drawings

The invention will be described in detail below with reference to the attached drawing figures, wherein:

fig. 1 is a graph showing measurement of Gd (compound 1) and Dy (compound 2) concentrations in blood of a rat (n ═ 5) with renal blood vessels ligated; and

FIG. 2 is an MR angiogram of a rabbit.

Detailed Description

The cascade polymer of the present invention can be represented by the general formula (I)

A-{X-[Y-(Z-<W-K_w>_z)_y]_x}_a (I)

Wherein

A represents a nitrogen-containing cascade parent nucleus with a basic branching number a,

x and Y are independent of each other and represent a direct bond or a cascade of elongated units having a branching number X or Y,

z and W are independent of each other and represent a cascade of elongated units having a branching number Z or W,

k represents a group containing a complexing agent,

a represents a number of 2 to 12 as a representative value,

x, t, z and w are independent of each other, and the representative value is a number of 1 to 4,

provided that at least two of the elongation units in cascade are different and

16. ltoreq. a.xyzw. ltoreq.64.

The following structure applies to cascade core a:

a nitrogen atom,

or

Wherein

m and n are numbers having a value of 1 to 10,

p is a number having a value of 1 to 10,

U¹represents Q¹Or E, the content of the first and second polymers,

U²represents Q²Or E, wherein

E denotes a group

Wherein

o is a number having a value of 1 to 6,

Q¹represents a hydrogen atom or Q²

Q²Represents a direct bond with the amino acid sequence of the amino acid sequence,

m represents C₁-C₁₀An alkylene chain optionally interrupted by 1 to 3 oxygen atoms and/or optionally substituted with 1 to 2 oxy groups,

r DEG represents branched or unbranched C₁-C₁₀Alkyl, nitro, amino, carboxyl or

Wherein Q²Is equal to the number of basic branches a.

The nitrogen atom is the simplest cascade core, with its three bonds (basic branching number a ═ 3) being replaced in the first "inner layer" (stage 1) by three cascade extension units X or Y (if X represents a direct bond) or Z (if X and Y each represent a direct bond), in other words: basic Cascade Starter Ammonia A (H)_a＝NH₃Is substituted with three cascaded elongation units X or Y or Z. At this point Q in cascaded core A²Is equal to the number of basic branches a.

Cascaded elongated units X, Y, Z and W comprising-NQ¹Q²Group, wherein Q¹Refers to a hydrogen atom or Q²And Q²Refers to a direct bond. Q contained in each cascade of elongate elements (e.g. X)²Is equal to the number of branches of the unit (e.g., X in X). The product of all branch numbers a.x.y.z.w represents the number of ligands K bonded in the cascade polymer. The polymer of the invention contains at least 16 and at most 64K groups per molecule, each group being capable of bonding to 1 and at most 3 (for divalent ions) ions, preferably to ions of an element of one of the above atomic numbers.

The last stage, i.e. the cascade of elongated units W to which the ligand K is bonded, is substituted with NH groups (-NQ)¹Q²Wherein Q is¹Denotes a hydrogen atom, and Q²Directly linked) to K, and the last-stage tandem extension unit can be used with NHQ²Radicals (e.g. by acylation) and NQ²Q²Linking groups (such as alkylation reaction).

The complexes of the cascade polymers according to the invention are of maximum 10 stages (i.e.cascade extension units X, Y and Z can also be present more than one in the molecule), but preferably are of stages 2 to 4, at least two of the cascade extension units in the molecule being different.

If it is not

The value of m is from 1 to 3, particularly preferably 1,

n has a value of 1 to 3, particularly preferably 1,

p has a value of 0 to 3, particularly preferably 1,

the value of o is 1 and the value of o,

m is-CH₂-CO or-CH₂A CO group, and

r DEG is-CH₂NU¹U²、CH₃Or NO₂And (c) a group, then the cascade core described by the above formula is the preferred cascade core a.

As more preferred cascade initiators A (H)_aHere, for example:

(in parentheses, in the case where the basic branch number a has a single or double substitution at the next stage of the construction)

Tris (aminoethyl) amine (a ═ 6 or 3);

tris (aminopropyl) amine (a ═ 6 or 3);

diethylenetriamine (a ═ 5 or 3);

triethylene tetramine (a ═ 6 or 4);

tetraethylenepentamine (a ═ 7 or 5);

1, 3, 5-tris (aminomethyl) benzene (a ═ 6 or 3);

trimesic acid triamide (a ═ 6 or 3);

1, 4, 7-triazacyclononane (a ═ 3);

1, 4, 7, 10-tetraazacyclododecane (a ═ 4);

1, 4, 7, 10, 13-pentaazacyclopentadecane (a ═ 5);

1, 4, 8, 11-tetraazacyclotetradecane (a ═ 4);

1, 4, 7, 10, 13, 16-hexaazacyclooctadecane (a ═ 6);

1, 4, 7, 13, 16, 19, 22, 25, 28-decaazacyclotriacontane (a ═ 10);

tetrakis (aminomethyl) methane (a ═ 8 or 4);

1, 1, 1-tris (aminomethyl) ethane (a ═ 6 or 3);

tris (aminopropyl) -nitromethane (a ═ 6 or 3);

2, 4, 6-triamino-1, 3, 5-triazine (a ═ 6 or 3);

1, 3, 5, 7-adamantanetetracarboxylic acid amide (a ═ 8 or 4);

3, 3 ', 5, 5', -diphenyl ether-tetracarboxylic acid amide (a ═ 8 or 4);

1, 2-bis [ phenoxyethane ] -3 ', 3 ", 5', 5" -tetracarboxylic acid amide (a ═ 8 or 4);

1, 4, 7, 10, 13, 16, 21, 24-octaazabicyclo [8.8.8] hexacosane (a ═ 6).

It is noted that the definition of cascade core a and thus the distinction between cascade core and first cascade extension unit is only of a purely formal sense and is therefore not relevant for the actual synthesis of the desired cascade polymer. Thus, a tris (aminoethyl) amine as used in example 4 can be seen as the cascade core a itself (first denoted as a in contrast to the general formula, where m ═ n ═ p ═ 1, U¹O is 1, and U¹＝U²＝Q²) However, the nitrogen atom can also be regarded as a cascade nucleus (cascade nucleus A) having three cascade extension units in the first stage

(definition of comparison to E).

With E

Or

Defining cascaded elongated elements X, Y, Z and W respectively,

wherein

U¹Represents Q¹Or E

U²Represents Q²Or E, wherein

E denotes a group

Wherein

o is a number having a value of 1 to 6,

Q¹represents a hydrogen atom or Q²，

U³represents C₁-C₂₀Alkylene chain which may optionally be interrupted by 1 to 10 oxygen atoms and/or 1 to 2-N (CO)_q-R₂1 to 2 phenylene radicals and/or 1 to 2 phenoxy radicals and/or optionally substituted by 1 to 2 oxy, thio, carboxyl, C₁-C₅Alkylcarboxyl radical, C₁-C₅Alkoxy, hydroxy, C₁-C₅Alkyl is substituted, wherein

q is a number having a value of 0 or 1, and

R²represents a hydrogen atom, a methyl or ethyl group, optionally substituted by 1 to 2 hydroxyl groups or 1 carboxyl group,

l represents a hydrogen atom or group

V represents methine

If U is simultaneously present⁴Is a direct bond or a group M, and U⁵With U³One kind of meaning or

V represents a group

If U is simultaneously present⁴And U⁵Likewise, a direct bond or a group M.

Preferred cascading extension units X, Y, Z and W are those mentioned above for the formula, group U³represents-CO-, -COCH₂OCH₂CO-，-COCH₂-，-CH₂CH₂-，-CONHC₆H₄-，-COCH₂CH₂CO-，-COCH₂-CH₂CH₂CO-, or-COCH₂CH₂CH₂CH₂CO-，

Group U⁴Represents a direct bond or-CH₂CO-，

Group U⁵Represents a direct bond or- (CH)₂)₄-，-CH₂CO-，-CH(COOH)-，-CH₂OCH₂CH₂-，-CH₂C₆H₄-, or-CH₂-C₆H₄OCH₂CH₂-，

The radical E represents a radical

The following examples of cascaded elongated units X, Y, Z and W may be taken in:

-CH₂CH₂NH-；-CH₂CH₂N＜；

-COCH(NH-)(CH₂)₄NH-；-COCH(N＜)(CH₂)₄N＜；

-COCH₂OCH₂CON(CH₂CH₂NH-)₂；-COCH₂OCH₂CON(CH₂CH₂N＜)₂；

-COCH₂N(CH₂CH₂NH-)₂；-COCH₂N(CH₂CH₂N＜)₂；

-COCH₂NH-；-COCH₂N＜；

-COCH₂CH₂CON(CH₂CH₂NH-)₂；-COCH₂CH₂CON(CH₂CH₂N＜)₂；

-COCH₂OCH₂CONH-C₆H₄-CH[CH₂CON(CH₂CH₂NH-)₂]₂；

-COCH₂OCH₂CONH-C₆H₄-CH[CH₂CON(CH₂CH₂N＜)₂]₂；

-COCH₂CH₂CO-NH-C₆H₄-CH[CH₂CON(CH₂CH₂NH-)₂]₂；

-COCH₂CH₂CO-NH-C₆H₄-CH[CH₂CON(CH₂CH₂N＜)₂]₂；

-CONH-C₆H₄-CH[CH₂CON(CH₂CH₂NH-)₂]₂；

-CONH-C₆H₄-CH[CH₂CON(CH₂CH₂N＜)₂]₂；

-COCH(NH-)CH(COOH)NH-；-COCH(N＜)CH(COOH)N＜；

or

The ligand K is represented by formula IA or IB:

wherein:

R¹each independently, represents a hydrogen atom or a metal ion of an element having an atomic number of 20-29, 39, 42-44 or 57-83,

R²represents a hydrogen atom, a methyl or ethyl group, optionally substituted with 1-2 hydroxyl groups or 1 carboxyl group,

R³represents a group

R⁴Represents straight-chain, branched, saturated or unsaturated C₁-C₃₀An alkyl chain optionally interrupted by 1 to 10 oxygen atoms, 1 phenylene group, 1 phenoxy group and/or optionally substituted by 1 to 5 hydroxyl groups, 1 to 3 carboxyl groups, 1 phenyl group,

R⁵represents a hydrogen atom or R⁴，

U⁶Represents straight-chain, branched, saturated or unsaturated C₁-C₂₀An alkylene chain optionally containing 1-5 imino groups, 1-3 phenylene groupsOxy, 1-3 phenylene imino groups, 1-5 amide groups, 1-2 hydrazide groups, 1-5 carbonyl groups, 1-5 ethyleneoxy groups, 1 urea group, 1 thiourea group, 1-2 carboxyalkylimine groups, 1-2 ester groups; 1 to 10 oxygen atoms, 1 to 5 sulphur atoms and/or 1 to 5 nitrogen atoms and/or optionally substituted by 1 to 5 hydroxyl groups, 1 to 2 mercapto groups, 1 to 5 oxy groups, 1 to 5 sulphur groups, 1 to 3 carboxyl groups, 1 to 5 carboxyalkyl groups, 1 to 5 ester groups and/or 1 to 3 amino groups, wherein the optionally contained phenylene group may be substituted by 1 to 2 carboxyl groups, 1 to 2 sulpho groups or 1 to 2 hydroxyl groups,

t represents a-CO-alpha, -NHCO-alpha or-NHCS-alpha group, and

α represents a bonding position of a terminal nitrogen atom of the last cascade unit W.

As preferred complexing agent groups K there may be mentioned, in the above formula IA, the group U⁶C of (A)₁-C₂₀Is preferably C₁-C₁₂The alkylene chain containing the group-CH₂，-CH₂NHCO，-NHCOCH₂O，-NHCOCH₂OC₆H₄，-N(CH₂CO₂H)，-NHCOCH₂C₆H₄，-NHCSNHC₆H₄，-CH₂OC₆H₄，-CH₂CH₂O and/or by a group-COOH, -CH₂COOH.

The following groups may be mentioned as U⁶Examples of (2):

-CH₂-，-CH₂CH₂-，-CH₂CH₂CH₂-，-C₆H₄-，-C₆H₁₀-，-CH₂C₆H₅-，

-CH₂NHCOCH₂CH(CH₂CO₂H)-C₆H₄，

-CH₂NHCOCH₂OCH₂-，

-CH₂NHCOCH₂C₆H₄-，

-CH₂NHCSNH-C₆H₄-CH(CH₂COOH)CH₂-，

-CH₂OC₆H₄-N(CH₂COOH)CH₂-，

-CH₂NHCOCH₂OCH₂CH₂O)₄-C₆H₄-，

-CH₂O-C₆H₄-，

-CH₂CH₂-O-CH₂CH₂-，-CH₂CH₂-O-CH₂CH₂-O-CH₂CH₂-，

the following groups may be indicated as R⁴Examples of (2):

-CH₃，-C₆H₅，-CH₂-COOH，

-CH₂-C₆H₅，-CH₂-O-(CH₂CH₂-O-)₆CH₃，-CH₂-OH

if the agents of the invention are to be used in NMR diagnostics, the complex salt central ion must be paramagnetic. Particularly divalent or trivalent ions of the elements having atomic numbers 21-29, 42, 44, and 58-70. Suitable ions are, for example, the chromium (III), iron (II), cobalt (II), nickel (II), copper (II), praseodymium (III), neodymium (III), samarium (III) and yttrium (III) ions. Gadolinium (III), terbium (III), dysprosium (III), holmium (III), erbium (III), manganese (II) and iron (II) ions are particularly preferred because of their very strong magnetic moment.

If the agents of the invention are to be used in radiodiagnosis, the central ion is derived from an element of higher atomic number in order to be able to absorb x-rays sufficiently. For this purpose, physiologically compatible, complex salt diagnostic reagents containing a central ion of an element having an atomic number of 21 to 29, 39, 42, 44, 57 to 83 have been found to be suitable; such as lanthanum (III) ions and the lanthanide ions mentioned above.

The cascade polymer complex of the present invention contains at least 16 ions of the above-mentioned atomic number elements.

The residual acidic hydrogen atoms, i.e. hydrogen atoms not substituted by the central ion, can optionally be substituted with cations of inorganic and/or organic bases, amino acids, or amino acid amides.

Suitable inorganic cations are, for example, lithium, potassium, calcium, magnesium and, in particular, sodium. Suitable organic base cations are those of primary, secondary or tertiary amines, such as ethanolamine, diethanolamine, morpholine, reduced glucamine, N-dimethylreduced glucamine, and in particular N-methylglucamine. Suitable cations of amino acids are, for example, those of lysine, arginine, and ornithine, as well as amides of other acidic or neutral amino acids.

The compounds of the present invention having a molecular weight of 10,000-80,000D, preferably 15,000-40,000D, exhibit the desired properties described above. They contain a large number of metal ions which are bound in stable form to the complex, as required for their use.

They accumulate in areas of high vascular permeability such as tumors, they make possible perfusion of the tissue, and with them it is possible to determine the blood volume in the tissue, selectively reducing the diastolic time or the blood density, illustrating the permeability of the blood vessels. And these physiological data are obtained by using extracellular contrast agents, such as Gd-DTPA [ Magnevist ]^(R)]But not available. In this way, they can also perform the functions of modern contrast technique nuclear spin tomography scanning and electronic computer tomography scanning: more characteristic diagnoses of malignancies, early treatment surveillance when using killer cells, anti-inflammatory or vasodilatory therapy, early identification of lower perfusion regions (as in the myocardium), angiography of vascular disease, and early identification and diagnosis of (non-bacterial or infectious) inflammation.

The complexes of the cascade polymers according to the invention are also very suitable for the imaging of lymphoid systems (interstitial or intravenous administration).

With extracellular contrast agents such as Gd-DTPA [ Magnevist ]^(R)]Another advantage that must be emphasized in comparison is the higher efficacy (stronger relaxation) as a nuclear spin tomography contrast agent; this ensures that the dose required for diagnosis is greatly reduced. At the same time, the contrast agent of the invention can be formulated as an isotonic solution in the blood, thereby reducing the osmotic pressure of the body, which reflects a reduced toxicity (higher toxicity threshold) of the substance in this respect. Smaller doses and higher toxicity thresholds have led to a significant increase in the reliability of contrast agents in modern imaging techniques.

In contrast to macromolecular contrast agents based on carbohydrates such as dextran (european patent application publication No. 0326226), they generally carry only about 5% of the signal enhancing paramagnetic cations as described above, whereas complexes of the polymers of the present invention can generally carry about 20% of the paramagnetic cations. Thus, a stronger signal enhancement can be produced per mole of macromolecule of the invention, while also allowing a significant reduction in the dose required for nuclear spin tomography relative to carbohydrate-based contrast agents.

Macromolecules having a uniformly defined molecular weight can be designed and produced using the polymer complexes of the invention. It is therefore also possible, surprisingly enough, to control the size of the macromolecules so that they are large enough to leave the vascular space only slowly, but small enough to pass through the renal capillaries of size 300- & 800 *.

The complexes of the cascade polymers of the present invention are characterized by improved drainage performance, higher efficacy, better stability, and/or better compatibility compared to other prior art-mentioned polymers.

Another advantage of the present invention is the possibility of providing complexes with hydrophilic or lipophilic, macrocyclic or open-chain, low-molecular or high-molecular weight ligands. It is therefore possible to control the compatibility and pharmacokinetics of these polymeric complexes by chemical substitution methods.

For preparing the inventive complexes of cascade polymers, compounds of the general formula (I

A-{X-[Y-(Z-<W-β_w>_z)_y]_x}_a (I′)

Wherein

x and Y, independently of one another, represent a direct bond or a cascade of extension units having a branching number X or Y,

z and W, independently of one another, represent a cascade of elongated units having a branching number Z and W,

a represents a number of 2 to 12 as a representative value,

x, Y, Z and w, independently of one another, represent a number from 1 to 4, and

beta represents the bonding position of the terminal NH group of the last-stage cascade extension unit W,

provided that at least two of the cascade extension units are different and that the product of the number of branches is guaranteed

16≤a·x·y·z·w≤64

With a complex or complexing agent K ' of the formula I ' A or I ' B

Wherein

R³' represents a group

R⁵represents a hydrogen atom or R⁴

U⁶Represents straight-chain, branched, saturated or unsaturated C₁-C₂₀Alkylene optionally containing 1 to 5 imino groups, 1 to 3 phenylene groups, 1 to 3 phenoxy groups, 1 to 3 phenylene imino groups, 1 to 5 amide groups, 1 to 2 hydrazide groups, 1 to 5 carbonyl groups, 1 to 5 ethyleneoxy groups, 1 urea group, 1 thiourea group, 1 to 2 carboxyalkylimine groups, 1 to 2 ester groups; 1 to 10 oxygen atoms, 1 to 5 sulphur atoms and/or 1 to 5 nitrogen atoms and/or optionally substituted by 1 to 5 hydroxyl groups, 1 to 2 mercapto groups, 1 to 5 oxy groups, 1 to 5 sulphur groups, 1 to 3 carboxyl groups, 1 to 5 carboxyalkyl groups, 1 to 5 ester groups and/or 1 to 3 amino groups, wherein the optionally contained phenylene group may be substituted by 1 to 2 carboxyl groups, 1 to 2 sulpho groups or 1 to 2 hydroxyl groups,

t, represents a-C^*O, -COOH, -N ═ C ═ O or-N ═ C ═ S group, and

C^*o represents an activated carboxyl group, and O represents an activated carboxyl group,

with the proviso that, if K' represents a complex, at least two (in the case of divalent ions) or three (in the case of trivalent metals) substituents R¹Represents an upper partMetal ions of the elements mentioned and optionally further carboxyl groups in the form of their inorganic and/or organic bases, amino acids or amino acid amide salts, with elimination of the optional protective groups, reaction of the resulting cascade polymer, if K' represents a complexing agent, with at least one metal oxide or metal salt of an element having an atomic number of 20 to 29, 39, 42, 44 or 57 to 83 in a manner known in the art, followed by optionally total or partial substitution of the acidic hydrogens in the complexes of the cascade polymer obtained with cations of inorganic and/or organic bases, amino acids or amino acid amides, and then optionally acylation of the free amino groups still present before or after the metal complexation.

Another aspect of the invention is the novel compounds of the general formula I' A:

wherein

R¹', independently of one another, represent a hydrogen atom, a metal ion of an element having an atomic number of 20 to 29, 39, 42 to 44 or 57 to 83 or an acid protecting group.

R³' represents a group

U⁶represents straight-chain, branched, saturated or unsaturated C₁-C₂₀Alkylene, optionally containing 1-5 imino groups1-3 phenylene groups, 1-3 phenoxy groups, 1-3 phenylene imino groups, 1-5 amide groups, 1-2 hydrazide groups, 1-5 carbonyl groups, 1-5 ethyleneoxy groups, 1 urea group, 1 thiourea group, 1-2 carboximido groups, 1-2 ester groups; 1 to 10 oxygen atoms, 1 to 5 sulphur atoms and/or 1 to 5 nitrogen atoms and/or optionally substituted by 1 to 5 hydroxyl groups, 1 to 2 mercapto groups, 1 to 5 oxy groups, 1 to 5 sulphur groups, 1 to 3 carboxyl groups, 1 to 5 carboxyalkyl groups, 1 to 5 ester groups and/or 1 to 3 amino groups, wherein the optionally contained phenylene group may be substituted by 1 to 2 carboxyl groups, 1 to 2 sulpho groups or 1 to 2 hydroxyl groups,

t' represents a-C^*O, -COOH, -N ═ C ═ O or-N ═ C ═ S group, and

C^*o represents an activated carboxyl group.

They are important intermediates for the preparation of complexes of cascade polymers of the general formula I.

Activated carbonyl groups C as complexes or complexing agents K^*As examples of O, mention may be made of acid anhydrides, p-nitrophenyl esters, esters of N-hydroxysuccinimide, pentafluorophenyl esters and acid chlorides.

The addition or acylation reaction for introducing the complexing agent unit is carried out on a substrate containing the desired substituent K, optionally with a leaving group bonded thereto, or which reacts to form the desired substituent.

As examples of addition reactions, mention may be made of the reaction of isocyanates with isothiocyanates, the reaction of isocyanates preferably being carried out in aprotic solvents, such as THF, dioxane, DMF, DMSO, dichloromethane, at temperatures between 0 and 100 ℃ and preferably between 0 and 50 ℃ with the optional addition of organic bases such as triethylamine, pyridine, lutidine, N-ethyldiisopropylamine, N-methylmorpholine. The reaction of isothiocyanates is generally carried out in a solvent such as water or lower alcohols such as methanol, ethanol, isopropanol and mixtures thereof, DMF or mixtures of DMF and water at a temperature of between 0 and 100 ℃, preferably between 0 and 50 ℃, optionally with the addition of an organic or inorganic base such as triethylamine, pyridine, lutidine, N-ethyldiisopropylamine, N-methylmorpholine, or alkaline earth metal hydroxides, alkali metal hydroxides such as lithium hydroxide, sodium, potassium, calcium, or carbonates thereof such as magnesium carbonate.

As examples of acylation reactions, there may be mentioned reactions of free carboxyl groups which are well known to those skilled in the art [ e.g.J.P.Greenstein, M.Winitz, Chemistry of the Amino Acids, John Wiley & Sons, N.Y. (1961), page 943-945 ]. It has proven advantageous to convert the carboxyl group into an activated form such as an anhydride, an activated ester or an acid chloride prior to the acylation reaction [ e.g., e.gross, j.meienhofer, the peptides, Academic Press, n.y. (1979), vol.1, pp.65-314; albertson, org.fact.12, 157 (1962).

For the reaction with the active ester, documents known to those skilled in the art can be cited [ e.g., Houben-Weyl, Methoden organic Chemie [ methods of organic chemistry ], Georg Thieme Verlag, Stuttgart, Volume E5(1985), 633 ]. The reaction may be carried out under the anhydride reaction conditions described above. But aprotic solvents such as dichloromethane, chloroform may also be used.

For the reaction of the acid chlorides, it is only possible to work with aprotic solvents such as dichloromethane, chloroform, toluene or THF, at temperatures between-20 and 50 ℃ and preferably between 0 and 30 ℃. Furthermore, documents known to the person skilled in the art are cited [ e.g.Houben-Weyl, Methoden der Organischen Chemie, Georg-Thieme-Verlag, Stuttgart, (1974), Volume 15/2, page 355-364 ].

If R is¹' represents an acid protecting group, and lower alkyl, aryl and aralkyl groups such as methyl, ethyl, propyl, butyl, phenyl, benzyl, benzhydryl, trityl, di (p-nitrophenyl) methyl, and trialkylsilyl groups are applicable.

Optionally, deprotection is carried out according to methods well known to those skilled in the art, such as hydrolysis, hydrogenolysis, base saponification of the ester with aqueous alcoholic base solution at temperatures between 0 ℃ and 50 ℃ or removal of tert-butyl ester by means of trifluoroacetic acid.

The terminal amino group, optionally not completely acylated by the ligand or complex, may optionally be converted to an amide or a hemiamide. Reaction with acetic anhydride, succinic anhydride or diglycolic anhydride may be mentioned as examples.

The desired metal ions can be introduced as disclosed in German patent application publication No. 3401052, by dissolving or suspending metal oxides or metal salts of elements having atomic numbers 20-29, 42, 44, 57-83 (e.g.nitrates, acetates, carbonates, hydrochlorides or sulfates) in water and/or lower alcohols (e.g.methanol, ethanol or isopropanol) and reacting with an equivalent amount of a solution or suspension of the complexing agent, optionally followed by substitution of the acidic hydrogen on the acidic group with cations of inorganic and/or organic bases, amino acids or amino acid amides.

The introduction of the desired metal ion can be carried out during the step of forming the complexing agent I 'A or I' B, i.e.before the coupling to the cascade polymer, or after the attachment of the unmetallized ligand I 'A or I' B.

In this case, neutralization is carried out with the aid of inorganic bases (e.g. hydroxides, carbonates or bicarbonates) such as sodium, potassium, lithium, magnesium or calcium and/or organic bases, such as primary, secondary and tertiary amines, such as ethanolamine, morpholine, reduced glucamine, N-methyl-and N, N-dimethylreduced glucamines, and also basic amino acids such as lysine, arginine and ornithine or amides of amino acids which are initially neutral or acidic, such as hippuric acid, glycine acetamide.

To prepare the neutral complexes, for example, a sufficient amount of the desired base can be added to an aqueous solution or suspension of the acid complex salt and a point of neutralization is reached. The resulting solution may then be evaporated to dryness under vacuum. Frequently, it is advantageous to add to the solution a water-miscible solvent, such as lower alcohols (methanol, ethanol, isopropanol and others), lower ketones (acetone and others), polar ethers (tetrahydrofuran, dioxane, 1, 2-dimethoxyethane and others), to precipitate the neutral salt, thereby obtaining crystals which are easy to isolate and purify. It has proven to be particularly advantageous to add the desired base as early as possible during the compounding of the reaction mixture, thus simplifying the work-up procedure.

If the acidic complex contains several free acidic groups, it is often suitable to prepare neutral mixed salts containing inorganic and organic cations as counterions.

Neutral mixed salts can be prepared, for example, by reacting the complexing agent in aqueous suspension or solution with the oxide or salt of the central ion-forming element and an organic base in an amount half that required for neutralization, and then separating the complex obtained, optionally purifying it and then adding the inorganic base in an amount required for complete neutralization. The order of base addition may also be reversed.

Optionally adding acid or base to adjust pH to 6 to 8, preferably about 7, and preferably using a membrane with appropriate pore size (e.g., Amicon)^(R)XM30、Amicon^(R)YM10、Amicon^(R)YM3) ultrafiltration or on Sephadex as applicable^(R)The resulting complex of the cascade polymer was purified by gel filtration on a gel.

For neutral complexes, it is often advantageous to use anion exchangers such as IRA 67 (OH)^-Type) and optionally adding a cation exchanger such as IRC 50 (H)⁺Type) as the ionic component of the polymeric complex.

The preparation of the cascade polymers having terminal amino groups, which are usually required for the coupling of the complexing agent K' (or other corresponding metal-containing complexes), starts from nitrogen-containing cascade starters A (H) which can be prepared by commercially available methods or according to or analogously to literature methods_a. March, Advanced Organic Chemistry, 3rd ed.; john Wiley&Sons，(1985)，364-381]X, Y, Z and W stages are introduced by acylation or alkylation of a protected amine of the desired structure with functional groups that can bond to the cascade nucleus, such as a carboxylic acid group, an isocyanate group, an isothiocyanate group or an activated carboxylic acid group (e.g., an anhydride group, an activated ester acid chloride group) or a halide group (e.g., a chloride group, a bromide group, an iodide group), an aziridine group, a mesylate group, a p-toluenesulfonate group or other leaving groups well known to those skilled in the art.

It is emphasized that the distinction between cascade core a and cascade elongated units is purely formal. Without formal cascade initiator A (H)_aInstead, the nitrogen atom which is defined to form part of the cascade nucleus is first introduced together with the first stage, which is synthetically advantageous. Thus, as in the synthesis of the compounds described in example 1b), a more advantageous solution is not to alkylate the cascade of the nuclear trimesic triamide (six-fold) on the form as benzyloxycarbonylaziridine, but with bis [2- (benzyloxycarbonylamino) -ethyl ] -ethyl]Reaction of the amine with trimesic acid triacyl chloride (threefold).

As amino-protecting groups, mention may be made of benzyloxycarbonyl, t-butoxycarbonylacyl, trifluoroacetyl, fluoromethoxycarbonyl, benzyl and formyl [ Th.W.Greene, P.G.M.Wuts, Protective Group in Organic Syntheses, 2nd ed., John Wiley and sons (1991), PP.309-385, which are well known to the person skilled in the art]. After removal of these protecting groups according to literature procedures, the next desired stage can be introduced into the molecule. In addition to the synthesis of stages in two stages (alkylation or acylation and deprotection) per stage of the cascade, it is also possible to introduce two stages simultaneously, e.g. X- [ Y ], using only two reactions]_xOr several stages, e.g. X- [ Y- (Z)_y]_x. These multi-stage units are synthesized by alkylating or acylating an unprotected amine having the desired structure of the cascade of elongated units ("cascade of elongated amines") with an amine group as a protected second cascade of elongated amines.

As a cascade initiator of the general formula A (H)_aThe compounds of (A) are commercially available or can be prepared by literature or analogous methods [ e.g., Houben-Weyl, Methoden der org. Chemie, Georg-Thieme-Verlag, Stuttgart (1957), volume 11/1; micheloni et al, inorg. chem. (1985), 24, 3702; atkins et al, org.Synth, vol.58 (1978), 86-98; the Chemistry of heterocyclic Compounds: bradshaw et al, Aza-Crown-Macrocycles, John Wiley& Sons，N.Y.(1993)]And (4) preparation. The following compounds may be cited as examples:

tris (aminoethyl) amine [ e.g., Fluka Chemistry [ Fluka Chemistry ] AG, Switzerland; Aldrich-Chemistry [ Aldrich Chemistry ], Germany ];

tris (aminopropyl) amine [ e.g., c.woemer et al, angelw.chem. [ Applied Chem ].]Int.Ed.Engl.(1993). 32，1306]；

Diethylenetriamine [ e.g., Fluka; aldrich ];

triethylenetetramine [ e.g., Fluka; aldrich ];

tetraethylenepentamine [ e.g., Fluka; aldrich ];

1, 3, 5-tris (aminomethyl) benzene [ e.g., T.M.Garrett et al, J.Am.chem.Soc. (1991),113，2965]；

trimesic acid triamide [ e.g., h.kurihara; jpn. kokai Tokyo JP 04077481; CA117，162453]；

1, 4, 7-triazacyclononane [ e.g., Fluka; aldrich ];

1, 4, 7, 10, 13-pentaazacyclopentadecane [ e.g., K.W.Aston, European patent application 0524161, CA120，44580]；

1, 4, 7, 10-tetraazacyclododecane [ e.g., Aldrich ];

1, 4, 8, 11-tetraazacyclotetradecane [ e.g., Fluka; aldrich ];

1, 4, 7, 10, 13, 16, 19, 22, 25, 28-decaazacyclotriacontane [ e.g., a. andres et al, j.chem soc. dalton Trans. (1993), 3507 ];

1, 1, 1-tris (aminomethyl) ethane [ e.g., R.J.Geue et al, Aust.J.chem. (1983),36，927]；

tris (aminopropyl) -nitromethane [ e.g., g.r.newkome et al, angelw.chem.1031205(1991), analogous to R.C. Larock, Comprehensive Organic Transformation, VCH Publishers, N.Y. (1989), 419-420]；

1, 3, 5, 7-adamantanetetraCarboxylic acid amides [ e.g., H.Stetter et al, Tetr.Lett.1967，1841]

1, 2-bis [ phenoxyethane]-3 ', 3 ", 5', 5" -tetracarboxylic acid amides [ e.g., j.p. collman et al; am chem soc (1988),1103477-86, analogously to the description of example 1b]；

1, 4, 7, 10, 13, 16, 21, 24-octaazabicyclo [8.8.8] hexacosane [ e.g., p.h.smith et al, j.org.chem. (1993), 58, 7939 ].

The cascade of elongated amines containing the desired functional groups of each of the above synthetic stages was prepared according to or analogously to methods known in the experimental part or from the literature.

Mention may be made, by way of example, of:

na, N xi-di-benzyloxycarbonyl-lysine-p-nitrophenyl ester [ see description of example 1c) ];

HOOC-CH₂OCH₂CO-N(CH₂CH₂NH-CO-O-CH₂C₆H₅)₂；

HOOC-CH₂N(CH₂CH₂NH-CO-O-CH₂C₆H₅)₂；

HOOC-CH₂CH₂CO-N(CH₂CH₂NH-COCF₃)₂[ prepared as described in example 3a) starting with bis (trifluoroacetamidoethyl) amine instead of bis (benzyloxycarbonylaminoethyl) amine and succinic anhydride instead of diethylene glycol anhydride]；

HOOC-CH₂OCH₂CONH-C₆H₄-CH[CH₂CON(CH₂CH₂NH-CO-O-CH₂C₆H₅)₂]₂[ preparation analogously to example 3a)]；

O＝C＝N-C₆H₄-CH[CH₂CON(CH₂CH₂NH-CO-O-CH₂C₆H₅)₂]₂

N-benzyloxycarbonyl-aziridine, prepared according to M.Zinic et al, J.chem.Soc, PerkinTrans 1, 21-26(1993),

n-benzyloxycarbonyl-lysine is commercially available from, for example, Bachem, California

According to the general formula of C.J.Cavallito et al, J.Amer.chem.Soc.1943,652140 starting from N-CO-OCH2C6H5- (2-bromoethyl) amine instead of benzyl chloride [ A.R.Jacobson et al, J.Med.chem (1991), 34, 2816]。

The preparation of the complexes or complexing agents of the formulae I 'A and I' B is carried out analogously to the experimental part or methods known from the literature (cf. for example, European patent application Nos. 0512661, 0430863, 0255471 and 0565930).

Thus, for the preparation of compounds of formula I ' A, e.g. in which the T ' group is used as precursor of the functional group T ', it can be said that the above-mentioned process can be used without affecting the acid protecting group R¹' in the case of protected acid functional Groups which are converted to free acid functional Groups, or means a protected acid functional group which can be converted to free acid functional Groups by methods known in the literature [ Th, W.Green, P.G.M.Wuts, Protective Groups in Organic Synthesis, 2nd Edition, John Wiley& Sons(1991)，PP.309-385]The deprotection group may then be converted into an isocyanate or isothiocyanate [ Methoden der org. Chemie (Houben-Weyl), E4, PP.742-749, 837-]Protected amine group of (a). These compounds can be prepared by monoalkylation of a cycloalkene (cyclo) with the appropriate alpha-haloamide according to or analogously to the procedure described in the experimental section [ in an aprotic solvent such as chloroform]。

For the preparation of compounds of the formula I' B, it is possible, for example, to use protected acid functions as reactive carboxyl groups-C^*Precursors of O, which can be prepared by the process described above without affecting the acid protecting group R¹' are converted to free acid functional groups and can be activated by methods known in the literature and described above. These compounds can be prepared according to or analogously to the methods described in the experimental section, for example by reacting an amino acid derivative of the formula II with an alkylating agent of the formula III.

Wherein

R⁵' and R⁵Have the same meaning, wherein R⁵Optionally containing hydroxyl or carboxyl groups, optionally in protected form, and

V¹is straight-chain or branched C₁-C₆Alkyl, benzyl, trimethylsilyl, triisopropylsilyl, 2, 2, 2-trifluoroethoxy or 2, 2, 2-trichloroethoxy, wherein V¹Different from V¹”，

Wherein

R¹"represents a protecting group, and

hal represents a halogen atom, for example Cl, Br or I, but preferably Cl [ see also m.a. williams, h.rapoport, j.org.chem.58，1151(1993)]。

Preferred amino acid derivatives are natural alpha-amino acids.

The reaction of compound (II) and compound (III) is preferably carried out in a buffered alkylation reaction system, wherein an aqueous phosphate solution is used as buffer. The reaction is carried out at a pH of 7 to 9, preferably at a pH of 8. The concentration of buffer may be between 0.1 and 2.5M, but it is preferred to use 2M phosphate buffer. The alkylation temperature may be between 0 and 50 c, with room temperature being preferred.

The reaction is carried out in a polar solution, such as acetonitrile, tetrahydrofuran, 1, 4-dioxane or 1, 2-dimethoxyethane, preferably using acetonitrile.

The agents of the invention can also be prepared according to methods known in the art, using the complexes of the invention, optionally with the addition of additives customary for plant preparations, suspended or dissolved in an aqueous medium and the suspension or solution optionally sterilized. Suitable additives are, for example, physiologically harmless buffers (e.g.trimethylammonio methane), and complexing or weakly complexing additives (e.g.complexes of diethylenetriaminepentaacetic acid or of the corresponding Ca-cascade polymers) or, if desired, electrolytes, such as sodium chloride or, if desired, antioxidants, such as ascorbic acid.

If it is desired to use suspensions or solutions of the agents of the invention in water or physiological saline for gastrointestinal administration or for other purposes, they are mixed with one or more adjuvants conventionally used in galenical preparations [ e.g. methylcellulose, lactose, mannitol]And/or surfactants [ e.g. lecithin, Tween^(R)、Myrj^(R)]And/or taste-modifying flavoring agents [ e.g. essential oils]And (4) mixing.

In principle, it is also possible to prepare the medicament of the invention without isolating the complexing agent salts. However, in any event, great care is taken in carrying out the chelation reaction so that the salts and salt solutions of the present invention are substantially free of toxic, noncoordinating forms of the metal ion.

In the preparation process, this requirement can be ensured, for example, by controlled titration with the aid of a coloured indicator, such as xylenol yellow. The invention therefore also relates to a process for preparing the complexes and their salts. The final precaution is to purify the isolated salt of the complex.

Preferably, the agent of the present invention contains 1. mu. mol to 1.3mol/l of the complex salt and is usually dosed in an amount of 0.0001 to 5 mmol/kg. They are designed for gastrointestinal and parenteral administration. Complexes of the invention

1. In the form of complexes of ions of elements having atomic numbers 21-29, 39, 42, 44 and 57-83, for use in NMR and radiodiagnosis;

2. in the form of complexes of ions of elements having atomic numbers 27, 29, 31, 32, 37-39, 43, 49, 62, 64, 70, 75 and 77, for use in radiodiagnosis and radiotherapy.

The agents of the present invention meet a variety of requirements needed for contrast agents for nuclear spin tomography. Therefore, after oral or parenteral administration, they are well suited for improving images by means of nuclear spin tomography, embodying their value in information by increasing the signal intensity. In addition, they have high potency to minimize the foreign substances required by the body, and good compatibility to maintain the non-invasive nature of the study.

The inventive agents have good water solubility and low permeability, which makes it possible to prepare highly concentrated solutions, so that the volume loaded by the circulation system can be controlled within reasonable limits, and compared to the dilution of body fluids, i.e. the NMR diagnostic agents must have 100 to 1000 times higher water solubility than is required for NMR spectroscopy. In addition, the agents of the invention have not only high stability in vitro, but also surprisingly good stability in vivo, so that the release or exchange of ions non-covalently bonded to the complex, which are toxic per se, proceeds very slowly over the time the novel contrast agent is completely excreted in vitro.

In general, the dosage of the agent of the present invention used as an NMR diagnostic agent is in the range of 0.0001 to 5mmol/Kg, preferably 0.005 to 0.5 mmol/Kg. Application details, e.g. in H.J. Weinmann et al, am.J. of Roentgenology142619 (1984).

Can be used as organ characteristic NMR diagnostic agent with particularly low dosage (less than 1mg/Kg body weight), such as tumor diagnosis and myocardial infarction diagnosis.

Furthermore, the complexes of the invention are advantageously used as sensitizers and shifting agents for in vivo NMR spectroscopy.

The agents of the invention are also suitable as radiodiagnostic agents because of their beneficial radioactivity and the good stability of the complexes they contain. Details of their use and dosage are described, for example, in Radiotracers for medical Applications, "CRC Press, Boca Raton, Florida.

Another imaging method using radioactive isotopes is positron emission tomography, which uses positron-emitting isotopes, such as⁴³Sc、⁴⁴Sc、⁵²Fe、⁵⁵Co and⁶⁸Ga(Heiss，W.D.；Phelps，M.E.；Positron Emission Tomography of Brain，Springer Verlag Berlin，Heidelberg，New York.1983)。

it is sufficiently surprising that the compounds of the invention can also distinguish malignant from benign tumours in areas where the blood brain barrier is absent.

They are also characterized by being completely eliminated from the body and thus having good tolerability.

Since the substances of the invention are concentrated on malignant tumors (do not diffuse into normal tissue, but have high permeability to tumor vessels), they may also contribute to the radiotherapy of malignant tumors. The latter is different from the corresponding diagnosis only by the amount and kind of isotope used. The aim is to destroy tumor cells in the shortest possible range of action by means of high-energy short-wave radiation. For this purpose, the interaction of the metals contained in the complex (e.g. iron or gadolinium) is used together with ionizing radiation (e.g. x-rays) or with neutron rays. Due to this effect, the radiation dose at the spot where the metal complex is present (e.g. in a tumor) is significantly increased. The use of these metal complexes allows malignant tumor tissue to receive the same radiation dose while substantially reducing the radiation dose on normal tissue, thereby avoiding side effects that afflict the patient. The metal complex conjugates of the invention are therefore also suitable as radiosensitizers in the treatment of malignant tumors (e.g. for the Mosler effect or in neutron capture therapy). Suitable beta-emitting ions are, for example⁴⁶Sc、⁴⁷Sc、⁴⁸Sc、⁷²Ga、⁷³Ga and⁹⁰and Y. Suitable alpha-emitting ions having a short half-life are, e.g.²²¹Bi，²¹²Bi，²¹³Bi and²¹⁴bi, among them, is preferably²¹²And (4) Bi. Suitable photon-and electron-emitting ions are¹⁵⁸Gd, which can be produced by neutron capture¹⁵⁷Gd is prepared.

If the agent of the invention is designed for use in a variation of the radiation therapy proposed by R.L. Mills et al (Nature Vol.336, (1988), P.787), the central ion must be derived from a Muslim isotope such as⁵⁷Fe or¹⁵¹Eu in the presence of a catalyst.

In the in vivo administration of the therapeutic agents of the present invention, the agents may be administered with a suitable carrier such as plasma or physiological common solution and another protein such as human serum albumin. The dose in this case depends on the type of cellular disorder, the type of metal ion used and the type of imaging method.

The therapeutic agents of the present invention are administered parenterally, preferably intravenously.

Details of the application of radiotherapeutic agents are discussed, for example, in r.w. kozak et al TIBTEC, October1986, 262.

The agents of the invention are well suited as x-ray contrast agents, especially for electronic Computed Tomography (CT), where particular emphasis is placed on the symptoms of anaphylactoid reactions of known iodine-containing contrast agents that are not detectable by biochemical-pharmacological studies. They are particularly useful due to the beneficial absorption properties in the region of higher tube voltages of the digital subtraction technique.

In general, the agent of the present invention is used in an amount of 0.1 to 5mmol/Kg, preferably 0.25 to 1mmol/Kg, as an X-ray contrast agent like diatrizoate.

Details of the use of x-ray contrast agents are discussed, for example, in Barke, Rontgen Kontras tmitel [ x-ray contrast agents ], G.Thieme, Leipzig (1970) and P.Thurn, E.Bucheler "Einfuhrung inde Rontgen diagnostic unstik [ diagnostic radiology entry ]" G.Thimem, Stuttgart, New York (1977).

In general, it has become possible to synthesize novel complexing agents, metal complexes and metal complex salts which open up new possibilities for diagnostic and therapeutic medicine.

The following examples serve to explain the objects of the invention in more detail.

Example 1

a) Bis [2- (benzyloxycarbonylamino) -ethyl ] -amine

51.5g (500mmol) of diethylenetriamine and 139ml (1mol) of triethylamine are dissolved in dichloromethane and 161g of benzylcyanoformate (Fluka) in dichloromethane are admixed at-20 ℃ and stirred overnight at room temperature. After completion of the reaction, the reaction mixture was evaporated and concentrated while being discharged, the residue was dissolved in diethyl ether, and the organic phase was washed with a sodium carbonate solution and dried over sodium sulfate. The filtrate was mixed with hexane, the precipitate was filtered off and dried.

Yield: 163.4g (88% of the theoretical yield)

Elemental analysis:

calculated values: C64.67H 6.78N 11.31

Measured value: C64.58H 6.83N 11.28

b) N, N, N ', N ', N ' -hexa [2- (benzyloxycarbonylamino) -ethyl ] -trimesic acid triamide

13.27g (50mmol) trimesic acid triacyl chloride (Aldrich) and 34.7ml (250mmol) triethylamine are dissolved in Dimethylformamide (DMF) and 65.0g (175mmol) of the amine described in example 1a) are admixed at 0 ℃ and stirred overnight at room temperature. The solution is concentrated by evaporation in vacuo and the residue is chromatographed on silica gel with ethyl acetate.

Yield: 39.4g (62% of theory)

Elemental analysis:

calculated values: C65.24H 5.95N 9.92

Measured value: C65.54H 5.95N 9.87

c)N^α，N^ξDi (N, N' -dibenzyloxycarbonyl-lysyl) -lysine, protected "trilysine"

3.6g (20mmol) of lysine hydrochloride and 6.95ml (50mmol) of triethylamine are dissolved in DMF and 26.8g (50mmol) of N are admixed^α，N^ξDibenzyloxycarbonyl-lysyl-p-nitrophenyl ester (Bachem) and stirred at room temperature for 2 days. After the reaction was complete, it was concentrated by evaporation in vacuo, the residue was dissolved in ethyl acetate and shaken with dilute hydrochloric acid and the layers were separated. The organic phase is dried over sodium sulfate, the solvent is evaporated off and the residue is chromatographed with a gradient of ethyl acetate/ethanol.

Yield: 10.7g (57% of theory)

Elemental analysis:

calculated values: C63.94H 6.65N 8.95

Measured value: C63.63H 6.69N 8.93

d) Fully protected benzyloxycarbonyl-24-polyamines based on N, N, N ', N ', N ' -hexa [2- (lysylamino) -ethyl ] -trimesic acid triamide

1.27g (1mmol) of the hexabenzyloxycarbonylamine described in example 1b) are dissolved in a glacial acetic acid solution and admixed with 33% HBr in glacial acetic acid with stirring. After 60 minutes, the initial precipitation was completed with ether, and the resultant hexamine-hydrobromide was washed with ether, dried in vacuo and used in the next reaction described below without purification.

Yield: 0.95g (quantitative)

7.0g (7.5mmol) of the protected "trilysine" described in example 1C), 1.2g (7.5mmol) of 1-hydroxybenzotriazole and 2.4g (7.5mmol) of 2- (1H-benzotriazol-1-yl) -1, 1, 3, 3-tetramethyluronium tetrafluoroborate (TBTU; peboc Limited, UK) was dissolved in DMF and stirred for 15 min. Then, 5.16ml (30mmol) of N-ethyldiisopropylamine and 0.95g (1mmol) of the above hexamine-hydrobromide were mixed into the solution, and stirred at room temperature overnight. After the reaction is complete, it is concentrated by evaporation in vacuo and the residue is chromatographed on silica gel using ethyl acetate/ethanol (2: 1).

Yield: 4.55g (76% of theory)

Elemental analysis:

calculated values: C64.35H 6.71N 10.52

Measured value: C64.08H 6.57N 10.29

e) 2-Bromopropionylglycine-benzyl ester

55.9g (326.1mmol) of 2-bromopropionyl chloride are added dropwise at 0 ℃ to a solution of 100g (296.4mmol) of benzyl glycinate-p-toluenesulfonate and 33.0g (326.1mmol) of triethylamine in 400ml of dichloromethane. The reaction temperature is not allowed to be higher than 5 ℃. After the addition was complete, the mixture was stirred at 0 ℃ for 1 hour and then at room temperature for two hours. 500ml of ice water are added and the aqueous phase is adjusted to pH 2 with 10% aqueous hydrochloric acid. The organic phase was separated and washed once with 300ml of 5% soda water solution and 400ml of water. The organic phase is dried over magnesium sulfate and evaporated to dryness in vacuo. The residue was recrystallized from isopropyl ether.

Yield: 68.51g (75% of theory) of a colorless crystalline powder

Melting point: 69-70 deg.C

Elemental analysis:

calculated values: C46.76H 7.19N 4.54 Br 25.92

Measured value: C46.91H 7.28N 4.45 Br 25.81

f) - [4- (benzyloxycarbonyl) -1-methyl-2-oxo-3-azabutyl ] -1, 4, 7, 10-tetraazacyclododecane

50g (162.2mmol) of the title compound from example 1e) are added to 55.8g (324.4mmol) of 1, 4, 7, 10-tetraazacyclododecane, dissolved in 600ml of chloroform and stirred at room temperature overnight. 500ml of water are added, the organic phase is separated off and washed twice with 400ml of water each time, dried over magnesium sulfate and evaporated to dryness in vacuo. The residue is chromatographed on silica gel (eluting solvent: chloroform/methanol/25% ammonia 10: 5: 1).

Yield: 40.0g of a pale yellow viscous oil [ 63% of the theoretical yield calculated on the amount of 1e) used ].

Elemental analysis:

calculated values: C61.36H 8.50N 17.89

Measured value: C61.54H 8.68N 17.68

g)10- [4- (benzyloxycarbonyl) -1-methyl-2-oxo-3-azabutyl ] -1, 4, 7-tris (tert-butoxycarbonylmethyl) -1, 4, 7, 10-tetraazacyclododecane (sodium bromide complex)

33g (169mmol) of tert-butyl bromoacetate are added to a solution of 20g (51.08mmol) of the expected compound from example 1f) and 17.91g (169mmol) of sodium carbonate in 300ml of acetonitrile and stirred at 60 ℃ for 24 hours. Cooled to 0 ℃, the formed salt is filtered off and the filtrate is evaporated to dryness. The residue is chromatographed on silica gel (elution solvent: ethyl acetate/ethanol 15: 1). The product-containing eluate is concentrated by evaporation and the residue is recrystallized from isopropyl ether.

Yield: 34.62g (81% of theory) of a colorless crystalline powder

Melting point: 116 ℃ C. (117 ℃ C.)

Elemental analysis:

calculated values: C54.54H 7.59N 8.37 Na 2.74 Br 9.56

Measured value: C54.70H 7.65N 8.24 Na 2.60 Br 9.37

h)10- (4-carboxy-1-methyl-2-oxo-3-azabutyl) -1, 4, 7-tris (tert-butoxycarbonylmethyl) -1, 4, 7, 10-tetraazacyclododecane (sodium bromide complex)

30g (35.85mmol) of the title compound from example 1g are dissolved in 500ml of isopropanol and 3g of palladium catalyst (10% Pd/C) are added. Hydrogenation was carried out at room temperature overnight. The catalyst was filtered off, the filtrate was evaporated to dryness in vacuo and recrystallized from acetone.

Yield: 22.75g (85% of theory) of a colorless crystalline powder

Melting point: 225 ℃ (decomposition)

Elemental analysis:

calculated values: C49.86H 7.69N 9.38 Na 3.07 Br 10.71

Measured value: C49.75H 7.81N 9.25 Na 2.94 Br 10.58

I) 24-monomer unit N- (5-DO 3A-yl-4-oxo-3-azahexanoyl) -cascade polyamide based on N, N, N ', N ', N ' -hexa [2- (trilysinylamino) -ethyl ] -trimesic acid triamide

6.0g (1mmol) of the polybenzyloxycarbonyl amine described in example 1d) are dissolved in glacial acetic acid and a 33% solution of hydrogen bromide in glacial acetic acid is mixed in with stirring. After 3 hours, the initial precipitation was completed with diethyl ether and the resulting 24-amine-hydrobromide salt was washed with ether and dried under vacuum.

35.84(48mmol) of the acid described in example 1h) above in DMF, 7.35g (48mmol) of 1-hydroxybenzotriazole, 15.41g (48mmol) of TBTU (Peboc Limited, UK) and 49.3ml (288mmol) of N-ethyldiisopropylamine are mixed and stirred at room temperature for 20 min. Subsequently, the above (1mmol) 24-amine-hydrobromide was mixed into the solution, and the mixture was stirred at room temperature for 4 days. The solution was concentrated by evaporation under vacuum and the residual oil was cooled in an ice bath and mixed with trifluoroacetic acid, stirred at room temperature overnight and then precipitated with ether, the precipitate was dried under vacuum, dissolved in water, adjusted to pH 7 and washed with YM3 Amicon^(R)The low molecular weight fraction was removed by ultrafiltration membrane and the retentate was finally membrane filtered and lyophilized.

Yield: 13.5g (83% of theory)

H₂O content (Karl-Fischer method): 6.2 percent

Elemental analysis (calculated as anhydride):

calculated values: C45.82H 6.09N 15.07 Na 10.79

Measured value: C45.56H 6.15N 14.80 Na 10.52

^*) DO3A ═ 1, 4, 7-tris (carboxymethyl) -1, 4, 7, 10-tetraazacyclododecane

k) 24-monomer unit-Gd-complexes of N- (5-DO 3A-yl-4-oxo-3-azahexanoyl) -cascade polyamides based on N, N, N ', N ', N ' -hexa [2- (trilysinylamino) -ethyl ] -trimesic acid triamide

8.13g (0.5mmol) of the polymerization agent acid described in example 1i) above are adjusted to pH 3 in water with dilute hydrochloric acid, 2.17g (6mmol) Gd₂O₃Stirring at 80 deg.C for 30min, cooling, adjusting pH to 7, and treating with YM3 AMICON^(R)Desalting with an ultrafiltration membrane. The final retentate was membrane filtered and lyophilized.

Yield: 8.89g (92.1% of theory)

H₂O content (Karl-Fischer method): 9.6 percent

Gd measurement (AAS method): 19.6 percent

Elemental analysis (calculated as anhydride):

calculated values: C40.26H 5.35N 13.24 Gd 21.62

Measured value: C39.98H 5.51N 13.42 Gd 21.37

Example 2

a) 2-bromopropionyl-beta-alanine-phenyl ester

53.65g (313mmol) of 2-bromopropionyl chloride are added dropwise at 0 ℃ to a solution of 100g (285mmol) of beta-alanine-phenyl ester-p-toluenesulfonate and 31.67g (313mmol) of triethylamine in 400ml of dichloromethane. The reaction temperature must not exceed 5 ℃. After completion of the dropwise addition, the mixture was stirred at 0 ℃ for 1 hour and then at room temperature for 2 hours. 500ml of ice water was added and the aqueous phase was adjusted to pH 2 with 10% aqueous hydrochloric acid. The organic phase was separated and washed once with 300ml of 5% aqueous hydrochloric acid, 300ml of 5% aqueous soda and 400ml of water. The organic phase is dried over magnesium sulfate and evaporated to dryness in vacuo. The residue was recrystallized from isopropyl ether.

Yield: 71.36g (78% of theory) of a colorless crystalline powder

Elemental analysis:

calculated values: C48.46H 7.51N 4.35 Br 24.80

Measured value: C48.29H 7.65N 4.25 Br 24.61

b)1- [5- (benzyloxycarbonyl) -1-methyl-2-oxo-3-azapentyl ] -1, 4, 7, 10-tetraazacyclododecane

50g (155.2mmol) of the title compound from example 2a) are added to a solution of 53.32g (310mmol) of 1, 4, 7, 10-tetraazacyclododecane and 600ml of chloroform and stirred at room temperature overnight. 500ml of water are added and the organic phase is separated and washed twice with 400ml of water each time. The organic phase is dried over magnesium sulfate and evaporated to dryness in vacuo. The residue was chromatographed on silica gel (eluting solvent: chloroform/methanol/25% aqueous ammonia: 10/5/1).

Yield: 38.39g [ 61% of the theoretical yield calculated on the amount of 2a) used ] of a pale yellow viscous oil.

Elemental analysis:

calculated values: C62.20H 8.70N 17.27

Measured value: C62.05H 8.81N 17.15

c)10- [5- (benzyloxycarbonyl) -1-methyl-2-oxo-3-azapentyl ] -1, 4, 7-tris (tert-butoxycarbonylmethyl) -1, 4, 7, 10-tetraazacyclododecane (sodium bromide complex)

31.8g (163mmol) of tert-butyl bromoacetate are added to a solution of 20g (49.32mmol) of the expected compound from example 2b) and 17.28g (163mmol) of sodium carbonate in 300ml of acetonitrile and stirred at 60 ℃ for 24 hours. Cooled to 0 ℃, the salt formed is filtered off and the filtrate is evaporated to dryness. The residue is chromatographed on silica gel (elution solvent: ethyl acetate/ethanol 10: 1). The product-containing eluate fraction is concentrated by evaporation and the residue is recrystallized from isopropyl ether.

Yield: 31.89g (76% of theory) of a colorless crystalline powder

Elemental analysis:

calculated values: C55.05H 7.70N 8.23 Na 2.69 Br 9.40

Measured value: C55.17H 7.85N 8.10 Na 2.51 Br 9.30

d)10- [5- (carboxy) -1-methyl-2-oxo-3-azapentyl ] -1, 4, 7-tris (tert-butoxycarbonylmethyl) -1, 4, 7, 10-tetraazacyclododecane (sodium bromide complex)

30g (35.26mmol) of the title compound from example 2C) are dissolved in 500ml of isopropanol and 3g of palladium catalyst (10% Pd/C) are added. Hydrogenation was carried out at room temperature overnight. The catalyst was filtered off, the filtrate was evaporated to dryness in vacuo and recrystallized from acetone.

Yield: 24.41g (91% of theory) of a colorless crystalline powder

Elemental analysis:

calculated values: C50.52H 7.82N 9.21 Na 3.01 Br 10.52

Measured value: C50.41H 7.95N 9.10 Na 2.91 Br 10.37

e) Cascade polyamides based on the 24-monomer unit N- (6-DO 3A-yl-5-oxo-4-azepinoyl) -trimesic acid triamide of N, N, N ', N ', N ' -hexa [2- (trilysinylamino) -ethyl ] -trimesic acid triamide

36.52(48mmol) of the acid described in example 2d) above in DMF, 7.35g (48mmol) of 1-hydroxybenzotriazole, 15.41g (48mmol) of TBTU (Peboc Limited, UK) and 49.3ml (288mmol) of N-ethyldiisopropylamine are mixed and stirred at room temperature for 20 min. Subsequently, the above (1mmol) 24-amine-hydrobromide was mixed into the solution, and the mixture was stirred at room temperature for 4 days. The solution was concentrated by evaporation in vacuo and the residual oil was cooled in an ice bath and mixed with trifluoroacetic acid, stirred overnight at room temperature and then precipitated with ether. Vacuum drying the precipitate, dissolving in water, adjusting pH to 7, and treating with YM3 Amicon^(R)Ultrafiltering and membrane removing low molecular weight fraction, and finally filtering and freeze drying retentate.

Yield: 14.4g (85% of theory)

H₂O content (Karl-Fischer method): 8.7 percent

Elemental analysis (calculated as anhydride):

calculated values: C46.82H 5.98N 14.79 Na 10.59

Measured value: C47.04H 6.23N 14.96 Na 10.26

f) 24-monomer unit-Gd-complexes of N- (6-DO 3A-yl-5-oxo-4-azepinoyl) -cascade polyamides based on N, N, N ', N ', N ' -hexa [2- (trilysinylamino) -ethyl ] -trimesic acid triamide

8.5g (0.5mmol) of the polymerization agent acid described in example 2e) above are adjusted to pH 3 in water with dilute hydrochloric acid, 2.17g (6mmol) Gd₂O₃Stirring at 80 ℃ for 30 minutesCooling, adjusting pH to 7 and adding YM3 AMICON^(R)Desalting with an ultrafiltration membrane. The final retentate was membrane filtered and lyophilized.

Yield: 8.50g (88% of theory)

H₂O content (Karl-Fischer method): 7.9 percent

Gd measurement (AAS method): 19.4 percent

Elemental analysis (calculated as anhydride):

calculated values: C41.12H 5.52N 12.99 Gd 21.21

Measured value: C40.86H 5.34N 13.25 Gd 20.95

Example 3

a) N, N' -bis (benzyloxycarbonyl) -3- [ carboxymethoxyacetyl ] -3-aza-pentane-1, 5-diamine

37.14g (100mmol) of the bis (benzyloxycarbonyl-aminoethyl) -amine described in example 1a) are dissolved in DMF and 17.4(150mmol) of diglycolic anhydride (Janssen Chimica) and 21ml (150mmol) of triethylamine are admixed in an ice bath and stirred at room temperature overnight. The solution is concentrated by evaporation in vacuo, the residue is dissolved in ethyl acetate and shaken with dilute hydrochloric acid and the layers are separated. The organic phase is dried over sodium sulfate, the drying agent is filtered off and crystallized by addition of hexane.

Yield: 41.4g (85% of theory)

Elemental analysis:

calculated values: C59.13H 6.00N 8.62

Measured value: C58.99H 5.93N 8.70

b) N, N', N "-tetrakis {8- (benzyloxycarbonylamino) -6- [ 2-benzyloxycarbonylaminoethyl ] -5-oxo-3-oxooctanoyl } cycloalkene

345mg (2mmol) of 1, 4, 7, 10-tetraazacyclododecane (cycloolefin; Fluka) are dried with toluene under reflux with water. A solution of 4.88g (10mmol) of N, N' -bis (benzyloxycarbonyl) -3- [ carboxymethoxyacetyl ] -3-aza-pentane-1, 5-diamine [ example 3a) ] in Tetrahydrofuran (THF) and 2.47g (10mmol) of 2-ethoxy-1-ethoxycarbonyl-1, 2-dihydroquinoline (EEDQ; fluka) was added to the cooled cycloolefintoluene solution and stirred overnight. After completion of the reaction, hexane was added to precipitate the product, the solvent was decanted and re-precipitated once with THF/hexane, followed by once with THF/toluene. After drying in vacuo, 2.78g (68% of theory) of a pale yellow solid are obtained.

Elemental analysis:

calculated values: C60.93H 6.29N 10.93

Measured value: C60.68H 6.40N 10.97

c) Based on the general formula of N, N' -tetrakis {8- (benzyloxycarbonylamino) -6- [ 2-benzyloxycarbonylaminoethyl group]N from (E) -5-oxo-3-oxaoctanoyl } cycloalkenes^α，N^ξFully protected benzyloxycarbonyl-32-polyamines of 32-amines condensed with-di (lysyl) -lysine ("trilysine

2.05g (1mmol) of the octabenzyloxycarbonylamine described in example 3b) are dissolved in a glacial acetic acid solution and admixed with 33% HBr in glacial acetic acid with stirring. After 90 minutes, the initial precipitation was completed with ether and the generated octaamine-hydrobromide was washed with ether, dried in vacuo and used in the next reaction described below without purification.

Yield: 1.6g (quantitative)

9.4g (10mmol) of the protected "trilysine" described in example 1C), 1.5g (10mmol) of 1-hydroxybenzotriazole and 3.2g (10mmol) of 2- (1H-benzotriazol-1-yl) -1, 1, 3, 3-tetramethyluronium tetrafluoroborate (TBTU; peboc Limited, UK) was dissolved in DMF and stirred for 15 min. Then, 5.16ml (30mmol) of N-ethyldiisopropylamine and 1.6g (1mmol) of the above-mentioned octamine-hydrobromide were mixed into the solution, and stirred at room temperature overnight. After completion of the reaction, it is concentrated by evaporation in vacuo and the residue is chromatographed on silica gel using dichloromethane/methanol (10: 1).

Yield: 6.0g (72% of theory)

Elemental analysis:

calculated values: C63.32H 6.76N 10.74

Measured value: C62.98H 6.91N 10.43

d) 32-monomer unit N- (5-DO 3A-yl-4-oxo-3-azahexanoyl) -cascade polyamides based on 32-monomer unit amines described in example 3c)

8.35g (1mmol) of the 32-mer unit benzyloxycarbonylamine described in example 3c) are dissolved in glacial acetic acid and admixed with a 33% solution of hydrogen bromide in glacial acetic acid with stirring. After 3 hours, the initial precipitation was completed with diethyl ether and the resulting 32-amine-hydrobromide salt was washed with ether and dried under vacuum.

47.8g (64mmol) of the acid described in example 1h) are dissolved in DMF, and 9.8g (64mmol) of 1-hydroxybenzotriazole, 20.5g (64mmol) of TBTU (Peboc Limited, UK) and 65.7ml (384mmol) of N-ethyldiisopropylamine are mixed and stirred at room temperature for 20 minutes. Subsequently, the above (1mmol) 32-amine-hydrobromide was mixed into the solution, and the mixture was stirred at room temperature for 4 days. The solution was concentrated by evaporation in vacuo and the residual oil was cooled in an ice bath and mixed with trifluoroacetic acid, stirred overnight at room temperature and then precipitated with ether. Vacuum drying the precipitate, dissolving in water, adjusting pH to 7, and treating with YM3 Amicon^(R)The low molecular weight fraction was removed by ultrafiltration membrane and the retentate was finally membrane filtered and lyophilized.

Yield: 17.2g (76.4% of theory)

H₂O content (Karl-Fischer method): 7.6 percent

Elemental analysis (calculated as anhydride):

calculated values: C45.73H 6.12N 15.08 Na 10.61

Measured value: C45.89H 6.30N 14.84 Na 10.31

e) 32-monomer unit-Gd-complexes of N- (5-DO 3A-yl-4-oxo-3-azahexanoyl) -cascade polyamides based on 32-monomer unit amines described in example 3c)

10.4g (0.5mmol) of the polymerization agent acid described in example 3d) above are adjusted to pH 3 in water with dilute hydrochloric acid, 2.89g (8mmol) Gd₂O₃Stirring at 80 deg.C for 30min, cooling, adjusting pH to 7, and mixing with YM3 AMICON^(R)Desalting with an ultrafiltration membrane. The final retentate was membrane filtered and lyophilized.

Yield: 12.1g (91.1% of theory)

H₂O content (Karl-Fischer method): 11.0 percent

Gd measurement (AAS method): 18.6 percent

Elemental analysis (calculated as anhydride):

calculated values: C40.26H 5.39N 13.28 Gd 21.30

Measured value: C40.10H 5.21N 13.04 Gd 21.03

Similarly from Yb₂(CO₃)₃Preparing an yttrium complex:

elemental analysis (calculated as anhydride):

calculated values: C39.42H 5.28N 13.00 Yb 22.94

Measured value: C39.29H 5.40N 12.81 Yb 22.65

Example 4

a) Hexaethylene glycol monomethyl ether-p-toluenesulfonate ester

14.3g (75mmol) of p-toluenesulfonyl chloride are added portionwise at 0 ℃ to a solution of 20g (67.49mmol) of hexaethyleneglycol monomethyl ether and 7.59g (75mmol) of triethylamine in 200ml of chloroform, and stirring is carried out for 4 hours at this temperature. Evaporated in vacuo and the residue chromatographed on silica gel (eluting solvent: chloroform/methanol: 5/1).

Yield: 27.67g (91% of theory) of a flaky, glassy solid

Elemental analysis:

calculated values: C53.32H 7.61S 7.12

Measured value: C53.15H 7.70S 7.03

b) 1-benzyloxy-5- (benzyloxycarbonyl) -2-chloro-3-oxo-4-azapentane

76g (326.1mmol) of 2-chloro-3- (benzyloxy) -propionyl chloride (prepared according to Inorg. chem. Vol.31; 2422, 1992) are added dropwise at 0 ℃ to a solution of 100g (296.4mmol) of benzyl glycinate-p-toluenesulfonate and 33.0g (326.1mmol) of triethylamine in 400ml of dichloromethane and stirred at this temperature for 2 hours. 500ml of ice water was added and the aqueous phase was adjusted to pH 2 with 10% aqueous hydrochloric acid. The organic phase was separated and washed once with 300ml of 5% aqueous hydrochloric acid, 300ml of 5% aqueous soda and 400ml of water. The organic phase is dried over magnesium sulfate and evaporated to dryness in vacuo. The residue was chromatographed on silica gel (eluting solvent: dichloromethane/hexane/acetone 15/5/1).

Yield: 75.07g (70% of theory) of a pale yellow viscous oil

Elemental analysis:

calculated values: C63.07H 5.57N 3.87 Cl 9.80

Measured value: C63.17H 5.65N 3.75 Cl 9.63

c)1- [4- (benzyloxycarbonyl) -1- (benzyloxymethyl) -2-oxo-3-azabutyl ] -1, 4, 7, 10-tetraazacyclododecane

70g (193.5mmol) of the title compound from example 4b) and 11.1g (64.5mmol) of 1, 4, 7, 10-tetraazacyclododecane are dissolved in 70ml of dimethylformamide and stirred at 50 ℃ for 2 days. Evaporated to dryness in vacuo, 700ml of water are added to the residue and extracted 2 times with 250ml of chloroform each time. The organic phase is dried over magnesium sulfate and evaporated to dryness in vacuo. The residue was chromatographed on silica gel (elution solvent: chloroform/methanol/25% aqueous ammonia: 10/5/1).

Yield: 13.16g [ 41% of the theoretical yield, calculated on the amount of cycloalkene used ] of a colorless viscous oil.

Elemental analysis:

calculated values: C65.17H 7.90N 14.07

Measured value: C65.24H 7.77N 14.18

d)10- [4- (benzyloxycarbonyl) -1- (benzyloxymethyl) -2-oxo-3-azabutyl ] -1, 4, 7-tris (tert-butoxycarbonylmethyl) -1, 4, 7, 10-tetraazacyclododecane (sodium bromide complex)

16.81g (86.2mmol) of tert-butyl bromoacetate are added to a solution of 13g (26.12mmol) of the expected compound from example 4c) and 9.14g (86.2mmol) of sodium carbonate in 200ml of acetonitrile and stirred at 60 ℃ for 24 hours. Cooled to 0 ℃, the salt formed is filtered off and the filtrate is evaporated to dryness. The residue is chromatographed on silica gel (elution solvent: ethyl acetate/ethanol 15: 1).

Yield: 19.46g (79% of theory) of a waxy solid

Elemental analysis:

calculated values: C57.32H 7.38N 7.43 Na 2.43 Br 8.47

Measured value: C57.22H 7.51N 7.27 Na 2.33 Br 8.29

e)10- (4-carboxy-2-oxo-1-hydroxymethyl-3-azetidinyl) -1, 4, 7-tris (tert-butoxycarbonylmethyl) -1, 4, 7, 10-tetraazacyclododecane (sodium bromide complex)

19g (20.15mmol) of the title compound from example 4d) are dissolved in 300ml of isopropanol and 3g of palladium catalyst (10% Pd/C) are added. Hydrogenation was carried out at room temperature overnight. The catalyst was filtered off, the filtrate was evaporated to dryness in vacuo and recrystallized from acetone.

Yield: 13.06g (85% of theory) of a colorless crystalline powder

Elemental analysis:

calculated values: C48.82H 7.53N 9.18 Na 3.00 Br 10.49

Measured value: C48.71H 7.68N 9.03 Na 2.81 Br 10.23

f)10- [4- (benzyloxycarbonyl) -1- (hydroxymethyl) -2-oxo-3-azabutyl ] -1, 4, 7-tris (tert-butoxycarbonylmethyl) -1, 4, 7, 10-tetraazacyclododecane

3.42g (20mmol) of benzyl bromide are added to a solution of 13g (17.04mmol) of the expected compound from example 4e) and 6.11g (18.75mmol) of anhydrous cesium carbonate in 70ml of dimethylformamide and stirred at 50 ℃ overnight. Cooled to 0 ℃ and 700ml of water are added. This is followed by 2 extractions with 300ml of dichloromethane each time. The organic phases were combined and washed 2 times with water, dried over anhydrous magnesium sulphate and evaporated to dryness in vacuo. The residue is chromatographed on silica gel (eluting solvent: ethyl acetate/ethanol).

Yield: 9.97g (78% of theory) of a colorless viscous oil.

Elemental analysis:

calculated values: C60.86H 8.47N 9.34

Measured value: C60.95H 8.62N 9.21

g)10- [4- (benzyloxycarbonylacyl) -1- (2, 5, 8, 11, 14, 17, 20-heptaoxaheneicosyl) -2-oxo-3-azabutyl ] -1, 4, 7-tris (tert-butoxycarbonylmethyl) -1, 4, 7, 10-tetraazacyclododecane

9.7g (12.93mmol) of the title compound from example 4f) are dissolved in 50ml of THF and 0.43g (14.22mmol) of sodium hydride (80% in paraffin oil) are added at-10 ℃. Stirring was carried out at 0 ℃ for 30 minutes. 11.65g (25.86mmol) of the title compound from example 4a) and 3.46g (25.86mmol) of lithium iodide are then added. Stirred at room temperature for 24 hours. 3ml of water are carefully added and then evaporated to dryness. The residue was chromatographed on silica gel (eluting solvent: chloroform/methanol 10: 1).

Yield: 12.1g (91% of theory) of a glassy solid.

Elemental analysis:

calculated values: C59.57H 8.72N 6.81

Measured value: C59.65H 8.91N 6.62

h)10- [1- (2, 5, 8, 11, 14, 17, 20-heptaoxaheneicosyl) -2-oxo-3-aza-4- (carboxy) butyl ] -1, 4, 7-tris (tert-butoxycarbonylmethyl) -1, 4, 7, 10-tetraazacyclododecane

12g (11.67mmol) of the title compound from example 4g) are dissolved in 300ml of isopropanol and 2g of palladium catalyst (10% Pd/C) are added. Hydrogenation was carried out at room temperature overnight. The catalyst was filtered off, the filtrate was evaporated to dryness in vacuo and recrystallized from acetone/isopropyl ether.

Yield: 10.18g (93% of theory) of a waxy solid

Elemental analysis:

calculated values: C56.33H 8.92N 7.46

Measured value: C56.20H 9.03N 7.35

i) 24-monomer units-Gd complexes of N- (5-DO 3A-yl-4-oxo-3-aza-7, 10, 13, 16, 19, 22, 25-heptaoxa-hexacosanyl) -cascade polyamides based on N, N, N ', N ', N ' -hexa [2- (trilysinylamino) -ethyl ] -trimesic acid triamide

6.0g (1mmol) of the 24-mer unit-benzyloxycarbonylamine described in example 1d) are dissolved in glacial acetic acid and admixed with a 33% solution of hydrogen bromide in glacial acetic acid with stirring. After 3 hours, the initial precipitation was completed with diethyl ether and the resulting 24-amine-hydrobromide salt was washed with ether and dried under vacuum. 45.03g (48mmol) of the acid described in example 4h) above in DMFTo this solution were mixed 7.35g (48mmol) of 1-hydroxybenzotriazole, 15.41g (48mmol) of TBTU (Peboc Limited, UK) and 49.3ml (288mmol) of N-ethyldiisopropylamine, and the mixture was stirred at room temperature for 20 minutes. Subsequently, the above (1mmol) 24-amine-hydrobromide was mixed into the solution, and the mixture was stirred at room temperature for 4 days. The solution was concentrated by evaporation in vacuo and the residual oil was cooled in an ice bath and mixed with trifluoroacetic acid, stirred overnight at room temperature and then precipitated with ether. The precipitate was dried under vacuum, dissolved in water, adjusted to pH 3 with dilute hydrochloric acid and admixed with 8.70g (24mmol) of Gd₂O₃Stirring at 80 deg.C for 4 hr, cooling, adjusting pH to 7, and mixing with YM3 AMICON^(R)The low molecular weight fraction is removed by ultrafiltration membranes. The final retentate was membrane filtered and lyophilized.

Yield: 19.6g (73.3% of theory)

H₂O content (Karl-Fischer method): 8.3 percent of

^GdMeasurement value (AAS method): 14.0 percent

Elemental analysis (calculated as anhydride):

calculated values: C43.94H 6.38N 9.43 Gd 15.39

Measured value: C44.27H 6.22N 9.29 Gd 15.09

Example 5

a)1, 7-bis (trifluoroacetyl) -1, 4, 7-triazaheptane

113.3g (790mmol) of ethyl trifluoroacetate are added dropwise at 80 ℃ under nitrogen to a solution of 41.14g (390mmol) of 1, 4, 7-triazaheptane and 350ml of tetrahydrofuran. Stir at room temperature overnight and concentrate by evaporation under vacuum. The residual oil was crystallized from hexane.

Yield: 115g (99.9% of theory)

Melting point: 68-70 deg.C

Elemental analysis:

calculated values: C32.55H 3.76F 38.62N 14.24

Measured value: C32.63H 3.75F 38.38N 14.19

b)1, 7-bis (trifluoroacetyl) -4-benzyloxycarbonyl-1, 4, 7-triazaheptane

14.75g (50mmol) of the trifluoroacetyl compound prepared in example 5a) and 8.3ml (60mmol) of triethylamine are dissolved in 120ml of dichloromethane and cooled to 0 ℃. 7.5ml (53mmol) of benzyl chloroformate (97%) dissolved in 20ml of dichloromethane are added dropwise with stirring. Stirring overnight at room temperature, extracting the resulting salt with distilled water, drying the dichloromethane solution over sodium sulfate and evaporating to dryness in vacuo, and crystallizing the residue with diethyl ether/hexane.

Yield: 18.40g (85.7% of theory)

Melting point: 131 ℃ C. and 132 ℃ C

Elemental analysis:

calculated values: C44.76H 3.99F 26.55N 9.79

Measured value: C44.87H 4.03F 26.62N 9.61

c)3, 9-bis (tert-butoxycarbonylmethyl) -6-benzyloxycarbonyl-3, 6, 9-triazaundecanedicarboxylic acid di-tert-butyl ester

4.29g (10mmol) of the trifluoroacetyl compound prepared in example 5a) are dissolved in 30ml of ethanol and mixed with a solution of 800mg (20mmol) of sodium hydroxide and 10ml of distilled water. After stirring at room temperature for 3 hours, the mixture was evaporated to dryness at 40 ℃ under vacuum, and the residual water was removed by refluxing with isopropanol and then dissolved in 30ml of dimethylformamide. 6.9g (50mmol) of potassium carbonate and 9.7g (50mmol) of tert-butyl bromoacetate are added and 4-benzyloxycarbonyl-1, 4, 7-triazaheptane is alkylated at room temperature overnight. The dimethylformamide is subsequently removed by means of an oil pump, the residue is partitioned between water and dichloromethane, the organic phase is dried over sodium sulfate and evaporated under vacuum, and the residue is purified by chromatography on silica gel. The title compound was eluted with ethyl acetate/hexane. The product is a bubble.

Yield: 6.49g (93.6% of theory)

Elemental analysis:

calculated values: C62.32H 8.57N 6.06

Measured value: C62.41H 8.66N 6.01

d)3, 9-bis (tert-butoxycarbonylmethyl) -3, 6, 9-triazaundecanedicarboxylic acid di-tert-butyl ester

3.5g (5mmol) of the compound prepared in example 5c) are dissolved in 100ml of ethanol and 200mg of Pearlman's catalyst (20% Pd on activated carbon) are added and hydrogenation is carried out until the calculated amount of hydrogen has been taken up completely. The catalyst was removed by suction filtration and the filtrate was evaporated to dryness in vacuo. The target compound was a white foam.

Yield: 2.80g (99.9% of theory)

Elemental analysis:

calculated values: C60.08H 9.54N 7.51

Measured value: C60.02H 9.62N 7.56

e)3, 9-bis (tert-butoxycarbonylmethyl) -6- [1- (ethoxycarbonyl) -ethyl ] -3, 6, 9-triazaundecanedicarboxylic acid di-tert-butyl ester

5.60g (10mmol) of the amino compound prepared in example 5d) are dissolved in 30ml of dimethylformamide. 1.66g (12mmol) of potassium carbonate and 2.17g (12mmol) of ethyl 2-bromopropionate were added at room temperature and stirred overnight. Then, the mixture was poured into ice water, extracted with ethyl acetate, and the organic solution was dried over sodium sulfate, evaporated to dryness in vacuo, and chromatographed on silica gel to give the desired compound. A mixture of ethyl acetate/hexane was used as eluent.

Yield: 4.18g (63.4% of theory)

Elemental analysis:

calculated values: C60.07H 9.32N 6.37

Measured value: C60.18H 9.40N 6.31

f)3, 9-bis (tert-butoxycarbonylmethyl) -6- [1- (carboxy) -ethyl ] -3, 6, 9-triazaundecanedicarboxylic acid di-tert-butyl ester

6.60g (10mmol) of the compound prepared in example 5e) are dissolved in 50ml of ethanol. A solution of 400mg (10mmol) of sodium hydroxide and 5ml of distilled water is added. Stirring was carried out at 50 ℃ for 3 hours. The saponification was determined quantitatively by thin layer chromatography. Evaporated to dryness in vacuo, traces of water were azeotropically removed with ethanol, and the residue was dried in vacuo at 40 ℃. The target compound was obtained as a white powder. The remaining white residue is dissolved in 80ml of aqueous ethanol (9: 1) and admixed with a solution of 535mg (10mmol) of ammonium chloride and 10ml of a distillation agent, with stirring. Evaporated to dryness in vacuo, the fraction soluble in butanol is dissolved out and evaporated again in vacuo. The residue was extracted with toluene. The organic phase was evaporated to dryness under vacuum and the title compound was a foam.

Yield: 5.35g (84.7% of theory)

Elemental analysis:

calculated values: C58.93H 9.09N 6.65

Measured value: C59.01H 9.16N 6.60

g) 24-monomer unit N- { N, N-bis [2- (N, N-bis (carboxymethyl)) -aminoethyl ] -glycyl } -cascade polyamide, sodium salt based on N, N, N ', N ', N ' -hexa [2- (trilysinylamino) -ethyl ] -trimesic acid triamide

6.0g (1mmol) of the polybenzyloxycarbonylamine described in example 1d) are dissolved in glacial acetic acid and a 33% solution of hydrogen bromide in glacial acetic acid is added with stirring. After 3 hours, the initial precipitation was completed with diethyl ether and the resulting 24-amine-hydrobromide salt was washed with ether and dried under vacuum.

30.33g (48mmol) ofThe acid described in example 5f) above was dissolved in DMF and 7.35g (48mmol) of 1-hydroxybenzotriazole, 15.41g (48mmol) of TBTU (Peboc Limited, UK) and 49.3ml (288mmol) of N-ethyldiisopropylamine were mixed and stirred at room temperature for 20 minutes. Subsequently, the above (1mmol) 24-amine-hydrobromide was mixed into the solution, and the mixture was stirred at room temperature for 4 days. The solution was concentrated by evaporation under vacuum and the residual oil was cooled in an ice bath and mixed with trifluoroacetic acid, stirred at room temperature overnight and then precipitated with ether, the precipitate was dried under vacuum, dissolved in water, adjusted to pH 7 and washed with YM3 Amicon^(R)The low molecular weight fraction was removed by ultrafiltration membrane and the retentate was finally membrane filtered and lyophilized.

Yield: 11.0g (86.3% of theory)

H₂O content (Karl-Fischer method): 8.2 percent of

Elemental analysis (calculated as anhydride):

calculated values: C42.87H 5.41N 11.96 Na 12.08

Measured value: C42.78H 5.66N 12.11 Na 11.89

h) 24-monomer unit-Gd complexes, sodium salts of N- { N, N-bis [2- (N, N-bis (carboxymethyl)) -aminoethyl ] -glycyl } -cascade polyamides based on N, N, N ', N' -hexa [2- (trilysinylamino) -ethyl ] -trimesic acid triamide

8.13g (0.5mmol) of the complexing agent acid described in example 5g) above are adjusted to pH 3 in water with dilute hydrochloric acid and 2.17g (6mmol) Gd₂O₃Stirring at 80 deg.C for 30min, cooling, adjusting pH to 7, and treating with YM3 AMICON^(R)Desalting with an ultrafiltration membrane. The final retentate was membrane filtered and lyophilized.

Yield: 8.0g (90.5% of theory)

H₂O content (Karl-Fischer method): 7.5 percent

Gd measurement (AAS method): 21.0 percent

Elemental analysis (calculated as anhydride):

calculated values: C35.93H 4.38N 10.03 Gd 23.09 Na 3.38

Measured value: C35.71H 4.65N 9.88 Gd 22.84 Na 3.50

Example 6

a)3, 9-bis (tert-butoxycarbonylmethyl) -6-benzyloxycarbonylmethyl-3, 6, 9-triazaundecanedioic acid ditert-butyl ester

5.60g (10mmol) of the amino compound prepared in example 5d) are dissolved in 30ml of dimethylformamide. Then 1.66g (12mmol) of potassium carbonate and 2.58(12mmol) of benzyl bromoacetate were added at room temperature and stirred overnight. Pouring into ice water, extracting with ethyl acetate, drying the organic phase with sodium sulfate, evaporating to dryness under vacuum, and separating by silica gel chromatography to obtain the target compound. The eluent was ethyl acetate/hexane.

Yield: 6.32g (89.3% of theory)

Elemental analysis:

calculated values: C64.65H 9.00N 5.95

Measured value: C64.62H 9.07N 5.90

b)3, 9-bis (tert-butoxycarbonylmethyl) -6-carboxymethyl-3, 6, 9-triazaundecanedioic acid ditert-butyl ester

7.08(10mmol) of the benzyl ester prepared in example 6a) are dissolved in 100ml of ethanol and 0.4g of Pearlman's catalyst (Pd 20%, C) are admixed. Hydrogenation until 224ml hydrogen is absorbed, suction filtration to remove the catalyst, and ethanol washing, solution in vacuum evaporation to dryness. The product was obtained as a foam and recrystallized from ether/hexane.

Yield: 6.87g (97.3% of theory)

Melting point: 73-75 deg.C

Elemental analysis:

calculated values: C57.85H 9.00N 5.95

Measured value: C57.91H 9.11N 6.01

c) 32-monomeric unit N- { N, N-bis [2- (N, N-bis (carboxymethyl)) -aminoethyl ] -glycyl } -cascade polyamide, nanosalt based on the 32-monomeric unit amine described in example 3c)

39.5g (64mmol) of the acid described in example 6b) are dissolved in DMF, and 9.8g (64mmol) of 1-hydroxybenzotriazole, 20.5g (64mmol) of TBTU (Peboc Limited, UK) and 65.7ml (384mmol) of N-ethyldiisopropylamine are mixed and stirred at room temperature for 20 minutes. Subsequently, the above (1mmol) 32-amine-hydrobromide was mixed into the solution, and the mixture was stirred at room temperature for 4 days. The solution was concentrated by evaporation in vacuo and the residual oil was cooled in an ice bath and mixed with trifluoroacetic acid, stirred at room temperature overnight and then precipitated with ether. Vacuum drying the precipitate, dissolving in water, adjusting pH to 7, and treating with YM3 Amicon^(R)Ultrafiltering and membrane removing low molecular weight fraction, and finally filtering and freeze drying retentate.

Yield: 15.7g (78.6% of theory)

H₂O content (Karl-Fischer method): 9.0 percent

Elemental analysis (calculated as anhydride):

calculated values: C41.77H 5.24N 12.33 Na 12.14

Measured value: C41.49H 5.36N 12.49 Na 11.93

e) 32-monomer unit-Gd complexes, sodium salts of N- { N, N-bis [2- (N, N-bis (carboxymethyl)) -aminoethyl ] -glycyl } -cascade polyamides based on the 32-monomer unit amines described in example 3c)

10.0g (0.5mmol) of the polymerization agent acid described in example 6c) above are adjusted to pH 3 in water with dilute hydrochloric acid, 2.89g (8mmol) Gd₂O₃Stirring at 80 deg.C for 30min, cooling, adjusting pH to 7, and treating with YM3 AMICON^(R)Desalting with an ultrafiltration membrane. The final retentate was membrane filtered and lyophilized.

Yield: 10.9g (90.9% of theory)

H₂O content (Karl-Fischer method): 9.5 percent

Gd measurement (AAS method): 20.9 percent

Elemental analysis (calculated as anhydride):

calculated values: C34.98H 4.24N 10.33 Gd 23.19 Na 3.39

Measured value: C35.20H 4.08N 10.46 Gd 22.89 Na 3.60

Example of in vivo contrast experiment with an extracellular contrast agent

The following experiments were carried out to show the suitability of the compounds described in example 1k) as contrast agents for retention in the blood.

Five male (Schering-SPF) mice weighing 300-350g were used as experimental animals. Before the experiment, the intestine was removed by opening the abdomen, and the bilateral renal vessels (artery + vein) were ligated with a surgical needle through the posterior peritoneum. The abdominal incision is then closed. 0.3ml (50 mmol/l each) of the following contrast agent solutions were administered intravenously to each animal: compound of example 1k), hereinafter compound 1, and 10- (1-hydroxymethyl-2, 3-dihydroxypropyl) -1, 4, 7-tris (carboxymethyl) -1, 4, 7, 10-tetraazacyclododecane, hereinafter compound 2, prepared analogously to european patent application EP 448191, in each case 1 part by weight. Blood samples were taken from the carotid artery with the catheter at the following times, 15, 30, 45, 60, 90 seconds, 3, 5, 10, 15 minutes after injection. The obtained blood samples were subjected to atomic emission spectrometry (ICP-AES) to measure the concentrations of gadolinium (Gd) and dysprosium (Dy) in a parallel manner. The fraction of injected contrast agent compound 1(Gd) and compound 2(Dy, reference substance) remaining in the blood was compared with the same animals which could be labeled differently. Since renal excretion is not possible, the decrease in blood concentration can only be attributed to dispersion into the blood fraction and diffusion into the interstitial spaces.

Results: the diffusion of compound 1 into the interstitial space is significantly reduced compared to the extracellular contrast agent compound 2 (see figure 1).

The extracellular contrast agent (compound 2) diffuses rapidly into the interstitial space of the body and reaches equilibrium (at a constant concentration in the blood) 3-5 minutes after injection. But on the contrary; the cascade polymer (compound 1) not only has a constantly high blood concentration (referring to a small volume of dispersion) but also does not reach equilibrium (pointing to very slow diffusion of interstitial tissue) throughout the 15 minute examination period. This indicates that compound 1 has the behavior of a contrast agent retained in the blood.

Example of MR angiography on rabbits

The compound of example 1k was used in an MR angiography study in rabbits (CH. R. Kisslegg, approximately 4Kg Body weight) (Ganzkorper [ white-Body ] MRT System Siemens Vision, 1.5 Tesla, FISP 3D; TR: 400 ms; TE15 ms; flip angle; (45 °; coronal)).

In the image without contrast agent (see fig. 2), only one to two major vessels (e.g. the abdominal aorta) are visible, and the contrast (signal intensity SI of these vessels against background) is relatively poor. After intravenous administration of 50 μmol Gd/kg body weight the contrast (vascular SI/background SI) was significantly enhanced and a variety of small and capillary vessels (e.g., femoral artery, vein, mesenteric tail artery, vein, renal artery, vein, and renal subunit artery, vein, etc.) were also visible that were not detectable without contrast agent.

Example of lymph node accumulation in guinea pig

The compounds of the invention of example 1k were tested in the following 30 minutes to 24 hours in stimulated guinea pigs (complete Freund's adjuct; 0.1ml per intramuscular injection of the left and right upper and lower extremities; two weeks before administration of the test substance) administered subcutaneously to determine lymph node accumulation at three consecutive lymph node locations (knee, inguinal, iliac). The results are given in the following table (gadolinium lymph accumulation by means of ICP-AES):

timing of lymph node resection	Gadolinium concentration in three consecutive lymph node locations [ mu. mol/l][% dose/gram of tissue]
					Knee	Groin	Ilium	Ratio of
	30min after injection	921μmol/l20.1％	387μmol/l8.5％	215μmol/l4.7％	Knee	Groin	Ilium	Ratio of	10∶4.2∶2.3
90min after injection	30min after injection	921μmol/l20.1％	387μmol/l8.5％	215μmol/l4.7％	659μmol/l14.4％	120μmol/l2.6％	68μmol/l1.5％	10∶1.8∶1.0	10∶4.2∶2.3
90min after injection	4h after injection	176μmol/l3.9％	79μmol/l1.7％	47μmol/l1.0％	659μmol/l14.4％	120μmol/l2.6％	68μmol/l1.5％	10∶1.8∶1.0	10∶4.5∶2.7
24h after injection	4h after injection	176μmol/l3.9％	79μmol/l1.7％	47μmol/l1.0％	62μmol/l1.4％	13μmol/l0.3％	28μmol/l0.6％	10∶2.1∶4.5	10∶4.5∶2.7

Claims

1. Compounds of the general formula I' A

Wherein

R²represents a hydrogen atom, and is represented by,

R³' represents a group

U⁶represents a methylene group or an ethylene group,

t' represents a-C^*O, -COOH, -N ═ C ═ O or-N ═ C ═ S group, and

C^*o represents an activated carboxyl group.