EP0853488A2

EP0853488A2 - Xsns-type bi-functional sulphide-containing sulphonamide chelating agents for radioactive isotopes

Info

Publication number: EP0853488A2
Application number: EP96945139A
Authority: EP
Inventors: Ludger Dinkelborg; Christoph Stephan Hilger; Wolfgang Kramp; Johannes Platzek; Bernd Radüchel; Sebastian Erber
Original assignee: Institut fuer Diagnostikforschung GmbH
Current assignee: Bayer Pharma AG
Priority date: 1995-09-21
Filing date: 1996-09-19
Publication date: 1998-07-22
Also published as: WO1997010852A2; WO1997010852A3; DE19536781A1; CA2232391A1; AU1435997A

Abstract

The invention pertains to novel compounds of the general formula (I): M - L in which M stands for a radioisotope of Tc or Re, L stands for a ligand of the general formula (II): B-CO-(CR1R2)n=1,2-S-CHR?3-CHR4-SO¿2-NH-CR?5R6-(CR7R8)¿m=1,2-SR9 in which R?1, R2, R3, R4, R5, R6, R7, R8 and R9¿ can have different meanings, and B stands for another group which is suitable both for the dative bond of metal ions and for coupling with selectively concentrating compounds. That coupling can alternatively be effected on R8. These novel compounds serve to form complexes of technetium and rhenium and are used in medical diagnostics and therapy.

Description

Bifunctional sulfide-containing sulfonamide chelating agents of the type XSNS for radioactive isotopes

The invention relates to new chelating agents containing sulfonamide groups, pharmaceutical compositions containing these compounds, their use in radiodiagnostics and radiotherapy, processes for producing these compounds and compositions, and conjugates of these compounds with substances which selectively accumulate in diseased tissue, in particular peptides.

The use of radiopharmaceuticals for diagnostic and therapeutic purposes has long been known in the field of biological and medical research. In particular, radiopharmaceuticals are used to represent certain structures such as the skeleton, organs or tissues. The diagnostic application presupposes the use of such radioactive agents, which are specific to those after application

Enrich structures in the patient to be examined. These locally accumulating radioactive agents can then be tracked, plotted or scintigraphed using suitable detectors, such as scintillation cameras or other suitable recording methods. The distribution and relative intensity of the detected radioactive agent characterizes the location of a structure in which the radioactive agent is located and can be the presence of abnormalities in structure and function, pathological

Represent changes etc. Similarly, radiopharmaceuticals can be used as therapeutic agents to irradiate certain pathological tissues or areas. Such treatment requires the production of radioactive therapeutic agents that accumulate in certain structures, tissues or organs. 4

Ion solutions or normal plasma ions such as Ca ² +, Na ⁺ , K ⁺ and Mg ²⁺ .

Because technetium can exist in a number of oxidation states ( ₊ 7 to -1), it is often necessary for radiopharmaceutical agents to contain additional agents known as stabilizers. These keep the radionuclide in a stable form until it has reacted with the ligand. These stabilizers can include agents known as transfer or auxiliary ligands that are particularly useful in stabilizing and complexing the metal in a well-defined oxidation state until the target ligand complexes the metal via ligand exchange. Examples of this type of auxiliary ligand are

(including their salts) gluconheptonic acid, tartaric acid, citric acid, or other common ligands, as detailed below.

Radionuclide is the standard

Radiopharmaceuticals are shown by first synthesizing the ligand and then reacting it with the metal radionuclide in a suitable manner in order to form a corresponding complex in which the ligand must necessarily be present unchanged after complexation, with the exception of the splitting off of any protective groups or hydrogen ions that may be present. Removal of these groups facilitates coordination of the ligand to the metal ion and thus leads to rapid complexation.

To form technetium-99m complexes, pertechnetate is first obtained from a nuclide generator and, using suitable reducing agents (eg SnCl2 S2O4 ^{2 "} etc.), converted to a lower oxidation level, which is then followed by a suitable one

Chelator is stabilized. Because technetium in a row Complex formation conditions implemented. If, for example, the production of a technetium-99m radiopharmaceutical is desired, the ligand produced is mixed with a pertechnetate solution with the addition of a suitable reducing agent and the corresponding technetium complex is produced under suitable reaction conditions. These complexes are then suitably administered to the patient by injection, inhalation or ingestion.

The solutions containing the radionuclide can, as in the case of technetium-99m, be obtained from an available Mo-99 / Tc-99m nuclide generator, or can be obtained from a manufacturer, as in the case of rhenium-186. The complex formation reaction is carried out at suitable temperatures (e.g. 20 ° -100 ° C) within a few minutes to several hours. To ensure complete complex formation, there is a large excess (more than 100-fold excess to the metal radionuclide) of the ligand produced and a sufficient amount of

Reducing agent required for a complete reduction of the radionuclide used.

Radiopharmaceuticals are made by combining the radionuclide complex, in an amount sufficient for diagnostic or therapeutic use, with pharmacologically acceptable radiological carriers. This radiological carrier should have favorable properties for the application of the radiopharmaceutical in the form of an injection,

Have inhalation or ingestion. Examples of such carriers are HSA, aqueous buffer solutions, for example tris (hydroxymethyl) aminoethane (or their salts), phosphate, citrate, bicarbonate etc., sterile water, physiological saline, isotonic chloride or dicarbonate 6

Suitable complexing agents for technetium and rhenium isotopes are e.g. cyclic amines as described by Volkert et al. (Appl. Radiol. Isot. 1982, 33; 891) and Troutner et al. (J. Nucl. Med. 1980, 21; 443), but they have the disadvantage that they are often only able to bind technetium-99m in good yields from a pH> 9. N2θ2 ~ Syst e (Pillai, M.R.A., Troutner, D.E. et al., - Inorg. Chem. 1990, 29; 1850) are in clinical use. Non-cyclic N4 systems such as the major disadvantage of HMPAO is their low complex stability. Because of its instability (Ballinger, JR et al., Appl. Radiat. Isot. 1991, 42; 315, Billinghurst, MW et al., Appl. Radiat. Isot. 1991, 42; 607), Tc-99m-HMPAO must be used within 30 minutes after it has been marked so that the proportion of decay products that have a different pharmacokinetics and excretion can be kept low. Such radiochemical contaminants make it difficult to identify diseases to be diagnosed. A coupling of these chelates or

Chelating agents to other substances that accumulate selectively in foci of disease cannot be solved by simple means, so that these are generally distributed nonspecifically in the organism.

2S2 chelators (Bormans, G. et al.; Nucl. Med. Biol. 1990, 17; 499) such as e.g. Ethylene dicysteine (EC; Verbruggen, AM et al .; J. Nucl. Med. 1992, 33; 551) meet the requirement for adequate stability of the corresponding technetium-99m complex, but only form when the pH of the complexing medium is> 9 Radio diagnostics with a purity greater than 69%. N3S systems (Fritzburg, A.; EP-0173424 and EP-0250013) form stable technetium-99m complexes, but must be used to form a uniform radiopharmaceutical

Temperatures of about 100 ° C are heated. of oxidation levels (+7 to -1), which can change the pharmacological properties by changing the charges of a complex, it is necessary to provide chelators or complex ligands for technetium-99m, which technetium safely, firmly and stably in a defined Can bind oxidation level in order to prevent undesired biodistribution from taking place in vivo by redox processes or technetium releases from the corresponding radio diagnostics, which complicates the reliable diagnosis of corresponding diseases.

The efficiency of radionuclides in in vivo diagnostics as well as therapy depends on the specificity and the selectivity of the labeled chelates to the target cell. These properties can be improved by coupling the chelates to biomolecules according to the "drug targeting" principle. Antibodies, their fragments, hormones, growth factors and substrates of receptors and enzymes are suitable biomolecules. The British patent application GB 2,109,407 describes the use of radioactively labeled monoclonal antibodies against tumor-associated antigens for tumor diagnosis in vivo. Likewise, direct protein labels via donor groups (amino, amide, thiol, etc.) of the protein (Rhodes, BA et al., J. Nukl. Med. 1986, 27, 685-693) or by introducing complexing agents (U.S. Patent 4,479,930 and Fritzberg, AR et al., Nucl. Med. 1986, 27, 957) with Technetium-99m. However, these experimental methods are not available for clinical use because on the one hand the selectivity is too low and on the other hand the background activity in the organism is too high to enable in vivo imaging. 8th

Substances have very different properties, as well as the mechanisms by which they are enriched, it is still necessary to vary the couplable chelating agent and to be able to adapt it to the physiological requirements of the coupling partner with regard to their lipophilicity, membrane permeability, etc.

The invention is therefore based on the object of providing stable complex compounds which are coupled or capable of coupling to different selectively enriching compounds without their specificity and selectivity being significantly affected. In addition, there is the task of providing couplable chelators or complexes which have a greater chemical range of variation of the substituents in order to be able to adapt them to the requirements referred to above. At the same time, the requirements for the use of these compounds in humans with regard to the absorbed radiation dose,

Stability and solubility of the compounds must be fulfilled.

According to the invention, this object is achieved in that new chelating agents containing bifunctional sulfonamide groups and their coupling products with specifically enriching compounds are made available.

The invention relates to compounds of the general formula (I)

M - L (I)

wherein In recent years, the need for radio-diagnostic agents specifically enriching in diseased tissues has increased. This can be achieved if complexing agents can easily be coupled to selectively enriching substances and do not lose their favorable complexing properties. However, since it is very common that after coupling a complexing agent using one of its functional groups to such a molecule a weakening of the complex stability is observed, the previous approaches for coupling chelating agents to substances which selectively enrich themselves are not very satisfactory, since a diagnostically not tolerable proportion of the isotope from the conjugate is released in vivo (Brechbiel, MW et al.; Inorg. formerly 1986, 25, 2772). It is therefore necessary to prepare bifunctional complexing agents which carry both functional groups for binding the desired metal ion and one (other, several) functional groups for binding the selectively enriching molecule, or to design complexing agents in such a way that they can only be coupled to one another selectively enriching substance the desired complexing agent structure is formed and thus a

Weakening of the complex stability is excluded. Such ligands enable a specific, chemically defined binding of technetium or rhenium isotopes to a wide variety of biological materials, even if so-called prelabeling is carried out. Some chelating agents coupled to monoclonal antibodies (eg EP-0247866 and EP-0188256) or fatty acids (EP-0200492) have been described. However, the 2S2 systems already mentioned are used as chelating agents, which are not very suitable due to their low stability. Because both the selectively enriching 10

Carboxy, aminocarbonyl, alkoxycarbonyl, amino, aldehyde or alkoxy groups with up to 20 carbon atoms is substituted and / or optionally interrupted by one or more heteroatoms from the series 0, N, S, P, As, Se and / or is substituted, stands,

R ⁷ and R ^{8 are the} same or different and each represents a hydrogen atom and / or a branched or unbranched Cι_g alkyl radical,

R ^{9 represents} a hydrogen atom, a branched or unbranched C 1-6 alkyl radical or a sulfur protecting group,

B for a radical -SR ¹¹ , -NHR ¹² or -OR ¹³ , wherein R ^{11 represents} a hydrogen atom, a branched or unbranched C 1-6 alkyl radical or one

Sulfur protection group stands,

R ^{12 represents} a hydrogen atom, an amino protecting group or a branched or straight-chain, cyclic or polycyclic C ₁ _3o-alkyl, alkenyl, polyalkenyl, alkynyl, polyalkynyl, aryl, alkylaryl or arylalkyl group, which are optionally with hydroxy , Oxy, oxo, carboxy, aminocarbonyl, alkoxycarbonyl, amino, aldehyde or alkoxy groups with up to 20 carbon atoms and / or optionally substituted by one or more heteroatoms from the series 0, N, S, P, As, Se is interrupted and / or substituted, M is a radioisotope of Tc or Re and L is a ligand of the general formula (II)

B-CO- (CR ¹ R ² ) _{n = 1/2} -S-CHR ³ -CHR ⁴ -S0 ₂ -NH-CR ⁵ R ⁶ - (CR ⁷ R ⁸ ) _{m = lf 2} "SR ⁹ (II)

means what

R ¹ , R ² , R ³ , R ⁴ and R ^{5 are the} same or different and each represents a hydrogen atom and / or a branched or unbranched C _] __g alkyl radical,

R6 for a hydrogen atom, for a branched or unbranched C _] __g alkyl radical or a radical -CO-R ^{1 (} - ⁾ , in which

R ^{10 is} a hydroxyl, a branched or straight-chain, cyclic or polycyclic Cτ__3Q alkoxy, alkenyloxy, polyalkenyloxy, alkynyloxy, polyalkynyloxy, aryloxy, alkylaryloxy or arylalkyloxy group, optionally with

Hydroxy, oxy, oxo, carboxy, aminocarbonyl, alkoxycarbonyl, amino, aldehyde or alkoxy groups with up to 20 carbon atoms are substituted and / or optionally by one or more heteroatoms from the series O, N, S , P, As, Se is interrupted and / or substituted or an N (R ^a R ^b ) group,

where R ^a and Rb ^{b are} identical or different and / or represent a hydrogen atom, a branched or straight-chain, cyclic or polycyclic Cι_3Q-alkyl, alkenyl, polyalkenyl, alkynyl, polyalkynyl, aryl, alkylaryl or arylalkyl radical, optionally with hydroxy, oxy, oxo, 12

Compounds according to the invention in which R, R ⁴ , R ⁷ and R ^{8 represent} hydrogen atoms are also particularly preferred.

Furthermore, the invention is particularly preferred

Compounds in which R ¹ and R ^{5 represent} hydrogen atoms.

Compounds according to the invention are particularly preferred in which B is a radical NHR ¹² or OR ¹³ , in which

R ¹² and R ^{13 have} the meaning given above

Another object of the invention relates to the new, bifunctional sulfur atom-interrupted sulfonamide ligands of the general formula (II)

B-CO- (CR ^ -R ² ) _{n = 1 r2} -S-CHR ³ -CHR ⁴ -S0 ₂ -NH-CR ⁵ R ⁶ - (CR ⁷ R ⁸ ) _{m = 1 f2} -SR ⁹ (II)

wherein R ¹ , R ² , R ³ , R ⁴ , R ⁵ , R ⁶ / R ⁷ , R ⁸ , R ⁹ , n, and B each have the meaning given above.

Preferred compounds of the general formula (II) are distinguished in that n and m each represent 1 and in that R ¹ , R ³ , R ⁴ , R ⁵ , R ⁷ and R ^{8 are} hydrogen atoms.

Particularly preferred compounds of the general formula (II) are distinguished in that n and m each represent 1 and in that R ¹ , R ³ , R ⁴ , R ⁵ , R ⁷ , R ⁸ and R ^{9 are} hydrogen atoms and R ^{6 is} a hydrogen atom, a branched or unbranched C] __ g alkyl radical or a radical -CO-R ¹⁰ , R ^{13 represents} a hydrogen atom or an alcohol protecting group.

Preferred compounds of the general formula (I) are characterized in that n and m each represent 1 and that R ¹ , R ³ , R ⁴ , R ⁵ , R ⁷ and R ^{8 are} hydrogen atoms.

Particularly preferred compounds of the general formula (I) are characterized in that n and m each represent 1 and that R ¹ , R ³ , R, R ⁵ , R ⁸ and R ^{9 are} hydrogen atoms and R ^{6 is} a hydrogen atom, one branched or unbranched C 1-6 alkyl radical or a radical -CO-R ¹⁰ , wherein

R ^{10 is} a hydroxyl, a branched or straight-chain C ₁ _30 alkoxy group or an N (R ^a R ^b ) group, wherein

R ^a and R ^{b are the} same or different and / or represent a hydrogen atom, a branched or straight-chain, C} __ 3o-alkyl radical, optionally with carboxy, aminocarbonyl, alkoxycarbonyl or amino groups with up to 20

Carbon atoms is substituted and / or optionally interrupted and / or substituted by one or more heteroatoms from the series 0, N, S.

Compounds according to the invention in which n and m each represent 1 are particularly preferred. 14

of angiotensins, angiotensin analogs, angiotensin derivatives and angiotensin antagonists as well as chemotactic peptides.

In further preferred conjugates according to the invention, the peptides have the following sequences

Cys-Ser-Cys-Ser-Ser-Leu-Met-Asp-Lys-Glu-Cys-Val-Tyr

Phe-Cys-His-Leu-Asp-Ile-Ile-Trp,

Cys-Ser-Cys-Ser-Ser-Trp-Leu-Asp-Lys-Glu-Cys-Val-Tyr-

Phe-Cys-His-Leu-Asp-Ile-Ile-Trp,

Cys-Thr-Cys-Phe-Thr-Tyr-Lys-Asp-Lys-Glu-Cys-Val-Tyr-

Phe-Cys-His-Leu-Asp-Ile-Ile-Trp,

Cys-Ser-Ala-Ser-Ser-Leu-Met-Asp-Lys-Glu-Ala-Val-Tyr-

Phe-Cys-His-Leu-Asp-Ile-Ile-Trp,

I I

Cys-Ser-Cys-Lys-Asp-Met-Thr-Asp-Lys-Glu-Cys-Leu-Asn-

Phe-Cys-Hiε-Gln-Asp-Val-Ile-Trp,

I I

Ala-Ser-Cys-Ser-Ser-Leu-Met-Asp-Lys-Glu-Cys-Val-Tyr- Phe-Ala-His-Leu-Asp-Ile-Ile-Trp, wherein

R ^{10 is} hydroxyl, branched or straight chain

C] __ 3o-alkoxy group or an N (R ^a R ^b ) group, where R ^a and Rb are the same or different and / or represent a hydrogen atom, a branched or straight-chain, C ₁ _3Q-alkyl radical which may be substituted with carboxy, Aminocarbonyl, alkoxycarbonyl or amino groups with up to 20 carbon atoms is substituted and / or optionally interrupted and / or substituted by one or more heteroatoms from the series 0, N, S, is.

The invention further relates to conjugates containing a compound of the general formula (I and / or II) and nucleotides of the DNA and RNA type and which accumulate selectively in diseased tissue

Substances with a convex bond between them and, in the case of substances containing carboxyl or amino groups, such as naturally occurring or modified oligonucleotides in which the degradation is prevented or made more difficult by naturally occurring nucleases, peptides, proteins, antibodies or fragments thereof amidically or is present in ester form in the case of hydroxyl-containing substances such as fatty alcohols or imidically in the case of substances containing aldehyde groups.

Particularly preferred conjugates are characterized in that the substances accumulating in the diseased tissue are peptides such as endotheline, partial sequences of endothelin, endothelin analogues, endothelin derivatives,

Endothelin antagonists or angiotensins, partial sequences 16

Phe-D-Trp-Leu-Asp-Ile-Ile-Trp,

Val-Tyr-Ile-His-Pro-Phe,

Val-Tyr-Ile-His-Pro,

or the cyclic amino acid sequences

Cyclo- (DTrp-DAsp-Pro-DVal-Leu),

Cyclo- (DGlu-Ala-alloDIle-Leu-DTrp).

on

The present invention furthermore further relates to compounds of the general formula (II)

B-CO- (CR ¹ R ² ) _{n = 1 2} -S-CHR ³ -CHR ⁴ -S0 ₂ -NH-CR ⁵ R ⁶ - (CR ⁷ R ⁸ ) _{m = 1 # 2} -SR ⁹ (II)

wherein

R ¹ , R ² , R ³ , R ⁴ and R ^{5 are the} same or different and each represents a hydrogen atom and / or a branched or unbranched C 1-6 alkyl radical,

R ^{6 represents} a hydrogen atom, a branched or unbranched C 1-6 alkyl radical or a radical -CO-R ¹⁰ , wherein

R ^{10 is} a hydroxyl, a branched or straight-chain, cyclic or polycyclic Cχ.30 alkoxy, alkenyloxy, polyalkenyloxy, alkynyloxy, polyalkynyloxy, aryloxy, alkylaryloxy or arylalkyloxy group, which may optionally be combined with hydroxyl, Ala-Ser-Ala-Ser-Ser-Leu-Met-Asp-Lys-Glu-Ala-Val-Tyr- Phe-Ala-His-Leu-Asp-Ile-Ile-Trp,

Cys-Ser-Cys-Ser-Ser-Leu-Met-Asp-Lys-Glu-Cys-Val-Tyr- Phe-Cys-His-Leu-Asp-Ile-Ile-Trp,

Cys-Thr-Cys-Phe-Thr-Tyr-Lys-Asp-Lys-Glu-Ala-Val-Tyr- Phe-Ala-His-Leu-Asp-Ile-Ile-Trp,

I I

Cys-Val-Tyr-Phe-Cys-His-Gln-Asp-Val-Ile-Trp,

N-acetyl-Leu-Met-Asp-Lys-Glu-Ala-Val-Tyr-Phe-Ala-His-Gin Asp-Val-Ile-Trp,

Asp-Arg-Val-Tyr-Ile-His-Pro-Phe-His-Leu,

Ac-Asp-Arg-Val-Tyr-Ile-His-Pro-Phe-His-Leu,

Asp-Arg-Val-Tyr-Ile-His-Pro-Phe,

Arg-Val-Tyr-Ile-His-Pro-Phe,

Arg-Val-Tyr-Ile-His-Pro-Phe-His-Leu,

Sar-Arg-Val-Tyr-Val-His-Pro-Ala,

For-Met-Leu-Phe,

For-Met-Leu-Phe-Lys,

the partial sequences

His-Leu-Asp-Ile-Ile-Trp,

D-Trp-Leu-Asp-Ile-Ile-Trp, 18th

R ^{12 represents} a hydrogen atom, an amino protecting group or a branched or straight-chain, cyclic or polycyclic Cι_3o-alkyl, alkenyl, polyalkenyl, alkynyl, polyalkynyl, aryl, alkylaryl or arylalkyl group, which optionally with hydroxy, oxy , Oxo, carboxy, aminocarbonyl, alkoxycarbonyl, amino, aldehyde or alkoxy groups with up to 20 carbon atoms and / or optionally substituted by one or more heteroatoms from the series O, N, S, P, As, Se is interrupted and / or substituted,

R ^{13 represents} a hydrogen atom or an alcohol protecting group,

whose conjugates with substances that selectively accumulate in diseased tissue, with a covalent bond between them and this in the case of

Substances containing carboxyl or amino groups, such as naturally occurring or modified oligonucleotides, in which the degradation is prevented or made more difficult by naturally occurring nucleases, peptides, proteins, antibodies or their fragments amidically or, in the case of substances containing hydroxyl groups, such as fatty alcohols, ester-like or in the case of aldehyde groups containing substances is imidic

as well as their complexes with radioisotopes of Tc or Re.

The compounds of the general formula (II) according to the invention are prepared by adding 2-chloroethanesulfonic acid chloride optionally substituted with R ³ and R ⁴ in a manner known per se in an aprotic solvent with addition of a Oxy-, oxo-,

Carboxy, aminocarbonyl, alkoxycarbonyl, amino, aldehyde or allkoxy groups with up to 20 carbon atoms and / or optionally interrupted by one or more heteroatoms from the series 0, N, S, P, As, Se and / or is substituted or an N (R ^a R ^b ) group, where R ^a and R ^{b are the} same or different and / or a hydrogen atom, a branched or straight-chain, cyclic or polycyclic Cι_3o-alkyl, alkenyl, polyalkenyl, alkynyl -, Polyalkynyl, aryl, alkylaryl or arylalkyl radical, optionally with hydroxy, oxy, oxo, carboxy, aminocarbonyl, alkoxycarbonyl,

Amino, aldehyde or alkoxy groups with up to 20 carbon atoms is substituted and / or optionally interrupted and / or substituted by one or more heteroatoms from the series 0, N, S, P, As, Se,

R ⁷ and R ^{8 are the} same or different and each represents a hydrogen atom and / or a branched or unbranched Cχ_g alkyl radical,

B for a residue -SR ¹¹ , -NHR ¹² or -OR ¹³ , wherein

R! 1 for a hydrogen atom, for a branched or unbranched Cι_g alkyl radical or for one

Sulfur protection group stands, 20th

wherein R ¹ , R ² , n and B have the meaning given above,

implements

and any protective groups present are split off in a manner known per se.

The invention furthermore relates to kits which are used to produce radiopharmaceuticals, consisting of a compound of the general formula (II) or a conjugate according to the invention containing compounds of the general formula (I and / or II) and substances which accumulate selectively in tissues, a Reducing agents and optionally an auxiliary ligand, which are in the dry state or in solution, as well as instructions for use with a reaction instruction for reacting the described compounds with technetium-99m or Re in the form of a pertechnetate solution or perrhenate solution.

The invention also relates to a radiopharmaceutical composition for the non-invasive in vivo presentation of organs, receptors and receptor-containing tissue and / or of atherosclerotic plaques, which contain a compound of the general formula (I) or a conjugate according to the invention containing compounds of the general formula (I and / or II) and contains substances which accumulate selectively in tissues, if appropriate with the additives customary in galenics, the compound being prepared in a kit with technetium-99m or Re in the form of a pertechnetate or perrhenate solution. suitable base with compounds of the general formula (III)

NH ₂ -CR ⁵ R ⁶ - (CR ⁷ R ⁸ ) _{m = 1 # 2} -SR ⁹

(III)

wherein R ⁵ , R ⁶ , R ⁷ , R ⁸ , R ⁹ and m have the meaning given above,

to compounds of the general formula (IV)

CR ³ = CR ⁴ -S0 ₂ -NH-CR ⁵ R ⁶ - (CR ⁷ R ⁸ ) _{m = 1/2} -SR ⁹

(IV)

wherein R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁷ , R ⁸ , R ⁹ and m have the meaning given above,

implements.

These reactions are carried out in polar and non-polar, aprotic solvents such as

Dichloromethane, tetrahydrofuran, chloroform, 1,4-dioxane, pyridine, DMF or DMSO at temperatures between -40 ° and 120 ° C with the addition of an auxiliary base to trap the acids released. Such auxiliary bases can, for example, tertiary amines, alkali and

Alkaline earth hydroxides, alkali and alkaline earth carbonates.

The resulting compounds of the general formula (IV) are optionally with the addition of a suitable auxiliary base, such as, for example, a tertiary amine, with compounds of the general formula (V)

B-CO- (CR ¹ R ² ) _{n = 1 2} -SH

(V) 22

often adversely affect the labeling yield and radiochemical purity and thus also the background due to non-specifically bound technetium. Sulfur protecting groups are established or split off using methods known to the person skilled in the art. The coupling to substances which accumulate selectively in diseased tissues also takes place according to methods known per se to the person skilled in the art (for example Fritzberg et al.; J. Nucl. Med. 26, 7 (1987)), for example by reacting electrophilic groups of the complex ligand with nuceophilic ones Centers of substances selectively accumulating in diseased tissues or by reaction of nucleophilic groups of the chelator with electrophilic groups of substances selectively accumulating in diseased tissues.

The coupling partners include different biomolecules used. So e.g. Ligands that bind to specific receptors and thus show changes in receptor density, these include Peptides,

Steroid hormones, growth factors and neurotransmitters. Coupling products with steroid hormone receptor-affine substances enable improved diagnosis of breast and prostate carcinomas (S.J. Brandes and J.A. Katzenellenbogen, Nucl .Med.Biol. 15, 53, 1988).

Variously, tumor cells have an altered density of receptors for peptide hormones or growth factors, e.g. the "epidermal growth factor" (EgF). The concentration differences can be used for the selective enrichment of cytostatics in tumor cells (E. Abound-Pirak et al.; Proc.Natl.Acad.Sci. USA 86: 3778 1989). Other biomolecules are metabolites that can be introduced into the metabolism of the cells and indicate a changed metabolism; these include e.g. Fatty acids,

Saccharides, peptides and amino acids. Fatty acids coupled In a method for performing a radio-diagnostic examination, the radiopharmaceutical composition is administered to a patient in an amount of 0.1 to 30 mCi, preferably 0.5 to 10 mCi per 70 kg of body weight, and the radiation emitted by the patient is recorded.

Surprisingly, many of the chelates synthesized and labeled with technetium-99m or Re show a higher stability than comparable N2S2 and N3S systems which are described in the literature. For example, in the case of a substance according to the invention (example 2) which was coupled to a partial endothelin sequence, no decomposition products are observed after 24 h. It was also possible to determine from petition experiments that the Tc-99m or re-chelators described in this invention complex better than the comparable N2S2, N3S and propylene amine oxime systems. The chelates and chelating agents described in the present invention are thus clearly more suitable for diagnostic and therapeutic purposes than the previously known systems. The mild marking conditions are a particular advantage. Thus, after the protective groups have been split off, the ligands according to the invention and their coupling products can be labeled on substances which accumulate selectively in diseased tissues at room temperature and at a physiological pH. The choice of suitable protective groups, which can be split off with different reaction conditions depending on the coupling product, always ensures that undesired side reactions cannot occur during the purification of the coupling products. This ensures that no undesired crosslinking reactions or oxidation of free sulfhydryl groups to form disulfides

Cleaning conditions occur. Such changes 24

It is immaterial whether the described chelating agents are labeled with technetium-99m before or after coupling to the selectively enriching molecule. For coupling to the selectively enriching molecule after complexation, however, it is a prerequisite that the reaction of the radioactive complex with the enriching compound proceeds quickly, under mild conditions and almost quantitatively, so that subsequent purification is not necessary.

The pharmaceutical compositions according to the invention are prepared in a manner known per se by adding the complexing agents according to the invention with the addition of a reducing agent, preferably tin (II) salts such as chloride, pyrophosphate or tartrate - and optionally with the addition of galenics usual additives - dissolves in aqueous medium and then sterile filtered. Suitable additives are, for example, physiologically harmless buffers (e.g. tromethamine), small additions of electrolytes (e.g. sodium chloride), stabilizers (e.g. gluconate, phosphates or phosphonates). The pharmaceutical composition according to the invention is in the form of a solution or in lyophilized form and is added shortly before application with a solution of Tc-99m pertechnetate, eluted from commercially available Mo / Tc generators, or a perrhenate solution.

When used in nuclear medicine in vivo, the pharmaceutical compositions according to the invention are dosed in amounts of 1x10 " ⁵ to 5xl0 ⁴ nmol / kg body weight, preferably in amounts between 1x10 ^{" 3} to 5xl0 ² nmol / kg body weight. Starting from an average body weight of 70 kg, the amount of radioactivity for diagnostic applications is between 0.05 to 50 mCi, preferably 5 to The less stable N2S2 systems have been described in EP-0200492. Other metabolic products such as saccharides, deoxyglucose, lactate and amino acids (leucine, methyl methionine, glycine) were modified with the help of the PET technique for imaging

Metabolism processes are used (R. Weinreich, Swiss Med. 8, 10, 1986). Non-biological substances such as misonidazole and its derivatives, which irreversibly bind to cell components in tissues or tissue parts with a reduced oxygen concentration, can also be used for the specific enrichment of radioactive isotopes and thus for imaging tumors or ischemic regions (ME Shelton, J. Nucl. Med. 30, 351, 1989). Finally, there is also the coupling of the new chelating agents to monoclonal ones

Antibodies or their fragments, polysaccharides such as dextrans or starches, bleomycins, hormones, enzymes, polypeptides such as polylysine and nucleotides of the DNA or RNA type are possible. Coupling products of the chelates according to the invention or their complexes with technetium-99m or Re with endothelines, endothelin derivatives or with partial sequences of the endotheline or their derivatives for the detection of atherosclerotic vascular diseases have proven particularly favorable. These derivatives were applied to WHHL rabbits which, due to a genetic defect in the LDL receptor, have high LDL concentrations in the blood and thus have atherosclerotic lesions. About 1 to 6 h after application of the derivatives in WHHL rabbits, a high accumulation in atherosclerotic plaques could be demonstrated. So far, only very late stages of atherogenesis have been diagnosed with invasive procedures. The compounds according to the invention therefore offer the decisive advantage of diagnosing much earlier stages of atherosclerosis using a non-invasive method. 26

(Silica gel, CH2CI2). 2.37 g of a slowly crystallizing oil remain. Yield: 66% Analysis: calc. : C 50.12 H 5.89 N 3.80 O 22.26 S 17.84 Found: C 49.97 H 6.01 N 3.62 S 17.56

N- {4 - [(N-methyl) carbamoyl1-3-thiabutylsulfonvn-S- (4-methoxybenzyl) cysteine ethyl ester (3) A solution of 3.59 g of vinyl sulfonic acid 2. (10 mmol) and 2 g triethylamine in 25 ml dichloromethane is cooled to 0 ° C. Then 1.05 g of N-methylmercaptoacetamide are added in a little dichloromethane and at RT 24

Hours stirred. The mixture is then diluted with 25 ml of dichloromethane, washed with dilute HCl and water, dried and concentrated.

Yield: 66%

Analysis:

Ber. : C 46.53 H 6.07 N 6.03 O 20.66 S 20.71 Found: C 46.42 H 6.18 N 6.09 S 21.03

N- {4 - [(N-methyl) carbamoyl] -3-thiabutylsulfonyI} - ethyl cysteine (4)

10 ml of HF are condensed into a 100 ml Teflon round-bottom flask at 2.degree. G of 3_ (5 mmol) and a drop of anisole at 0 ° C. with exclusion of moisture. The mixture is stirred at 0 ° C. for 30 min and then hydrogen fluoride is carefully distilled off. The residue is taken up in dichloromethane, washed with sodium bicarbonate solution and water, dried and concentrated. The oily residue is crystallized by trituration with diethyl ether. Yield: 64% Analysis:

Ber. : C 34.87 H 5.85 N 8.13 O 23.22 S 27.93 Found: C 34.55 H 5.91 N 8.42 S 27.37 30 mCi per 70 kg application. Between 5 and 500 mCi, preferably 10 to 350 mCi, are applied for therapeutic applications. The application is usually carried out by intravenous, intraarterial, peritoneal or intertumor injection of 0.1 to 2 ml of a solution of the agent according to the invention. Intravenous administration is preferred.

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

Example 1

S- (4-methoxybenzyl) ystein ethyl ester (1)

HCl is introduced into a solution of 27.7 g of S-4-methoxybenzylcysteine in 250 ml of absolute EtOH until it is saturated and heated to boiling. After the reaction is complete, the mixture is filtered after cooling to room temperature and the mother liquor is concentrated again. 28.4 g of white crystals remain. Yield: 93% Analysis: calc. : C 51.06 H 6.59 N 4.58 O 15.70 S 10.49 opp. : C 50.88 H 6.83 N 4.45 S 10.15

N-vinylsulfonyl-S- (4-methoxybenzyl) cysteine ethyl ester (2) An ice-cooled solution of 3.06 g (10 mmol) of the S-protected cysteine derivative 1 and 1.79 g of chloroethanesulfonyl chloride (11 mmol) in 10 ml dichloromethane dry pyridine (44 mmol) is slowly added while cooling with ice. The mixture is allowed to warm to RT and, after the reaction has ended, 20 ml of dilute HCl are added and the dichloromethane phase is separated off. The aqueous phase is extracted several times with dichloromethane, washed with water, dried, concentrated and chroographed 28

(HOOC-Trp-Ile-Ile-Asp-D-Trp-Phe-Gly) -5-ca hamr.yl -4- methyl-3-thiabutγlsulfonyl} -S- (4-m thoyy-benzy.) - cysteine ethyl ester ( 6)

To a solution of 523 mg of the cysteine derivative 5_ (1 mmol), 101 mg of triethylamine and 115 mg of N-hydroxysuccinimide (1 mmol) in 10 ml of anhydrous dimethylformamide are 211 mg of EDC (1.1 mol) in with stirring at -10 ° C 2 ml of anhydrous dimethylformamide were added dropwise and the mixture was stirred at 0 ° C. for 2 hours. A solution of 967 mg of Phe-D-Trp-Asp-Ile-Ile-Trp (1.1 mmol) in DMF is then added dropwise within 30 minutes. The mixture is first stirred at 0 ° C. for a further 2 hours and stirred at room temperature for 12 hours. The solvent is evaporated in vacuo and taken up in dichloromethane. After renewed filtration, the mixture is washed twice with 0.5 N HCl and saturated sodium hydrogen carbonate solution, dried over magnesium sulfate and the solvent is stripped off. The residue is crystallized by trituration with diethyl ether. Yield: 29% Analysis:

Ber. : C 58.16 H 6.27 N 10.12 O 18.50 S 6.95 Found: C 57.78 H 6.42 N 10.03 S 7.06

N- r (HOOC-Trp-Ile-Ile-Asp-D-Trp-Phe-Glv) -carbamoγl-4-methyl-3-thiabutylsulfonyll -cysteine ethyl ester (7)

10 ml of HF are condensed into a 100 ml round-bottom flask at 1.degree. The mixture is stirred at 0 ° C. for 30 min and then hydrogen fluoride is carefully distilled off. The residue is crystallized by trituration with diethyl ether. Yield: 39% Analysis:

Calc .: C 56.09 H 6.22 N 11.09 0 18.99 S 7.61 Found: C 56.19 H 6.46 N 11.16 S 7.95 N- .4-f (N-methyl) carbamoyl] -3-thiabutylaul fonyl} - ethyl cysteine. Technetium-99m complex 10 mg of compound 4. are dissolved in 1.0 ml of ethanol. 50 ul of this ligand solution are mixed with 100 ul ethanol, 150 ul phosphate buffer pH 8.5, 50 ul of a deoxygenated aqueous citrate solution (50 mg / ml), 2.5 ul of a deoxygenated tin (II) chloride solution (5 mg / ml 0.1 N HCl) and 100 μl of a pertechnetate solution (400-1000 μCi) are added. After an incubation time of 10 min, the reaction mixture is examined for the purity of the Tc complex formed by means of HPLC: LiChrospher RP-18 column, 5 μ, 125 × 4.6 mm; Gradient elution from 100% A to 100% B within 15 min (eluent A: acetonitrile / sodium phosphate 5 mM, pH 2.0 (10/90); eluent B: acetonitrile / sodium phosphate 5 mM, pH 2.0 (75/25);

1 ml / min. The radiochemical purity is> 98%.

Example 2

N- {4 - [(N-Glycyl) carbamoyl] -4-methyl-3-thiabutylsulfonyl} - S- (4-methoxγ-benzγl) cysteine ethyl ester (5) A solution of 3.59 g of vinylsulfonic acid 2. (10 mmol) and 2 g triethylamine in 25 ml THF is cooled to 0 ° C. This is followed by 1.63 g of mercaptopropionylglycine

(10 mmol) in a little THF and stirred for 24 hours at RT. The solvent is then stripped off, the residue is taken up in 50 ml of dichloromethane, the organic phase is washed with dilute HCl and water, dried and concentrated and chromatographed (silica gel, CH ₂ C1 ₂ ). Yield: 56% Analysis: calc. : C 45.96 H 5.79 N 5.36 0 24.49 S 18.41 Found: C 45.61 H 5.90 N 5.44 S 18.07 30th

Analysis:

Ber. : C 58.31 H 6.36 N 3.40 0 15.54 S 7.78

Found: C 57.89 H 6.66 N 3.21 S 7.47

N-vinylsulfonyl-S-bis (4-methoxyphenyl) methyl cysteine ethyl ester (9)

An ice-cooled solution of 4.12 g (10 mmol) of the S-protected cysteine derivative £ and 1.79 g of chloroethanesulfonyl chloride (11 mmol) in 10 ml of dichloromethane is slowly mixed with dry pyridine (44 mmol) while cooling with ice. The mixture is allowed to warm to RT and, after the reaction has ended, 20 ml of dilute HCl are added and the dichloromethane phase is separated off. The aqueous phase is extracted several times with dichloromethane, washed with water, dried, concentrated and chromatographed (silica gel, CH ₂ C1 ₂ ). Yield: 66% Analysis: calc. : C 56.76 H 5.85 N 3.01 0 20.62 S 13.78 Found: C 56.81 H 5.70 N 2.89 S 14.02

N- (4 - [(N-Glycyl) carbamoyll-4-methyl-3-thiabuty] sulfonyl} - S-bis (4-rn thoxyphenγl) methvl-cysteine ethyl ester (10) A solution of 4.66 g of vinylsulfonic acid 2 , (10 mmol) and 2 g of triethylamine in 25 ml of dichloromethane is cooled to 0 ° C. Then 1.63 g of mercaptopropionylglycine are added and the mixture is stirred for 24 hours at RT, followed by dilution with 25 ml of dichloromethane, with dilute HCl and water washed, dried, concentrated and chromatographed (silica gel, CH2Cl2 / Me0H). Yield: 45% analysis:

Ber. : C 51.58 H 5.77 N 4.46 O 22.90 S 15.30 Found: C 51.11 H 6.05 N 4.49 S 14.98 N- r (HQQC-Trp-Ile-Ile-Asp-D-Trp-Phe-Glv) -carbamπy -4-methyl-3-thiabutylsulfonyll -cysteine ethyl ester. Technetium-99m complex

10 mg of compound 2 are dissolved in 1.0 ml of ethanol. 50 μl of this ligand solution are mixed with 250 μl phosphate buffer pH 8.5, 50 μl of a deoxygenated aqueous citrate solution (50 mg / ml), 2.5 μl of a deoxygenated tin (II) chloride solution (5 mg / ml 0.1 N HCl) and 100 μl of a pertechnetate solution (400-1000 μCi) were added. After an incubation time of 10 min, the reaction mixture is examined for the purity of the Tc complex formed by means of HPLC: LiChrospher RP-18 column, 5 μ, 125 × 4.6 mm; Gradient elution from 100% A to 100% B within 15 min (eluent A: acetonitrile / sodium phosphate 5 mM, pH 2.0 (10/90); eluent B: acetonitrile / sodium phosphate 5 mM, pH 2.0 (75/25); 1 ml / min. The radiochemical purity is> 96%.

Example 3

S-bis (4-methoxyphenyl) ethyl cysteine ethyl ester (8) 1.86 g of anhydrous cysteine ethyl ester hydrochloride (10 mmol) are suspended in 10 ml of glacial acetic acid and about 2.76 g of S-bis (4-methoxyphenyl) - methanol (15 mmol) and 2.1 ml of BF _ß -diethyl etherate (15 mmol) were added. The mixture is stirred at RT for 2 h, with almost everything clearly dissolving. The mixture is then concentrated on a rotary evaporator at a bath temperature of 40 ° C. The oily residue is dissolved in ethyl acetate. By shaking with saturated

Sodium acetate solution precipitates the protected cysteine derivative, which is suctioned off and washed well with water and acetone. Yield: 79% 32

Example 4

N- (4-Carboxv-3-thiabutγlsulfonvl. -S-bi (4-methoxypheny]) - methyl cysteine ethyl ester (12) To a stirred solution of 920 mg mercaptoacetic acid (10 mmol) and 1.5 g Triton B solution 5.6 mg of the vinyl sulfonic acid 9_ (1.2 mmol) were added with cooling and the mixture was stirred at RT for 20 h, then acidified with 1 N HCl and extracted with ethyl acetate, and the combined organic phases were saturated

Washed saline, dried and concentrated. The remaining residue is recrystallized from Me0H / CH2Cl2. Yield: 61% Analysis:

Found: C 51.69 H 5.60 N 2.51 0 22.95 S 17.25. Found: C 51.28 H 5.90 N 2.27 S 17.86

N- {4-Carboxy-3-thiabutylsulfonvl} cysteine ethyl ester (13) 558 mg of the protected S-compound 12 (1 mmol) and a trace of anisole are added to 10 ml of trifluoroacetic acid at RT and a trace of anisole is added and the mixture is stirred at 50 ° C. for 2 hours. After finished

The trifluoroacetic acid is removed in vacuo, the residue is taken up in ethyl acetate, washed with saturated sodium chloride solution, dried and concentrated. The oily residue is crystallized by trituration with diethyl ether.

Yield: 59%

Analysis: Calc .: C 32.62 H 5.17 N 4.23 O 28.97 S 29.03

Found: C 32.75 H 5.28 N 4.45 S 29.67

N- carboxy-3-thiabutylsulfone. -cysteine ethyl ester. Technetium-9m complex 10 mg of compound 12 are dissolved in 1.0 ml of ethanol. 50 μl of this ligand solution are mixed with 100 μl ethanol, N- {4-f (N-Glycyl) carbamoyl] -4-methyl-3-thiabutylsulfnnyl} - S-cysteine ethyl ester (11)

At RT 628 mg of the protected S-compound iQ. Are added to 10 ml of trifluoroacetic acid. (1 mmol) and a trace of anisole and stirred at 50 ° C for 2 hours. After finished

The trifluoroacetic acid is removed in vacuo, the residue is taken up in ethyl acetate, washed with saturated sodium chloride solution, dried and concentrated. The oily residue is crystallized by trituration with diethyl ether. Yield: 34% Analysis:

Ber. : C 35.81 H 5.51 N 6.96 0 27.83 S 23.90 Found: C 35.32 H 5.84 N 6.63 S 23.95

N- {4- [(N-Glycyl) carbamoyl] -4-methyl-3-thiabutylsulfonyl} - S-cysteine ethyl ester. Technetium-99m-Kompl x 10 mg of compound i are dissolved in 1.0 ml of ethanol. 50 ul of this ligand solution are mixed with 100 ul ethanol, 150 ul phosphate buffer pH 8.5, 50 ul of a deoxygenated aqueous citrate solution (50 mg / ml), 2.5 ul of a deoxygenated tin (II) chloride solution (5 mg / ml 0.1 N HCl) and 100 μl of a pertechnetate solution (400-1000 μCi) are added. After an incubation time of 10 min, the reaction mixture is examined for the purity of the Tc complex formed by means of HPLC: LiChrospher RP-18 column, 5 μ, 125 × 4.6 mm; Gradient elution from 100% A to 100% B within 15 min (eluent A: acetonitrile / sodium phosphate 5 mM, pH 2.0 (10/90); eluent B.- acetonitrile / sodium phosphate 5 mM, pH 2, 0 (75/25); lml / min. The radiochemical purity is> 95%. 34

211 mg EDC (1.1 mol) in 5 ml of anhydrous dimethylformamide were added dropwise with stirring at -10 ° C. and the mixture was stirred at 0 ° C. for 2 hours. Then a solution of 572 mg of the cysteine derivative 2Λ (1 mmol) in DMF is added dropwise within 30 minutes. The mixture is first stirred at 0 ° C. for a further 2 hours and stirred at room temperature for 12 hours. The product is filtered off from the N, N'-dicyclohexylurea and the filtrate is evaporated in vacuo and taken up in dichloromethane. After renewed filtration, the mixture is washed twice with 0.5 N HCl and saturated sodium hydrogen carbonate solution, dried over magnesium sulfate and the solvent is stripped off. The residue is crystallized by trituration with diethyl ether. Yield: 39% Analysis:

Ber. : C 54, 53 H 6, 30 N 8, 48 0 17, 76 S 12, 94

Gef. : C 54, 21 H 6, 49 N 8, 53 S 12, 75

For-Met-Leu-Phe- {{N-TN '- (5-amino- thiapentylsulfonyl) - cysteinyl -2-aminoethyl} amide} (16)

991 mg of the peptide 1 pound (1 mmol) is treated for 45 minutes at 0 ° C. with 10 ml of anhydrous HF in the presence of 5 ml of anisole and 3.5 ml of diethyl sulfide. After evaporating the

Acid, the remaining residue is taken up in 5% acetic acid, washed several times with diethyl ether and lyophilized. Chromatographic purification via Sephadex

G-10 with 0.2 M acetic acid gives an oil.

Yield: 29%

Analysis: Calculated: C 47.10 H 6.32 N 10.99 0 18.82 S 16.77

Ge f. : C 46, 81 H 6, 57 N 11, 25 S 16, 81

For-Met-Leu-Phe-f {N-FN '- (5-amino-3-thiapentvlsulfonyl) - cysteinyll 2-aminoethvllamidl. Technetium-99m-Kormolex 10 mg of the compound if_ are dissolved in 1.0 ml of ethanol. 50 μl of this ligand solution are mixed with 100 μl ethanol, 150 ul phosphate buffer pH 8, 5, 50 ul of a deoxygenated aqueous citrate solution (50 mg / ml), 2.5 ul of a deoxygenated tin (II) chloride solution (5 mg / ml 0.1 N HCl) and 100 ul one Pertechnetate solution (400-1000 μCi) added. The reaction mixture is after a

Incubation time of 10 min examined by HPLC for the purity of the Tc complex formed: LiChrospher RP-18 column, 5 μ, 125 x 4.6 mm; Gradient elution from 100% A to 100% B within 15 min (eluent A: acetonitrile / sodium phosphate 5 mM, pH 2.0 (10/90); eluent B: acetonitrile / sodium phosphate 5 mM, pH 2.0 (75/25); 1 ml / min. The radiochemical purity is> 97%.

Example 5

N- (4-carboxy-3-thiabutylsulfonyl} -S-bis (4-methoxyphenyl) - methyl-cysteine- .N- (2-aminoethyl) amidi (14)

A solution of 5.58 g of the cysteine ester 12 (10 mmol) in 50 ml of toluene / dioxane is slowly added dropwise at 95 ° C. to a solution of 25 g of ethylenediamine (240 mmol) and the mixture is boiled under reflux for 2 hours. After cooling to room temperature, excess ethylenediamine is distilled off in vacuo and the residue is recrystallized from methanol / CH Cl2. Yield: 80% Analysis:

Ber. : C 50.42 H 5.82 N 7.35 0 19.59 S 16.83 Found: C 51.01 H 5.99 N 7.41 S 16.56

For-Met-LF.u-Phe-.N-TN- (4-carboxy- thiabutylsulfonyl) - £ - bis (4-methoxyphenyl) methylcysteinyl -2-aminoethyl} amiri

(15)

To a solution of 622 mg For-Met-Leu-Phe (1.1 mmol), 101 mg triethylamine and 115 mg N-hydroxysuccinimide

(1.0 mmol) in 10 ml of anhydrous dimethylformamide 36

0 ° C stirred for 6 h. The mixture is allowed to warm to RT and, after the reaction has ended, dilute HCl is added and the dichloromethane phase is separated off. The aqueous phase is extracted several times with dichloromethane, washed with water, dried and concentrated. Yield: 58% Analysis:

Ber. : C 64.22 H 5.40 N 3.00 0 13.69 S 13.72 Found: C 64.02 H 5.57 N 3.09 S 13.52

N- (4-carboxy-3-thiabutylsulfonvll-S-trityl-cyst .nmethyl - ester (19)

To a stirred solution of 920 mg of mercaptoacetic acid (10 mmol) and 1.5 g of Triton B solution, 4.68 g of the vinylsulfonic acid (10 mmol) are added with cooling and the mixture is stirred at RT for 20 h. It is then acidified with 1N HCL and extracted with ethyl acetate. The combined organic phases are washed with saturated sodium chloride solution, dried and concentrated. The remaining residue is recrystallized from MeOH / CH2Cl2. Yield: 54% Analysis:

Fitting : C 57, 94 H 5, 22 N 2, 50 O 17, 15 S 17, 19

Gef. : C 57, 43 H 5, 40 N 2, 41 S 17, 58

N- {4- rHOOC-Leu-His-Phe-Pro-His-Ile-Tyr-Val-Arσ-NH-carbonyll -3-thiabutylsulfonyl. -S-trityl-cvstein- methyl ester (20) To a solution of 560 mg of acid ü (1 mmol), 280 μl triethylamine and 115 mg N-hydroxysuccinimide (1.0 mmol) in 10 ml of anhydrous dimethylformamide are stirred with - 10 ° C. 211 mg of EDC (1.1 mol) in 5 ml of anhydrous dimethylformamide were added dropwise and the mixture was stirred at 0 ° C. for 2 hours. A solution of 1.18 g of H2N-Arg-Val-Tyr-Ile-His-Por-Phe-His-Leu-COOH (1 mmol) in DMF is then added dropwise within 30 minutes. It will be first 150 ul phosphate buffer pH 8, 5, 50 ul of a deoxygenated aqueous citrate solution (50 mg / ml), 2.5 ul of a deoxygenated tin (II) chloride solution (5 mg / ml 0.1 N HCl) and 100 ul one Pertechnetate solution (400-1000 μCi) added. The reaction mixture is after a

Incubation time of 10 min examined by HPLC for the purity of the Tc complex formed: LiChrospher RP-18 column, 5 μ, 125 x 4.6 mm; Gradient elution from 100% A to 100% B within 15 min (eluent A: acetonitrile / sodium phosphate 5 mM, pH 2.0 (10/90); eluent B: acetonitrile / sodium phosphate 5 mM, pH 2.0 (75/25); 1 ml / min. The radiochemical purity is> 95%.

Example 6

S-tritylcysteine methyl ester (17)

The solution of 2.79 g of triphenylchloromethane in DMF is slowly added dropwise to a solution of 1.71 g of the cysteine methyl ester (10 mmol) and triethylamine (10 mmol) in DMF and the mixture is stirred at RT for 12 hours. Water is then added, the mixture is made weakly alkaline with saturated sodium hydrogen carbonate solution and extracted with dichloromethane. The organic phase is dried and concentrated. The remaining residue is chromatographed (silica gel, CH2CI2). Yield: 67% Analysis: calc. : C 66.73 H 5.84 N 3.38 0 7.73 S 7.75 Found: C 66.46 H 5.96 N 3.44 S 7.59

N-vinylsulfonyl-S-trityl-cysteine methyl ester (18) An ice-cooled solution of 8.28 g (20 mmol) of the S-trityl cysteine derivative 12th and 4.89 g of chloroethanesulfonyl chloride (30 mmol) in 50 ml of dichloromethane is slowly added

Ice cooling with 20 ml of dry pyridine and 38 offset. After an incubation time of 10 min, the reaction mixture is examined for the purity of the Tc complex formed by means of HPLC: LiChrospher RP-18 column, 5 μ, 125 × 4.6 mm; Gradient elution from 100% A to 100% B within 15 min (eluent A: acetonitrile / sodium phosphate 5 mM, pH 2.0 (10/90); eluent B: acetonitrile / sodium phosphate 5 mM, pH 2.0 (75/25); 1 ml / min. The radiochemical purity is> 96%.

stirred for a further 2 hours at 0 ° C. and stirred for 12 hours at room temperature. The product is filtered off from the urea and the filtrate is evaporated in vacuo and taken up in dichloromethane. After renewed filtration, the mixture is washed twice with 0.5 N HCl and saturated sodium hydrogen carbonate solution, dried over magnesium sulfate and the solvent is stripped off. The residue is crystallized by trituration with diethyl ether. Yield: 28% Analysis:

Ber. : C 59.25 H 6.49 N 13.82 0 14.86 S 5.58 Found: C 59.47 H 6.73 N 13.57 S 5.66

N- {4- rHOOC-Leu-His-Phe-Pro-His-Tle-Tyr-Val-Arσ-NH-carbonyll -3-thiabutylsulfonyl1-cysteine methyl ester (21) 1.72 g of the peptide 2SL (1 mmol) Treated for 45 minutes at 0 ° C with 20 ml of anhydrous HF in the presence of 5 ml of anisole and 3.5 ml of diethyl sulfide. After the acid has been evaporated off, the remaining residue is taken up in 5% acetic acid, washed several times with diethyl ether and lyophilized. Chromatographic purification on Sephadex G-10 with 0.2 M acetic acid gives 252 mg of an oil. Yield: 17% Analysis: calc. : C 53.53 H 6.60 N 16.08 0 17.29 S 6.50 Found: C 53.12 H 6.86 N 15.78 S 6.33

W- {4- rHOOC-T.P. -His-Phe-Pro-His-Ile-Tyr-Val-Arσ-NH- carbonyll -3- hiabutylsul fonyll-cysteine ethyl ester. Technetium-99m complex

10 mg of compound 21 are dissolved in 1.0 ml of ethanol. 50 ul of this ligand solution are mixed with 100 ul ethanol, 150 ul phosphate buffer pH 8.5, 50 ul of a deoxygenated aqueous citrate solution (50 mg / ml), 2.5 ul of a deoxygenated tin (II) chloride solution (5 mg / ml 0.1 N

HCl) and 100 μl of a pertechnetate solution (400-1000 μCi) 40

20 carbon atoms and / or is optionally interrupted and / or substituted by one or more heteroatoms from the series O, N, S, P, As, Se or an N (R ^a R ^) group, where R ^a and R ^ are the same or different and / or a hydrogen atom, a branched or straight-chain, cyclic or polycyclic Cι_3o-alkyl, alkenyl, polyalkenyl, alkynyl, polyalkynyl, aryl,

Represent alkylaryl or arylalkyl, which is optionally substituted with hydroxy, oxy, oxo, carboxy, aminocarbonyl, alkoxycarbonyl, amino, aldehyde or alkoxy groups having up to 20 carbon atoms and / or optionally by one or more heteroatoms from the series 0, N, S, P, As, Se is interrupted and / or substituted, stands,

B for a residue -SR ¹¹ , -NHR ¹² or -OR ¹³ , wherein

R ^{11 represents} a hydrogen atom, a branched or unbranched C 1-6 alkyl radical or a sulfur protecting group,

Claims

claims

1. Compounds of the general formula (I)

M - L (I)

wherein

M for a radioisotope of Tc or Re and L for a ligand of the general formula (II)

B-CO- {C ~. ¹ ~. ² ) _{TIB 1 I 2} -S-CHR ³ -CHR ⁴ -S0 ₂ -NH-CR ⁵ R ⁶ - (CR ⁷ R ⁸ ) _{jn = 1 2} -SR ⁹

(II)

means what

R ^ for a hydrogen atom, for a branched or unbranched Cι_g alkyl radical or a radical -CO-R ¹⁰ , wherein

R ¹ ^ is a hydroxyl, a branched or straight-chain, cyclic or polycyclic C ₁ _3 _Q - alkoxy, alkenyloxy, polyalkenyloxy, alkynyloxy, polyalkynyloxy, aryloxy,

Alkylaryloxy or arylalkyloxy group, optionally with hydroxy, oxy, oxo, carboxy, aminocarbonyl, alkoxycarbonyl, amino, aldehyde or allkoxy groups with up to 42

6. Ligands of the general formula (II)

^B - ^CO - ^(C R ¹ R ²⁾ _{n = lf2} - ^S - ^CHR3 - ^CHR4 - ^S0 ₂ - ^NH - ^C R ^5R6 - ^(CR7 R ⁸⁾ --_ _{!: Lf} 2- ^SR9 (II)

wherein R ¹ , R ² , R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁷ , R ^β , R9. _n , m and B each have the meaning given in claim 1.

7. Ligand according to claim 6, characterized in that n and m each represent 1.

8. Ligands according to claim 6, characterized in that R ¹ , R ³ , R ⁴ , R ⁷ and R ^{8 represent} hydrogen atoms.

9. Ligand according to claim 6, characterized in that R ¹ and R ⁵ each represent hydrogen atoms.

10. Ligand according to claim 6, characterized in that B for a radical NHR ¹² or OR ¹³ , wherein

R ¹² and R ^{13 have} the meaning given under claim 1.

11. Conjugates containing a compound of the general formula (I and / or II) and substances which accumulate selectively in diseased tissue, there being a covalent bond between them and, in the case of substances containing carboxyl or amino groups, such as naturally occurring or modified oligonucleotides in which the degradation is prevented or made difficult by naturally occurring nucleases, peptides, proteins, R ¹² for a hydrogen atom, an amino protecting group or a branched or straight-chain, cyclic or polycyclic CI_3 _Q - alkyl, alkenyl, polyalkenyl, alkynyl,

Polyalkynyl, aryl, alkylaryl or arylalkyl group which is optionally substituted with hydroxyl, oxy, oxo, carboxy, aminocarbonyl, alkoxycarbonyl, amino, aldehyde or alkoxy groups having up to 20 carbon atoms and / or is optionally interrupted and / or substituted by one or more heteroatoms from the series O, N, S, P, As, Se,

R ^{13 represents} a hydrogen atom or an alcohol protecting group.

2. Compounds according to claim 1, characterized in that n and m each represent 1.

3. Compounds according to claim 1 or 2, characterized in that R ³ , R, R ⁷ and R ^{8 represent} hydrogen atoms.

4. Compounds according to claim 3, characterized in that R ¹ and R ^{5 are} hydrogen atoms.

5. Compounds according to claim 3, characterized in that B for a radical NHR ¹² or OR ¹³ , wherein

R ¹² and R ^{13 have} the meaning given under claim 1. 44

Cys-Ser-Cys-Lys-Asp-Met-Thr-Asp-Lys-Glu-Cys-Leu-Asn-

Phe-Cys-His-Gln-Asp-Val-Ile-Trp,

Ala-Ser-Cys-Ser-Ser-Leu-Met-Asp-Lys-Glu-Cys-Val-Tyr- Phe-Ala-His-Leu-Asp-Ile-Ile-Trp,

Ala-Ser-Ala-Ser-Ser-Leu-Met-Asp-Lys-Glu-Ala-Val-Tyr- Phe-Ala-His-Leu-Asp-Ile-Ile-Trp,

l 1

Cys-Val-Tyr-Phe-Cys-His-Gln-Asp-Val-Ile-Trp,

N-acetyl-Leu-Met-Asp-Lys-Glu-Ala-Val-Tyr-Phe-Ala-His-Gln-Asp-Val-Ile-Trp,

Asp-Arg-Val-Tyr-Ile-His-Pro-Phe-His-Leu,

Ac-Asp-Arg-Val-Tyr-Ile-His-Pro-Phe-His-Leu,

Asp-Arg-Val-Tyr-Ile-His-Pro-Phe,

Arg-Val-Tyr-Ile-His-Pro-Phe,

Arg-Val-Tyr-Ile-His-Pro-Phe-His-Leu,

Sar-Arg-Val-Tyr-Val-His-Pro-Ala, Antibodies or their fragments are amidic or, in the case of substances containing hydroxyl groups such as fatty alcohols, ester-like or imidic in the case of substances containing aldehyde groups.

12. Conjugates according to claim 11, characterized in that the substances accumulating in diseased tissue are peptides such as endotheline, partial sequences of endothelin, endothelin analogues, endothelin.

Derivatives, endothelin antagonists or angiotensins, partial sequences of angiotensins, angiotensin analogs, angiotensin derivatives and angiotensin antagonists and chemotactic peptides mean.

13. Conjugates according to claim 11, characterized in that the peptides have the following sequences or parts thereof I Cys-Ser-Cys-Ser-Ser-Leu-Met-Asp-Lys-Glu-Cys-Val-Tyr

I.

Phe-Cys-His-Leu-Asp-Ile-Ile-Trp,

Cys-Ser-Cys-Ser-Ser-Trp-Leu-Asp-Lys-Glu-Cys-Val-Tyr-

Phe-Cys-His-Leu-Asp-Ile-Ile-Trp,

l i Cys-Thr-Cys-Phe-Thr-Tyr-Lys-Asp-Lys-Glu-Cys-Val-Tyr-

¹ 1

Phe-Cys-His-Leu-Asp-Ile-Ile-Trp,

Cys-Ser-Ala-Ser-Ser-Leu-Met-Asp-Lys-Glu-Ala-Val-Tyr- ι_ ι Phe-Cys-His-Leu-Asp-Ile-Ile-Trp, 46 in which R ¹ , R ² , R ³ , R ⁴ , R5, R6, R ⁷ , R8 _# R9, _{nm and B have} the meaning given in claim 1,

implements.

15. A process for the preparation of ligands of the general formula (II), characterized in that 2-chloroethane-sulfonyl chloride in a manner known per se in a protic or aprotic solvent with the addition of a suitable base with compounds of the general formula (III)

NH ₂ -CR ⁵ R ⁶ - (CR ⁷ R ⁸ ) _{m =} χ, ₂ "SR ⁹

(III)

wherein R ⁵ , R ⁶ , R ⁷ , R ⁸ , R ⁹ and m have the meaning given in claim 1,

at temperatures from -20 ° to 180 ° C to compounds of the general formula (IV)

CR ³ = CR ⁴ -S0 ₂ -NH-CR ⁵ R ⁶ - (CR ⁷ R ⁸ ) _{m = lf 2} -SR ⁹

(IV)

wherein R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁷ , R ⁸ , R ⁹ and m the in

Have the meaning given,

implements

and these compounds of the general formula (IV) optionally with the addition of a suitable auxiliary base at temperatures from -20 ° to 180 ° C in a manner known per se with compounds of the general formula (V) For-Met-Leu-Phe,

For-Met-Leu-Phe-Lys,

the partial sequences

His-Leu-Asp-Ile-Ile-Trp,

D-Trp-Leu-Asp-Ile-Ile-Trp,

Phe-D-Trp-Leu-Asp-Ile-Ile-Trp,

Val-Tyr-Ile-His-Pro-Phe,

Val-Tyr-Ile-His-Pro,

or the cyclic amino acid sequences

Cyclo- (DTrp-DAsp-Pro-DVal-Leu),

Cyclo- (DGlu-Ala-alloDIle-Leu-DTrp).

exhibit.

14. A process for the preparation of a compound of general formula (I), characterized in that technetium-99m or Re in the form of pertechnetate or perrhenate in the presence of a reducing agent and optionally an auxiliary ligand with a compound of general formula (II)

B-CO- (CR ¹ R ² ) _{n = 1 2} -S-CHR ³ -CHR ⁴ -S0 ₂ -NH-CR ⁵ R ⁶ - (CR ⁷ R ⁸ ) _{m = 1/2} -SR ⁹

(II) 48

a pertechnetate or perrhenate solution is prepared.

18. A method for radiodiagnostic examination, characterized in that a radiopharmaceutical composition according to claim 17 in an amount of 0.1 to 30 mCi, preferably from 0.5 to 10 mCi per 70 kg body weight is administered to a patient and the radiation emitted by the patient is recorded becomes.

B-CO- (CR ¹ R ² ) -. _{= lf 2} -SH

(V)

wherein R ¹ , R, n and B have the meaning given in claim 1,

implements

and any protective groups present are split off in a manner known per se.

16. Kit for the production of radiopharmaceuticals, consisting of a compound of general formula (II) according to one of claims 6 to 10 or a conjugate according to one of claims 11 to 13 and a reducing agent and optionally an auxiliary ligand, which is in the dry state or in solution are available, as well as instructions for use with a reaction instruction for the implementation of the described compounds with Technetium-99m or Re in the form of a pertechnetate solution or perrhenate solution.

17. Radiopharmaceutical composition for the non-invasive in vivo presentation of organs, receptors and receptor-containing tissue and / or of atherosclerotic plaques, characterized in that it contains a compound according to one of claims 1 to 5 or a conjugate according to claims 11 to 13 and optionally contains the additives customary in galenics, the compound being in a kit according to claim 16 with technetium-99m or Re in the form