EP0929570A1

EP0929570A1 - Modified cytostatic agents

Info

Publication number: EP0929570A1
Application number: EP97910326A
Authority: EP
Inventors: Hans-Georg Lerchen; Karsten Von Dem Bruch; Jörg Baumgarten; Michael Sperzel
Original assignee: Bayer AG
Current assignee: Bayer AG
Priority date: 1996-10-04
Filing date: 1997-09-22
Publication date: 1999-07-21
Also published as: WO1998015571A1; US20020173468A1; DE19640970A1; AU4776697A; JP2001502308A

Abstract

The invention concerns conjugates of cytostatic agents and N-thiocarbonyl-modified amino acids and peptides. The invention further concerns processes for their preparation and their use as medicaments, in particular in connection with cancers.

Description

Modified cytostatics

The present invention relates to conjugates of cytostatics and N-thiocarbonyl-modified amino acids or peptides, processes for their preparation and their use as medicaments, in particular in connection with cancer.

Chemotherapy for tumor diseases is accompanied by mostly serious side effects due to the toxicity of chemotherapeutic agents on proliferating cells of other tissues. Have been busy for many years

Scientists with the problem of improving the selectivity of active substances used. A frequently pursued approach is the synthesis of prodrugs which are more or less selective in the target tissue, for example by changing the pH (for example Tietze et al., DE 4 229 903), by enzymes (for example glucuronidases; Jacquesy et al., EP 511 917; Bosslet et al., EP 595 133) or by

Antibody-enzyme conjugates (Bagshawe et al., WO 88/07378; Senter et al., US Pat. No. 4,975,278; Bosslet et al., EP 595 133) are released. Problems with these approaches include the lack of stability of the conjugates in other tissues and organs and in particular the ubiquitous distribution of active substances, which follows the extracellular release of active substances in the tumor tissue.

In the following, three cytostatically active basic bodies from different substance classes, which have serious side effects, are presented as examples.

The heterocyclic amine batracylin (1) shows a good antitumor effect in various colon cancer models (US Pat. No. 4,757,072).

(1)

Peptide conjugates of (1) with good in vitro activity and more favorable solubility properties (US Pat. No. 4,180,343) are less tolerable in animal experiments than batracylin itself. For example, the fucose described in EP 501 250 Conjugates very strongly in the liver on glycoconjugates of cytostatics, as described in our application PCT / 96/01279, which is also pending, have more favorable properties, but are synthetically accessible only with great effort

The quinolone-a (2) 7 - [(3aRS, 4RS, 7aSR) -4- amino- 1,3,3 a, 4,7,7a-hexahy dro-i so-indol-2-yl] -8- chloro-l-cyclopropyl-6-fluoro-l, 4-dihydro-4-oxo-3-quinolinecarboxylic acid shows not only an excellent anti-bacterial activity but also a very good effectiveness against different tumor cell lines (EP 520 240, JP 4 253 973) However, there are considerable toxicological problems (e.g. genotoxicity, bone marrow toxicity, high acute toxicity in vivo etc.)

(2) (Quinolone-a)

20 (S) -Camptothecιn (3) is a pentacyclic alkaloid that was isolated by Wall et al in 1966 (J Amer Chem Soc 88 (1966) 3888). It has a high antitumor activity potential in numerous in vitro and in -vivo tests Unfortunately, the realization of the promising potential in the clinic failed due to toxicity and solubility problems

By opening the E-ring lactone and forming the sodium salt, a water-soluble compound was obtained which is in a pH-dependent equilibrium with the ring-closed form. Clinical studies have so far not been successful here either

NaOH

About 20 years later it was found that the biological activity is due to an enzyme inhibition of topoisomerase I. Since then, the research activities have been intensified again in order to find more contractual and in vivo effective camptothecin derivatives

In order to improve the lack of water, salts of A-ring and

B-ring-modified camptothecin derivatives and of 20-O-acyl derivatives with ionizable groups are described (Vishnuvajjala et al, US 4,943,579). The latter prodrug concept was later also applied to modified camptothecin derivatives (Wani et al, WO 96/02546) The 20-O-acyl prodrugs described, however, have a very short half-life in vivo and are split very quickly into the basic body

We have now found that the modification of cytostatics such as Batracyhn, anti-tumor-active quinolones (such as quinolone-a) or camptothecin or camptothecin derivatives with N-thiocarbonyl-modified amino acids to new compounds with surprising, highly interesting Properties leads

The conjugates obtained in this way are easily accessible synthetically and show in vitro a similarly high activity towards different tumor cell lines and tumor xenografts as the underlying toxophore

Depending on the composition of the N-thiocarbonyl-modified amino acids, the conjugates according to the invention show significantly improved solubility properties compared to the underlying cytostatics

They have a higher tolerance and tumor selectivity than the underlying toxophores

- In vivo they show good to very good therapeutic activity

They are much more stable in extracellular medium and in blood than the pure amino acid prodrugs from Batracyhn, quinolons or from camptothecin derivatives described above In the case of 20-O-acylation of camptothecin derivatives, the ester-like linkage of the carrier residues with the 20-hydroxy group stabilizes the lactone ring which is important for the action

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

S

II

(Ar-X-NH-C-) _n MC

wherein

^ ⁿ for 1 to n 'identical or different groups

S

II

Ar- ^λ -NH-C is where n is a number 1 to n 'and n' is the maximum

Corresponds to the number of possible connecting points of M,

wherein

Ar stands for an aryl radical with up to 10 carbon atoms, which, in addition to X, can optionally be mono- or polysubstituted by alkyl with up to 6 carbon atoms, alkoxy with up to 6 carbon atoms, alkoxycarbonyl with up to 6 carbon atoms, hydroxy, carboxy, carboxyalkyl with up to 6 carbon atoms, cyano, nitro, isocyanato,

Isothiocyanato, halogen, sulfonyl and / or sulfonamide,

X represents a direct single bond or alkylene with up to 6 carbon atoms,

M stands for a mono-, di-, tri- or tetrapeptide which has the same or different groups via the α-amino group and / or via amino and / or hydroxy groups of the side chains S

II

Ar-X-NH-C is linked, with further functional groups of

Peptide can optionally carry protective groups,

C represents a residue of a cytostatic or a cytostatic derivative which is linked to M via an amino function or via an oxygen atom,

and their stereoisomers, stereoisomer mixtures and salts.

C can be an intercalating substance, a topoisomerase inhibitor, an anti-metabolite, an alkylant, a tubulin inhibitor, a tyrosine phosphokinase inhibitor, a protein kinase C inhibitor or an active substance with a different or unknown cytostati or see cytotoxic mechanism of action. C can be, for example, a nucleoside, an endiin antibiotic, a quinolone or naphthyridone carbon acid or a cytotoxic peptide antibiotic e.g. be from the class of Dolastatine. C can batracylin, quinolone-a, 5-fluorouracil, cytosine arabinoside, methotrexate, etoposide, camptothecin, a camptothecin derivative, daunomycin, doxorubicin,

Taxol, vinblastine, vincristine, dynemicin, calicheamycin, esperamycin, quercetin, suramin, erbstatin, cyclophosphamide, mitomycin C, melphalan, cisplatin, bleomycin, staurosporine or another anti neoplastic active ingredient.

Unless otherwise stated, the term “alkyl groups” is intended to include straight-chain, branched, cyclic and cycloalkyl radicals containing alkyl radicals. This definition should also apply analogously to all other radicals containing alkyl groups, such as alkoxy etc.

Preferred compounds of the formula (I) are those in which

Ar stands for a phenyl radical which is para to X, also hydroxyl, carboxy,

Can carry isothiocyanato or halogen.

Also preferred are compounds of formula (I), wherein X represents a single bond or methylene

Also preferred are compounds of formula (I)

wherein

M stands for a mono-, di- or Tπpeptide which has the same or different groups S via the α-amino group and / or via amino and / or hydroxy groups of the side chains

II

Ar-X-Nπ-C is linked, with further functional groups of

Peptide can optionally carry protective groups

The peptides M preferably consist of amino acid residues which are derived from alanine, aspartic acid, glutamic acid, glycine, leucine, histidine, lysine, arginine,

Ornithine, senn, tyrosine, valine, diaminopropionic acid, γ-diaminobutyric acid or phenylalamine, where several amino acid residues can be linked peptically both via the α-amino group and optionally via the side chain amino functions or via both functions

If M carries further functional groups, these are preferably unblocked

Also preferred are compounds of the formula (I) in which C for a batracyhn, methotrexate, quinolone a, etoposide, melphalan, taxol, camptothecin radical, a camptothecin modified in the A ring or B ring Deπvat, a Dauomycin- or Doxorubicin residue stands, whereby C is linked to M via an amino or hydroxy function. Very particularly preferred examples of C are

Residues of Batracyhn, Chinolon-a and Doxorubicin, Camptothecin, 7-Ethylcampto-thecin, 10, l l- (Methylenedιoxy) camptothecιn, 7-Hydroxymethylcamptothecιn and 7-Ethyl-10-hydroxy camptothecin

The compounds according to the invention can exist in stereoisomeric forms, for example as enantiomers or diastereomers, or as mixtures thereof, for example as a racemate. The invention relates both to the pure stereoisomers and to their mixtures If necessary, the stereoisomer mixtures can be separated into the stereoisomerically uniform constituents in a known manner, for example by chromatography or by crystallization processes

The amino acid residues can each be in the D-form or in the L-form

The amino acid nomenclature follows the rules established by IUPAC

If the stereochemistry was not specified, amino acids of the L form were used

The compounds according to the invention can occur in rotation isomeric forms or as mixtures thereof as a result of a rotation inhibition

If necessary, rotational isomer mixtures can be separated into the uniform constituents by known methods, for example by chromatography (for example HPLC) or by crystallization processes. This is possible not only at the final stage of the conjugates but also, if appropriate, at intermediate stages. If appropriate, the rotamer-pure end stages can be prepared from the rotamer-pure intermediates by suitable synthesis

The compounds according to the invention can also be present in the form of their salts. In general, salts with organic or inorganic bases or acids and internal salts may be mentioned here.

The acids which can be added preferably include hydrohalic acids, such as, for example, hydrochloric acid and hydrobromic acid, in particular hydrochloric acid, furthermore phosphoric acid, nitric acid, sulfuric acid, mono- and bifunctional carboxylic acids and hydroxycarboxylic acids, such as, for example, acetic acid, trifluoroacetic acid, Maleic acid, malonic acid, oxalic acid,

Gluconic acid, succinic acid, fumaric acid, tartaric acid, citric acid, salicylic acid, sorbic acid and lactic acid as well as sulfonic acids, such as p-toluenesulfonic acid, 1,5-naphthane disulfonic acid or camphorsulfonic acid

Physiologically acceptable salts can also be metal or ammonium salts of such compounds according to the invention which have a free carboxyl group Particularly preferred are, for example, sodium, potassium, magnesium or calcium salts, and also ammonium salts which are derived from ammonia or organic amines such as, for example, ethylamine, di- or tπethylamine, di- or tπethanolamine, dicyclohexylamine, dimethylaminoethanol, arginine, lysine, Ethylenediamine or phenethylamine

The invention further relates to a process for the preparation of compounds of the general formula (I), characterized in that compounds of the general formula (II)

M'-C (II),

in which C has the meaning given above and M 'represents a radical M which carries hydrogen atoms at the desired linking sites and whose remaining potential linking sites are blocked by protective groups,

with compounds of the general formula (III)

Ar - X- N = C - S (III)

in suitable solvents in the presence of a base to give compounds of the general formula (Ia)

S ιι (Ia)

Ar-X-NH-C-M "-C

implements

in which Ar, X and C have the meanings given above and M "stands for a radical M whose further potential linking sites are blocked by protective groups,

S

II and in the case of the introduction of further groups Ar-X-NH-C _^

which differ from the one (s) initially introduced, the corresponding protective groups, if appropriate, selectively from the compounds of the formula (Ia) splits off, in the manner indicated above, with further compounds of the general formula (III) which differ from those initially introduced, and, if appropriate, this reaction sequence for the introduction of further

S

II repeats the introduced residues of various residues Ar-X-NH-C,

and that any remaining protecting groups may be split off.

The conjugates according to the invention can be prepared, for example, by linking cytostatics derivatives carrying hydroxyl or amino groups (e.g. batracylin, quinolones or camptothecins) to activated carboxyl components, which in turn can be parts of protected amino acids, peptides or N-thiocarbonyl-modified peptides.

The compounds of the general formula (II) can be obtained by linking protected amino acid building blocks to amino or hydroxy functions of C using customary methods of peptide chemistry and, if appropriate, building up a peptide chain by gradually introducing further amino acid building blocks. Alternatively, peptide building blocks carrying protective groups can also be linked to C using customary methods.

The reactions can be carried out at various pressure and temperature ratios, for example from 0.5 to 2 bar or from -30 to + 100 ° C., in suitable solvents such as dimethylformamide (DMF), tetrahydrofuran (THF), dichloromethane, chloroform, and lower alcohols , Acetonitrile, dioxane, water or in mixtures of the solvents mentioned. As a rule, reactions in DMF or THF / dichloromethane at normal pressure and at a temperature of 0 to 60 ° C., in particular at about room temperature, are preferred.

For the activation of the carboxyl groups come the coupling reagents known in peptide chemistry, such as e.g. in Jakubke / Jeschkeit: amino acids,

Peptides, proteins; Verlag Chemie 1982 or Tetrahedr. Lett. 34, 6705 (1993). For example, acid chlorides, N-carboxylic acid anhydrides or mixed anhydrides are preferred.

Also preferred for activating the carboxyl groups is the formation of adducts with carbodiimides, for example N, N'-diethyl, N, N'-diisopropyl, N, N'-dicyclo- hexylcarbodiimide, N- (3-dimethylaminopropyl) -N'-ethyl-carbodiimide hydrochloride, N-cyclohexyl-N '- (2-morpholinoethyl) -carbodiimide-metho-p-toluenesulfonate, or carbonyl compounds such as carbonyldiimidazole, or 1, 2- Oxazolium compounds such as 2-ethyl-5-phenyl-l, 2-oxazolium-3-sulfate or 2-tert-butyl-5-methyl-isoxazolium perchlorate, or acylamino compounds such as 2-ethoxy-l-ethoxycarbonyl-

1,2-dihydroquinoline, or propanephosphonic anhydride, or isobutylchloroform, or benzotriazoilyloxy-tris (dimethylamino) phosphonium hexafluorophosphate, 1-hydroxybenzotriazole or hydroxysuccinimide ester.

Triethylamine, Hünig base, ethyldiisopropylamine, pyridine, N, N-dimethylaminopyridine or others can be used as bases, for example.

The protective groups known in peptide chemistry, for example of the urethane, alkyl, acyl, ester or amide type, can be used as protective groups for any further reactive functions in the cytostatic part or for third functions of the amino acids.

Amino protecting groups in the context of the invention are the usual amino protecting groups used in peptide chemistry.

These preferably include: benzyloxycarbonyl, (Cbz) 3,4-dimethoxybenzyloxycarbonyl, 3,5-dimethoxybenzyloxycarbonyl, 2,4-dimethoxybenzyloxycarbonyl, 4-methoxybenzyloxycarbonyl, 4-nitrobenzyloxycarbonyl, 2-nitrobenzyloxycarbonyl, 2-nitro-4,5 -dimethoxybenzyloxycarbonyl, methoxycarbonyl, ethoxycarbonyl, tert-butoxycarbonyl, (Boc) allyloxycarbonyl, vinyloxycarbonyl, 3,4,5-trimethoxybenzyloxycarbonyl, phthaloyl, 2,2,2-trichloroethoxycarbonyl, 2,2,2-trichloro-tert -butoxycarbonyl, menthyloxycarbonyl, 4-nitrophenoxycarbonyl, fluoro-enyl-9-methoxycarbonyl (Fmoc), formyl, acetyl, propionyl, pivaloyl, 2-chloroacetyl, 2-bromoacetyl, 2,2,2-trifluoroacetyl, 2,2, 2-trichloroacetyl, benzoyl, benzyl,

4-chlorobenzoyl, 4-bromobenzoyl, 4-nitrobenzoyl, phthalimido, isovaleroyl or benzyloxymethylene, 4-nitrobenzyl, 2,4-dinitrobenzyl, 4-nitrophenyl or 2-nitrophenylsulfenyl. Particularly preferred protecting groups are Fmoc, Boc and Cbz.

Protective groups can be split off in the corresponding reaction steps, for example by exposure to acid or base, hydrogenolytically or in some other way reductively. Biological testing

1. Growth inhibition test to determine the cytotoxic properties

The human colon tumor cell lines SW 480 and HT 29 (ATCC No. CCL 228 and HBT-38) and the mouse melanoma cell line B16F10 were grown in Roux dishes in RPMI 1640 medium with the addition of 10% FCS. It was then trypsinized and taken up in RPMI plus 10% FCS to a cell number of 50,000 cells / ml. 100 μl of cell suspension / well were placed in a 96 microwell plate and incubated for 1 day at 37 ° C. in a C0 ₉ incubator. A further 100 μl of RPMI medium and 1 μl of DMSO with the test substances were then added.

The growth was checked after day 3 and day 6. This became everyone

Microwell 40 μl MTT solution (3- (4,5-dimethyIthiazol-2-yl) -2,5-diphenyltetrazoline bromide) with an initial concentration of 5 mg / ml H., 0 was added. It was

Incubated for 5 hours in the C0 ₂ incubator at 37 ° C. The medium was then suctioned off and 100 μl of i-propanol / well were added. After 30 min. The absorbance at 540 nm was shaken with 100 μl H ₂ 0 using a Titertek Multiskan

MCC / 340 (flow) measured.

The cytotoxic effect is given in Table 1 as an IC ₅₀ value for the SW 480 and HT 29 and B16F10 cell lines:

Table 1:

IC ₅₀ / μM IC ₅₀ / μM IC ₅₀ / μM

Example SW 480 HT 29 B16F10

1.2) 25 40 -

1.3) 45 70 -

1.4) 40 40 30

1.5) 250 400 -

1.6) 550 800

^" eispi ιc ₅ o μM IC ₅₀ / μM

B el ιc ₅₀ / μM

SW 480 HT 29 B 16F10

2.1) 20 9 9

2.2) 15 6 4

2.3) 0.9 0.7 0.2

2.4) 0.8 0.9 1

2.5) 200> 200 200

2.6) 0.2 0.3 0.06

2.8) 0.2 o, ι 0.1

2.9) 2 2 1

2.10) 2 2 0.4

2.11) 60 150 30

3) 3 2 1

4.1) 0.01 0.02 o, ι

4.2) 0.07 0.06 0.3

4.3) 0.02 0.02 o, ι

4.4) 0.3 0.2 0.6

4.5) 0.3 0.2 0.8

4.6) 0.2 0.15 0.5

4.7) 0.1 0.06 0.3

4.8) 0.3 0.15 0.8

4.9) 0.02 0.015 0.2

4.10) 0.02 0.01 0.2

4.1 1) 0.06 0.03 0.2

4.12) 0.04 0.03 0.2

4.13) 0.06 0.04 0.2

4.14) 0.16 0.075 0.75 1

13 -

Example ιc ₅₀ / μM ιc ₅₀ / μ ιc ₅₀ / μ

SW 480 HT 29 B16F10

4.15) 0.09 0.06 0.2

4.16) 0.15 0.12 0.6

4.17) 0.3 0.17 0.8

4.18) 0.3 0.12 0.4

4.19) 0.08 0.04 0.4

4.20) 0.07 0.06 0.3

4.21) 0.7 0.3 3

4.22) 0.04 0.04 0.1

4.23) 0.08 0.07 0.15

5.1) 0.025 0.02 0.05

5.2) 0.5 0.3 0.9

6) 0.005 0.003 0.015

7) 0.06 0.08 1

8) 0.15 0.2 3.0

2. Hematopoietic activity of conjugates compared to the underlying drug

Material and methods:

Bone marrow cells were rinsed from the mouse femur. 10 ⁵ cells are in McCoy 5A medium (0.3% agar) together with recombinant murine GM

CSF (Genzyme; stem cell colony formation) and the substances (10 ^"4 to 100 μg / ml) were incubated at 37 ° C. and 7% CO _2. 7 days later the colonies (<50 cells) and clusters (17-50 cells ) counted.

Results: As shown in Table 2, the investigated conjugates show a drastically reduced inhibition of bone marrow stem cell proliferation compared to the underlying active ingredient.

Table 2: Inhibition of CSF-induced proliferation of mouse bone marrow stem cells

Example IC ₅₀ [ng / ml]

Quinolone-a 0.2

2.4) 60.0

Camptothecin 0.4

4.4) 10

4.9) 22

In vivo inhibition of tumor growth in the nude mouse model

Material:

Athymic nude mice (NMRI nu / nu strain) were used for all in vivo experiments to investigate the inhibition of tumor growth. The selected large cell lung carcinoma LXFL 529 was by serial passage in Nude mice developed. The human origin of the tumor was proven by iso-enzymatic and immunohistochemical methods.

Experimental setup:

The tumor was implanted subcutaneously in both flanks of 6 to 8 week old nu / nu nude mice. Depending on the doubling time, the treatment was started as soon as the tumors had reached a diameter of 5-7 mm. The mice were randomized to the treatment group and the control group (5 mice per group with 8-10 evaluable tumors). The individual tumors in the control group all grew progressively.

The size of the tumors was measured in two dimensions using a slide gauge. The tumor volume, which correlates well with the cell number, was then used for all evaluations. The volume was calculated according to the formula "length x width x width / 2" ([axb ² ] / 2, a and b stand for two diameters arranged at right angles).

Relative tumor volume (RTV) values were calculated for each tumor by dividing the tumor size on day X by the tumor size on day 0 (at the time of randomization). The mean values of the RTV were then used for the further evaluation.

The final measured value was the inhibition of the increase in tumor volume (tumor volume of the test group / control group, T / C, in percent).

Treatment:

The compounds were applied intraperitoneally (i.p.) on days 1, 2 and 3 after randomization.

Results: The compound from Example 4.4) shows the therapeutic activity of the conjugates according to the invention against the large-cell human lung tumor xenograph LXFL 529. Therapy leads to tumor remission at the maximum tolerable dose (MTD) and half of the MTD. Table 3:

Dose survival number of the relative relative

Therapy ht [mg / kg / day] time tumors tumor volume Köipergewic [days] [day 21] on day 21 on day 21 [% of day 0] [% of day 0]

> 39 35

Control group - 35> 18 16 1137 1 11.6> 35

7> 43

Example 6.25 4.4) (MTD)> 43> 43 9 0.2 1 13.0> 43

Example> 43> 43 4.4) 3.125 7 69.5 105.8> 43> 43

The compounds according to the invention have a surprisingly strong cytotoxic activity against various, both in vitro and in vivo

Tumors, especially those of the lungs and colon, are associated with a high selectivity towards non-malignant cells.

They are therefore suitable for the treatment of cancer, particularly those of the lungs and colon.

The present invention includes pharmaceutical preparations which, in addition to non-toxic, inert pharmaceutically suitable excipients, contain one or more compounds according to the invention or which consist of one or more active compounds according to the invention, and processes for the preparation of these preparations.

The active ingredient (s) can optionally also be in microencapsulated form in one or more of the above-mentioned carriers.

The therapeutically active compounds should be present in the pharmaceutical preparations listed above preferably in a concentration of about 0.1 to 99.5, preferably of about 0.5 to 95% by weight of the total mixture. In addition to the compounds according to the invention, the pharmaceutical preparations listed above can also contain further active pharmaceutical ingredients.

In general, it has proven to be advantageous in both human and veterinary medicine to use the active ingredient (s) according to the invention in total amounts of about 0.5 to about 500, preferably 5 to 100 mg / kg of body weight per 24 hours, optionally in the form multiple doses to achieve the desired results. A single dose contains the active ingredient (s) according to the invention preferably in amounts of about 1 to about 80, in particular 3 to 30 mg / kg of body weight.

Synthesis examples

All thiocarbonyl-amino acid or thiocarbonyl peptide conjugates which are the subject of this invention are synthesized according to the following general procedure:

A solution of 1 mmol of the underlying amino acid or peptide conjugate in 50 ml of absolute dimethylformamide is mixed with 1.1 mmol of the corresponding isothiocyanate per free amino group. After adding 1.74 ml (10 mmol) of ethyldiisopropylamine, the mixture is stirred at room temperature until no more amino acid or peptide conjugate can be detected in the thin-layer chromatogram, but for a maximum of 16 h. One constricts i. Vak. and cleans the residue after drying i. High vacuum by flash chromatography on silica gel e.g. with an ethyl acetate / petroleum ether or a dichloromethane / methanol system. Repeated overturning from dichloromethane / methanol 1: 1 (v / v) with diethyl ether often results in pure products.

Remaining protective groups are then removed in a second step by methods known from the literature (a fluorenyl-9-methoxycarbonyl group e.g. with piperidine in absolute dimethylformamide at room temperature; a tert-butoxycarbonyl group e.g. with trifluoroacetic acid in absolute dichloromethane at room temperature).

The corresponding isothiocyanates can be purchased from chemical retailers or are synthesized using methods known from the literature. Precursor: Amino acid or peptide conjugates

Example 1.1

N- [N ^R - (Fuorenyl-9-methoxycarbonyl) -Iysyl] batracylin, trifluoroacetate

1.1. a) N- [N ^α - (tert-butoxycarbonyl) -N ^ε - (fluorenyI-9-methoxycarbonyl) - lysyl] - batracylin:

N ^α - (tert-butoxycarbonyl) -N ^ε - (fluorenyl-9-methoxycarbonyl) -lysine (5.3 g,

1 1.3 mmol) and 2-isobutoxy-l-isobutoxycarbonyl-l, 2-dihydroquinoline (4 ml,

14 mmol) are dissolved in 40 ml dichloromethane. After 20 min stirring at room temperature, a solution of batracylin (2.5 g, 10 mmol) in dimethylformamide (80 ml) is added and the mixture is stirred for a further 24 h at room temperature is concentrated in vacuo until crystallization begins. The suspension obtained is mixed with ethanol (500 ml) and boiled under reflux for 1 h. After cooling to room temperature, the product is filtered off and washed with acetone and then with diethyl ether. Yellow crystals are obtained

(5.9 g, 84%) [TLC (ethyl acetate) R _{ = 0.57, mp = 158 ° C (dec))

1.1) N- [N ^ε - (Fiuorenyl-9-methoxycarbonyl) -lysyl] -batracyIin, trifluoroacetate:

An anhydrous trifluoroacetic acid (25 ml) is added to a suspension of the above compound (5.6 g, 8 mmol) in dichloromethane (75 ml) and the resulting Solution stirred for 90 min at room temperature.After concentration in vacuo, the residue is crystallized by adding diethyl ether (200 ml) .The precipitate is filtered off and washed intensively with diethyl ether.After several drops of dichloromethane / methanol 1.1 with diethyl ether, yellow-orange crystals are obtained (5 , 13 g, 90%) [TLC (ethyl acetate) R _f = 0.05; Mp = 162 ° C (dec.)].

Example 1.2

N- [Seryl-D-alanyI] batracylin, trifluoroacetate

I.2.a) N- [N-benzyloxycarbonyl-D-alanyl] batracylin:

N-Benzyloxycarbonyl-D-alanine (3.9 g, 17.5 mmol) is reacted with batracylin (4.1 g, 16.4 mmol) analogously to Example I 1 a. After concentration in vacuo to 50 ml, the mixture is treated with ethyl acetate made up to 300 ml and immediately heated to boiling for 10 min. The mixture is then allowed to cool to room temperature, filtered and the filter material is boiled again with ethyl acetate (200 ml) by cooling to 0 ° C. with stirring and filtration gives yellow crystals. The crystals (6, 4 g, 80%) are separated off by filtration and the combined filtrates, after concentration in vacuo, are purified by flash chromatography [petroleum ether / ethyl acetate 3 2-1 1]. A further 1.35 g (17%) [TLC (ethyl acetate) - Rf = 0.45, mp = 256 ° C, [α] ²⁰ = + 75.1 ° (c = 1.0 / CH ₂ C1 ₂ + 0.5% CH ₃ OH)]

I.2.b) N- [D-AlanyI] -batracylin:

Compound 1.2 a (1 1.4 g, 25 mmol) is dissolved in a 33% solution of hydrogen bromide in glacial acetic acid (100 ml). After 30 min at room temperature, the mixture is concentrated in vacuo to 30 ml and then with vigorous stirring in saturated sodium bicarbonate solution (1000 ml) poured 8/15571 20 -

stir for 10 min. continue, filter off and wash with water, a little isopropanol and diethyl ether. The product is obtained in yellow crystals (7.87 g, 98%) [TLC (ethyl acetate): R _f = 0.06; Mp = 267 ° C (dec.)].

I.2.c) N- [N- (tert-butoxycarbonyl) -seryl-D-alanyl] -batracylin:

Representation in analogy to Example I.l.a from N- (tert-butoxycarbonyl) -serine and N-

[D-alanyl] batracylin (Example I.2.b); Education: 77%.

1.2) N- [Seryl-D-alanyl] batracylin, trifluoroacetate:

Representation in analogy to Example 1.1 from compound I.2.c; Education: 98%.

Example 1.3

N- [N ^ε - (Fuorenyl-9-methoxycarbonyl) -lysyI-D-aIanyl] -quinolone-a,

Trifluoroacetate

1.3.a) N- [N- (tert-butoxycarbonyl) -D-alanyl] -quinolone-a:

N- (tert-Butoxycarbonyl) -D-alanine (3.6 g, 19.2 mmol) and 2-isobutoxy-1-isobutoxycarbonyl-1, 2-dihydroquinoline (5.8 g, 19.2 mmol ) are dissolved in 200 ml of dimethylformamide. After stirring at room temperature for 8 h, quinolone-a (4 g, 9.6 mmol) and ethyldiisopropylamine (3.3 ml) are added and the mixture is treated with ultrasound for 10 h. The mixture is concentrated, the residue is taken up in dichloromethane and precipitated with ether. After filtration, washing with ether and drying in a high vacuum, 4.58 g (81%) of the target product are obtained, which is reacted without further purification.

I.3.b) N- [D-alanyl] -quinone-a, trifluoroacetate:

4.56 g (7.75 mmol) of the compound from the above example are dissolved in a mixture of dichloromethane (50 ml) and anhydrous trifluoroacetic acid (50 ml) at 0 ° C. and stirred at this temperature for 1 h. The mixture is concentrated, the mixture is distilled with dichloromethane and the residue is reprecipitated from methanol with diethyl ether. 4.07 g (87%) of the crystalline target product are obtained [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2): R _f = 0.34].

I.3.c) N- [N ^α - (tert-butoxycarbonyl) -N ^ε - (fluorenyl-9-methoxycarbonyl) - lysyl-D-alanyl] -quinolone-a:

N ^α - (tert-Butoxycarbonyl) -N ^ε - (fluorenyl-9-methoxycarbonyl) -lysine (1.57 g, 3.36 mmol) is dissolved in dimethylformamide (25 ml) and at 0 ° C with N-

Hydroxysuccinimide (600 mg, 5.04 mmol) and N, N'-dicyclohexylcarbodiimide (820 mg, 4.03 mmol) were added. After 3 h, the urea formed is filtered off, 1.5 g (2.86 mmol) of the compound from Example 1.3.b) are added to the filtrate and the mixture is stirred at room temperature for 16 h. Residual urea is filtered off and the filtrate is purified by flash chromatography [dichloromethane / methanol 97.5: 2.5 → 90: 10]. Then one falls over from dichloromethane / methanol 1: 1 with diethyl ether. Yield: 1.5 g (56%) [TLC (dichloromethane / methanol 9: 1): R _f = 0.47]. 1.3) N- [N ^ε - (FluorenyI-9-methoxycarbonyl) -lysyI-D-aIanyl] - quinolone-a, trifluoroacetate

Cleavage of the tert-butoxycarbonyl group from compound I.3.c in analogy to Example II and reprecipitation of the crude product from methanol with diethyl ether gives yellow crystals. Yield: 80% [TLC (dichloromethane / methanol / ammonia (17%) 15: 4: 0.5): R _f = 0.36].

Example 1.4

20-O- [N ^ε - (Fluorenyl-9-methoxycarbonyl) -lysyl-alanyl] -camptothecin, trifluoroacetate

CF ₃ COOH

I.4.a) 20-O- (alanyl) camptothecin, trifluoroacetate:

Camptothecin (500 mg, 1.44 mmol) are dissolved in absolute dimethylformamide (20 ml) and then with 4-dimethylaminopyridine (50 mg) and N-tert-butoxycarbonylalanine-N-carboxy-anhydride (775 mg, 3, 6 mmol) was added. After 3 h, a further 775 mg (3.6 mmol) of N-tert-butoxycarbonyl-alanine-N-carboxy-anhydride are added and the suspension is treated with ultrasound for 16 h. The mixture is concentrated, the raw material is taken up in dichloromethane (50 ml) and 5 ml of trifluoroacetic acid are added at 0 ° C. After 30 min. Stirring is concentrated again and the product is purified by flash chromatography (acetonitrile / water 20: 1). The corresponding fractions are collected, concentrated and after dissolving in dioxane / water lyophilized. 712 mg (93%) of the target compound are obtained [FAB-MS: m / z = 420 (M + H) ⁺ ],

1.4) 20-O- [N ^ε - (Fluorenyl-9-methoxycarbonyl) -lysyl-alanyl] - camptothecin, trifluoroacetate:

The conjugate from Example I.4.a is linked according to standard instructions (see Example I. la) with N ^α - (tert-butoxycarbonyl) -N ^ε - (fluorenyl-9-methoxycarbonyl) lysine and then in analogy to Example Il unblocked at the α-amino function. The target compound is obtained in a yield of 24% [TLC (acetonitrile / water 20: 1): R _f - 0.15].

Example 1.5

7-ethyl-20-O- (lysyl-alanyl) camptothecin, di-trifluoroacetate

I.5.a) 7-ethyl-20-O- [N- (tert-butoxycarbonyl) alanyl] camptothecin:

A solution of 1.88 g (5.0 mmol) of 20 (S) -7-ethyl-camptothecin (S. Sawada et al., Chem. Pharm.Bull. 39 (1991) 1446-1454) in 100 ml of absolute dimethylformamide 2.15 g (10.0 mmol) of N- (tert-butoxycarbonyl) alanine-N-carboxylic acid anhydride and 150 mg (1.2 mmol) of 4- (N, N-dimethylamino) pyridine are added with stirring. After 3 h at room temperature, a further 2.15 g (10.0 mmol) of N- (tert-butoxycarbonyl) alanine-N-carboxylic acid anhydride and 150 mg (1.2 mmol) of 4- (N, N-dimethylamino) - pyridine and stir overnight at room temperature.

The mixture is then concentrated in vacuo and the residue is purified by flash chromatography [petroleum ether / ethyl acetate 2: 1 → 1: 1 → ethyl acetate]. you receives 2.02 g (73.8%) of colorless crystals [TLC (ethyl acetate) R ₍ - 0.56, mp = 206-212 ° C, FAB-MS m / z = 548 (M + rf)]

1.5. b) 20-O-alanyl-7-ethyl-camptothecin, trifluoroacetate:

A solution of compound 15 a (1.81 g, 3.3 mmol) in a mixture of 70 ml dichloromethane and 7 ml anhydrous trifluoroacetic acid is stirred for 90 min at room temperature. After concentration in vacuo to a small volume, the product is precipitated with diethyl ether and washed thoroughly with diethyl ether. 1.34 g (72.3%) of light yellow crystals are obtained [TLC (ethyl acetate) R _t = 0.05, mp = 242 ° C (Zers)]

I.5.c) 7-ethyl-20-O- [N ^α , N ^ε -di- (tert-butoxycarbonyl) -IysyI-aIanyl] - camptothecin:

1.57 g (4.55 mmol) of N, N-di- (tert-butoxycarbonyl) lysine and 923 mg (6.83 mmol) of 1-hydroxy-1H-benzotriazole hydrate are dissolved in 35 ml of dimethylformamide. After addition of 1 , 09 g (5.7 mmol) of N-ethyl-N '- (dimethylaminopropyl) carbodimide hydrochloride and 990 μl (5.7 mmol) of ethyl-diisopropylamine are stirred for

30 min at room temperature Then a solution of compound 15 b (1.3 g, 2.32 mmol) in 35 ml of dimethylformamide and 408 μl (2.32 mmol) of ethyldiisopropylamine is added and the mixture is stirred for a further 16 h at room temperature After concentration in vacuo and purification by flash chromatography [petroleum ether / ethyl acetate 2 1 -> 1 1 - ethyl acetate], light yellow is obtained

Crystals yield 1.38 g (75.3%) [TLC (ethyl acetate). R _f = 0.53, mp - 125 ° C (decomp)]

1.5) 7-EthyI-20-O- (lysylalanyι) camptothecin, di-trifluoroacetate:

An anhydrous trifluoroacetic acid (5 ml) is added to a suspension of the above compound (1.18 g, 1.5 mmol) in dichloromethane (50 ml) and the resulting solution is stirred for 1 h at room temperature. After concentration to a small volume in vacuo the product precipitated by adding diethyl ether. The precipitate is filtered off and recrystallized from ethyl acetate. 862 mg (71.5%) of yellow crystals are obtained [TLC (ethyl acetate) R _f = 0.05, mp = 137 ° C. (dec.)]. Example 1.6:

7-IN ^e - [Fluorenyl-9-methoxycarbonyl] -L-lysyl-L-valyloxymethyl) camptothecin, trifluoroacetate

I.6.a) 7-hydroxymethyl camptothecin:

This compound is used according to the instructions of Miyasaka et al (Chem Pharm

Bull 39 (1991) 2574)

I.6.b) 7-L-ValyIoxymethyl) -camptothecin, trifluoroacetate:

1 g (2.64 mmol) of 7-hydroxymethyl-camptothecin is dissolved in 100 ml of DMF and then mixed with 100 mg of 4-N, N-dimethylaminopyridine and one equivalent of N-tert-butoxycarbonyl-L-valine-N-carboxy-anhydride and the suspension at 16 h

Room temperature stirred, the mixture is concentrated and the residue is purified by flash chromatography on ethyl acetate / petroleum ether 1 l and later 1.5 l. The purified material is taken up in 30 ml dichloromethane and mixed with 5 ml trifluoroacetic acid at 0 ° C. After 30 min stirring, the mixture is concentrated and the amino-blocked product from dichloromethane / ether is precipitated. The target compound is obtained in a

Overall yield of 55% [TLC (acetonitrile / water / glacial acetic acid 5 1 0.2) R _f = 0.37]

1.6 7-IN ^ε - [fluorenyl-9-methoxycarbonyl] -L-lysyl-L-valyloxymethylJ-camptothecin, trifluoroacetate

560 mg of the conjugate from Example I 6 b become a solution of 560 mg (1.5 Aq) N ^α , N ^e -bis- (tert-butoxycarbonyl) -L-lysine, 239 mg N-hydroxybenzotriazole and 271 mg N-ethyl-N '- (3-dimethylaminopropyl) carbodiimide hydrochloride is added to 50 ml of dimethylformamide and stirred for two hours at room temperature. The mixture is concentrated, taken up in dichloromethane and extracted three times with water. After drying the organic phase, the mixture is concentrated and by flash chromatography ( Petroleum ether / ethyl acetate 1 1> ethyl acetate)

The product obtained is then taken up in 20 ml of dichloromethane, 3 ml of trifluoroacetic acid are added at 0 ° C. and the mixture is stirred for one hour at room temperature. After concentration and precipitation from dichloromethane / ether, the product is obtained Target compound in 62% yield [DC acetonitrile / water / glacial acetic acid 5 1 0.2 R, = 0.62]

1.7 10, l l-methylenedioxy-20-O- {N ^ε - [FluorenyI-9-methoxycarbonyl] -syl-leucylj-camptothecin, trifluoroacetate

I.7.a) 10,11-methylenedioxy-camptothecin:

This camptothecin derivative is produced according to Wall et al (J Med Chem 29 (1986), 2358) from the S-configured, enantiomerically pure tricyclic compound, which can be obtained, for example, by racemate resolution

I.7.b) 10.1 l- (methylenedioxy) -20-O-leucyl-camptothecin, trifluoroacetate

150 mg (0.382 mmol) of 10,11-methylenedioxy-camptothecin are dissolved in 20 ml of DMF and then mixed with 20 mg of 4-N, N-dimethylaminopyridine and 10 equivalents of N-tert-butoxycarbonyl-L-leucine-N-carboxy-anhydride and the suspension was stirred at 40 ° C. for 16 h. The mixture is concentrated and purified by flash chromatography on ethyl acetate / petroleum ether 2 1. The purified material is taken up in 15 ml of dichloromethane and 2 ml of trifluoroacetic acid are added at 0 ° C. After 30 min

Stirring is concentrated and the amino-blocked product is precipitated from dichloromethane / methanol with ether. The target compound is obtained in a total yield of 35%.

I.7.c) 10, U-methylenedioxy-20-O- {N ^e - [fluorenyl-9-methoxycarbonyl] -IysyI- leucylj-camptothecin, trifluoroacetate

The conjugate from Example I 7 b is linked according to standard instructions (see Example I 1 a with N ^α - (tert-butoxycarbonyl) -N ^E - (fluorenyl-9-methoxycarbonyl -) - lysιn and then to the α-amino function by the action of Trifluoroacetic acid deblocks 69% yield over 2 steps [DC Acetomtril / Water 10 1 R _f = 0.4] 8/15571 27

Example 1.8

20-O- (Lysyl-aspartyI) camptothecin, di-hydrobromide

I.8.a) 20-O- [N- (tert-Butoxycarbonyl) -aspartyl- (γ-benzyl ester)] - camptothecin:

A suspension of 5.23 g (15.0 mmol) of 20 (S) -camptothecin in 400 ml of absolute dimethylformamide is stirred with 10.45 g (30.0 mmol) of N- (tert-butoxycarbonyl) -aspartic acid- (γ-benzyl ester) -N-carboxylic acid anhydride and 367 mg (3.0 mmol) of 4- (N, N-dimethylamino) pyridine were added. After stirring for 8 h at 60 ° C., a further 5.23 g (15.0 mmol) of N- (tert-butoxycarbonyl) -aspartic acid (γ-benzyl ester) -N-carboxylic acid anhydride and 183.5 mg (1.5 mmol ) 4- (N, N-Dimethylamino) pyridine and stir for three days at room temperature. The mixture is then concentrated in vacuo and the residue is purified by flash chromatography [petroleum ether / ethyl acetate 1: 2]. 2.3 g (23.4%) of orange-yellow crystals are obtained [TLC (ethyl acetate): R _f - 0.59; Mp = 130 ° C (dec.)].

I.8.b) 20-O-aspartyl- (γ-benzyl ester) -camptothecin, trifluoroacetate:

A solution of compound I.8.a (2.22 g, 3.4 mmol) in a mixture of 70 ml dichloromethane and 7 ml anhydrous trifluoroacetic acid is for 90 min. stirred at room temperature. After concentration in vacuo to a small volume, the product is precipitated with diethyl ether and washed thoroughly with diethyl ether. 1.08 g (72.3%) of beige crystals are obtained [TLC (ethyl acetate): R _f = 0.14, mp = 216 ° C. (dec.)]. 1.8.c) 20-O- [N, N ^ε -di- (tert-butoxycarbonyl) -lysyl-aspartyl- (γ-benzyl ester)] - camptothecin:

433 mg (1.25 mmol) of N, N-di- (tert-butoxycarbonyl) lysine and 338 mg (2.50 mmol) of 1-hydroxy-1H-benzotriazole hydrate are dissolved in 15 ml of dimethylformamide. After adding 360 mg (1.88 mmol) of N-ethyl-N '- (dimethylaminopropyl) carbodimide hydrochloride and 500 μl (3.0 mmol) of ethyl-diisopropylamine, the mixture is stirred for 15 min. at room temperature. A solution of compound I.8.b (500.7 mg, 0.75 mmol) in 15 ml of dimethylformamide and 200 μl (1.13 mmol) of ethyl-diisopropylamine is then added and the mixture is stirred for a further 16 h in the room - temperature. After concentration in vacuo, the residue is taken up in dichloromethane and the solution is washed once with water. It is dried over MgSO4 and the residue which remains after concentration in vacuo is purified by flash chromatography [petroleum ether / ethyl acetate 1: 2] to give beige crystals. Yield: 473.8 mg (70.5%) [TLC (ethyl acetate): R _f = 0.42; Mp = 99 ° C (dec.)].

1.8) 20-O- (lysyl-aspartyl) camptothecin, di-hydrobromide:

A solution of the above compound (462 mg, 0.52 mmol) in dichloromethane (25 ml) is mixed with a 33% solution of hydrogen bromide in acetic acid (5 ml) and the suspension resulting after a few minutes is stirred for 1 h at room temperature . It is decanted from the precipitated product and the residue is washed thoroughly with diethyl ether. For cleaning is after

Solution in warm ethanol reprecipitated by adding diethyl ether. 391 mg (100%) of yellow crystals are obtained [TLC (acetonitrile / water 5: 1): R _f = 0.05; Mp = 225 ° C (dec.)].

Example 1.9

20-O- (lysylseryl) camptothecin, di-hydrobromide

I.9.a) 20-O- [O-BenzyI-N- (tert-butoxycarbonyl) -seryl] -camptothecin:

A suspension of 5.23 g (15.0 mmol) 20 (S) -camptothecin in 400 ml absolute

Dimethylformamide is stirred with 9.64 g (30.0 mmol) of O-benzyl-N- (tert-butoxycarbonyl) -serine-N-carboxylic acid anhydride and 367 mg (3.0 mmol) of 4- (N, N-dimethylamino) -pyridine added. After stirring for 8 h at 60 ° C., a further 4.82 g (15.0 mmol) of O-benzyl-N- (tert-butoxycarbonyl) -serine-N-carboxylic acid anhydride and 183.5 mg (1.5 mmol) 4 are added - (N, N-Dimethylamino) pyridine and stir for three

Days at room temperature. The mixture is then filtered, the filtrate is concentrated in vacuo and the residue is purified by flash chromatography [petroleum ether / ethyl acetate 2: 1 → 1: 1 → 1: 2]. 6.66 g (70.9%) of a yellow foam are obtained [TLC (acetonitrile ethyl acetate 1: 1): R = 0.66; FAB-MS: m / z = 626 (M + H ⁺ )].

I.9.b) 20-O- [O-benzylseryl] camptothecin, trifluoroacetate:

A solution of compound I.9.a (2.5 g, 4.0 mmol) in a mixture of 20 ml dichloromethane and 4 ml anhydrous trifluoroacetic acid is stirred for 1 h at room temperature. After concentration in vacuo to a small volume, the product is precipitated with diethyl ether and washed thoroughly with diethyl ether. 2.51 g (98.1%) of yellow crystals are obtained [TLC (acetonitrile / ethyl acetate 1: 1): R _f = 0.17; Mp = 198 ° C (dec.)]. 8/15571 _ ₃₀

1.9.c) 20-O- [N ^α , N ^F -di (tert-butoxycarbonyl) -Iysyl- (O-benzyl) -seryl] - camptothecin:

1.73 g (5.0 mmol) of N, N-di- (tert-butoxycarbonyl) lysine and 1.35 g (10 mmol) of 1-hydroxy-1H-benzotriazole hydrate are dissolved in 50 ml of dimethylformamide. After adding 1.44 g (7.5 mmol) of N-ethyl-N '- (dimethylaminopropyl) carbodiimide

Hydrochloride and 2.0 ml (12 mmol) of ethyl diisopropylamine are stirred for 15 min. at room temperature. A solution of compound I.9.b (1.92 g, 3.0 mmol) in 50 ml of dimethylformamide and 790 μl (4.5 mmol) of ethyldiisopropylamine is then added and the mixture is stirred for a further 16 h at room temperature. After concentration in vacuo, the residue is by flash chromatography

[Petroleum ether / ethyl acetate 3: 1 - »1: 1 -> 1: 3] to yellow crystals. Yield: 2.32 g (89.1%) [TLC (ethyl acetate): R _f = 0.45; Mp = 130 ° C (dec.)].

1.9) 20-O- (lysylseryl) camptothecin, di-hydrobromide:

A solution of the above compound (2.13 g, 2.46 mmol) in dichloromethane (120 ml) is washed with a 33% solution of hydrogen bromide in acetic acid

(25 ml) were added and the suspension resulting after a few minutes was stirred at room temperature for 1 h. It is decanted from the precipitated product and the residue is washed thoroughly with diethyl ether. After cleaning in dichloromethane / methanol, the mixture is reprecipitated 1: 1 by adding diethyl ether. 1.78 g (100%) of yellow crystals are obtained [TLC (acetonitrile / water 5: 1):

R _f - = 0.05].

Example 1.10

7-ethyl-20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) lysylvalyl] camptothecin, trifluoroacetate

1.10.a) 20-O- [N- (tert-Butoxycarbonyl) -valyl] -7-ethyl-camptothecin:

The compound is prepared from 1.88 g (5.0 mmol) of 20 (S) -7-ethyl-camptothecin (S. Sawada et al., Chem. Pharm. Bull. 39 (1991) 1446-1454) and 2.43 g (10.0 mmol) of N- (tert-butoxycarbonyl) -valine-N-carboxylic anhydride. 1.46 g (51%) of beige crystals are obtained [TLC (acetonitrile): R _f = 0.86; M.p. = 224-227 ° C (dec.); FAB-MS: m / z = 576 (M + FT)].

I.lO.b) 7-ethyl-20-O-valyl-camptothecin, trifluoroacetate:

From compound I.lO.a (1.44 g, 2.5 mmol) the N-

(tert-Butoxycarbonyl) group split off. 626 mg (43%) of yellow crystals are obtained [TLC (acetonitrile): R _f = 0.45; Mp = 160 ° C (dec.)].

I.lO.c) 20 -O - [N ^α - (tert-Bu to xy ca rb o nyI) - N ^ε - (fI uoren yl - 9 - methoxycarbonyl) -lysyl-valyI] -7-ethyl-camptothecin:

Analogous to 1.5. c 797 mg (1.7 mmol) N ^α - (tert-butoxycarbonyl) -N ^ε -

(Fluorenyl-9-methoxycarbonyl) lysine reacted with compound I.lO.b (590 mg, 1.0 mmol). After concentration in vacuo and purification by flash chromatography [petroleum ether / ethyl acetate 1: 2], beige crystals are obtained. Yield: 287 mg (31%) [TLC (ethyl acetate): R _f = 0.50; Mp = 172 ° C (dec.)]. 1.10) 7-ethyl-20-O- [N ^f - (fIuorenyl-9-methoxycarbonyl) -lysyl-vaIyl] - camptothecin, trifluoroacetate:

The above compound (277.8 mg, 0.3 mmol) is deprotected as described with trifluoroacetic acid in dichloromethane. 209 mg (74%) of yellow crystals are obtained [TLC (ethyl acetate). R _f = 0.06, mp = 199 ° C (dec.)]

Example 1.11

7-ethyl-20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) lysylvalyl] -10-hydroxycamptothecin, trifluoroacetate

I.ll.a) 20-O- [N- (tert-Butoxycarbonyl) -valyI] -7-ethyl-10-hydroxy-camptothecin:

The compound is prepared using the method described in I 5 a from 392.4 mg (1.0 mmol) of 20 (S) -7-ethyl-10-hydroxy-camptothecin (S Sawada et al, Chem Pharm Bull 39 (1991) 3183-3188) and a total of 2.43 g (10.0 mmol) of N- (tert-butoxycarbonyl) -valine-N-carboxylic acid anhydride within 6 days. Flash chromatography [petroleum ether / ethyl acetate 5 1 -> 2 1 -> 1 1] 353 mg (45%) of light yellow crystals [TLC (acetonitrile / ethyl acetate 1 1) R _f = 0.63, mp = 95-97 ° C]

1.1 l.b) 7-ethyl-10-hydroxy-20-O-valyl-camptothecin, trifluoroacetate:

The compound l i l a (340 mg, 0.43 mmol) is used as described under I 5 b

Cleaving the N- (tert-butoxycarbonyl) group 255 mg (98%) of yellow crystals are obtained [TLC (acetonitrile / ethyl acetate 1 l) R _t = 0.04, mp = 189 ° C. (dec)) I.ll.c) 20-O- [N ^α - (tert-butoxycarbonyl) -N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-valyI] -7-ethyl-10-hydroxy-camptothecin:

Analogously to I.5.C, 562.3 mg (1.2 mmol) of N - (tert-butoxycarbonyl) -N ^ε - (fluorenyl-9-methoxycarbonyl) -lysine with compound 1.11.b (242.2 mg, 0.4 mmol) implemented. After concentration in vacuo and purification by flash chromatography [petroleum ether / ethyl acetate 5: 1 → 3: 1 → 1: 1], yellow crystals are obtained. Yield: 251 mg (67%) [TLC (acetonitrile / ethyl acetate 1: 1): R _f = 0.68; Mp = 163 ° C (dec.)].

1.11) 7-Ethyl-20-O- [N ^ε - (fIuorenyI-9-methoxycarbonyl) -Iysyl-valyl] -10-hydroxy-camptothecin, trifluoroacetate:

The above compound (244.9 mg, 0.26 mmol) is deprotected as described with trifluoroacetic acid in dichloromethane. 115 mg (46%) of yellow crystals are obtained [TLC (acetonitrile / ethyl acetate 1: 1): R _f = 0.05; Mp = 196 ° C (dec.)].

Examples 1.1 - 1.3

Batracylin conjugates with an amino acid; general formula:

1.1) N- [N- (4-Hydroxyphenylamino-thiocarbonyl) -D-alanyl] batracylin

Educt: N- (D-alanyl) -batracylin yield: 76% [TLC (ethyl acetate / glacial acetic acid 100: 1): R _f = 0.53; Mp: 185 ° C]

1.2) N- [N ^α - (4-HydroxyphenyIaminino-thiocarbonyl) Iysyl] batracylin

Starting material: N- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl] -batracylin, trifluoroacetate

Yield: 68% over 2 steps [TLC (dichloromethane / methanol 5: 1): R _f = 0.31; Mp: 162 ° C (dec.)] 1.3) N- [N ^ε - (4-Hydroxy-phenylamino-thiocarbonyl) -lysyl] -batracyIin

Starting material: N- [N ^α - (tert-butoxycarbonyl) lysyl] -batracylin

Yield: 71% over 2 steps [TLC (dichloromethane / methanol 5: 1): R, = 0.30;

Mp: 162 ° C (dec.)]

Examples 1.4 - 1.8

Batracylin conjugates with two amino acids; general formula:

1.4) N- [N ^α - (4-hydroxy-phenylamino-thiocarbonyl) -lysyl-D-alanyl] - batracylin starting material: N- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-D-alanyl] -batracylin ,

Trifluoroacetate,

Yield: 70% over 2 steps [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2): R _f =

0.36]

1.5) N- [N- (4-HydroxyphenylIamino-thiocarbonyl) seryI-D-alanyl] batracylin

Educt: N- (seryl-D-alanyl) batracylin, trifluoroacetate

Yield: 45% [TLC (dichloromethane / methanol / ammonia 17% 15: 2: 0.2):

R _f - 0.32]

1.6) N- [N- (4-HydroxyphenylIamino-thiocarbonyI) glutamyl-D-alanyl] batracylin

Educt: N- (glutamyl-D-alanyl) -batracylin yield: 70% [TLC (dichloromethane / methanol / ammonia 17% 15: 8: 0.8):

R _f - 0.68] 98/15571 35

1.7) N- [N ^α , N ^ε -bis- (4-hydroxy-phenylamino-thiocarbonyl) -lysyl-seryl] -batracylin

Educt N- (lysyl-seryl) -batracyhn, di -trifluoroacetate yield 46% [TLC (dichloromethane / methanol / ammonia 17% 15 3 0.2)

R _f = 0.24, mp 155-157 ° C (dec))

1.8) N- {N ^α - [N ^α , N ^ε -bis- (4-hydroxy-phenylamino-thiocarbonyl) -lysyI] - α, ß-diaminopropionyl} -batracylin

Educt N- [N ^α -Lysyl-Nß- (fluorenyl-9-methoxycarbonyl) -α, ß-diamιno-propionyl] -batracylin, di-trifluoroacetate yield 39% over 2 steps [TLC (acetonitrile / water / glacial acetic acid 5 1 0 , 2) R _f =

0.54]

Examples 2.1 - 2.10

Quinolone-a conjugates with an amino acid; general formula:

2.1) N- [N- (4-hydroxyphenylamino-thiocarbonyι) alanyl] - quinolone-a

Educt N- (alanyl) quinolone-a, trifluoroacetate

Yield 48% [TLC (acetonitrile / water 10 1) R _f = 0.55]

2.2) N- [N- (4-Hydroxy-phenylamino-thiocarbonyl) -D-alanyl] quinolone-a

Educt N- (D-alanyl) -quinolone-a, trifluoroacetate yield 61% [TLC (dichloromethane / methanol / glacial acetic acid 90 10 1) R _f = 0.38] 2.3) N- [N ^α - (4-Hydroxy-phenylamino-thiocarbonyl) -α, γ-diamino-butyryl] -quinolone-a, hydrochloride

Educt: N- [N ^γ - (fluorenyl-9-methoxycarbonyl) -, γ-diaminobutyryl] - quinolone-a, trifluoroacetate salt-free precursor: 60% over 2 steps [DC

(Dichloromethane / methanol / ammonia 17% 10: 10: 3): R _f - 0.51; Mp: 221 ° C (dec.)]

Hydrochloride: The compound is suspended in water and the pH

Value adjusted to 2-3 with 0.1 N hydrochloric acid. After filtration, the filtrate is lyophilized.

2.4) N- [N ^α - (4-hydroxyphenylamino-thiocarbonyl) lysyl] quinolone-a, hydrochloride

Educt: N- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl] -quinolone-a,

Trifluoroacetate yield: 74% over 2 steps [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2): R _f =

0.33]

2.5) N- [N ^α , N ^ε -bis- (4-hydroxyphenylamino-thiocarbonyl) -D-lysyl] quinolone-a

Educt: N- (D-lysyl) -quinolone-a, di-trifluoroacetate yield: 59% [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2): R _f = 0.33; Mp

186 ° C]

2.6) N- [N ^α - (4-Hydroxy-phenylamino-thiocarbonyl) -ornithyl] -quinolone-a, hydrochloride

Educt: N- [N - (fluorenyl-9-methoxycarbonyl) -ornithyl] -quinolone-a, trifluoroacetate salt-free precursor: 47% over 2 stages [TLC (dichloromethane / methanol / ammonia 17% 10: 10: 3 ): R _f = 0.36; Mp: 211 ° C (dec.)] Hydrochloride: The compound is suspended in water and the pH

2.7) N- [N ^α - (phenylamino-thiocarbonγI) -lysyl] -quinolone-a educt: N- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl] -quinolone-a, trifluoroacetate 98/15571 37 -

Educ. 58% over 2 steps [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2): R _f = 0.48]

2.8) N- [N ^α - (4-isothiocyanato-phenylamino-thiocarbonyI) -Iysyl] - quinolone-a educt: N- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl] -quinolone-a,

Trifluoroacetate

Yield: 73% over 2 steps [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2): R _f =

0.38]

2.9) N- [N - (4-carboxy-phenylamino-thiocarbonyI) -lysyl] -quinolone-a educt: N- [N * - (fluorenyl-9-methoxycarbonyl) -lysyl] -quinolone-a,

Trifluoroacetate yield: 62% over 2 steps [TLC (acetonitrile / water / glacial acetic acid 10: 3: 1.5):

R _f = 0.6]

2.10) N- [N ^α - (phenyl-methylamino-thiocarbonyI) -lysγl] -chinoIon-a

Educt: N- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl] -quinolone-a,

Trifluoroacetate

Yield: 59% over 2 steps [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2): R _f ^■

0.44]

Example 2.11

Quinolone-a conjugates with two amino acids; general formula:

COOH

2.11) N- [N ^α - (4-hydroxyphenylamino-thiocarbonyl) lysyl-D-alanyl] quinolone-a

Starting material: N- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-D-alanyl] -quinolone-a

Trifluoroacetate yield: 53% over 2 steps [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2) R _f =

0.33]

Example 3

Doxorubicin conjugates; general formula:

3) N- [N ^α - (4-HydroxyphenyIamino-thiocarbonyI) -lysyI-aIanyI] - doxorubicin

Educt: N- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-alanyl] -doxorubicin,

Trifluoroacetate,

Yield: 46% over 2 steps [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2): R _f =

0.2; FAB-MS: m / z = 894 (M + H) ⁺ ] 98/15571 39 -

Examples 4.1 - 4.11

Conjugates of 20 (S) camptothecin; general formula:

4.1) 20-O- [N ^α - (4-Hydroxyphenylamino-thiocarbonyl) lysyl alanyl] camptothecin

Educt: 20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-alanyl] -camptothecin,

Trifluoroacetate yield: 80% over 2 steps [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2): R _f =

0.32]

4.2) 20-O- [N ^α - (4-Hydroxy-phenylamino-thiocarbonyl) -lysyI-Ieucyl] - camptothecin, hydrochloride

Educt: 20-O- [ ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-leucyl] -camptothecin,

Trifluoroacetate salt-free precursor: 71% over 2 stages [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2): R _f = 0.48]

Hydrochloride: The compound is dissolved in dioxane / water and converted into the hydrochloride with one equivalent of 0.1 N hydrochloric acid. The resulting solution is then lyophilized.

4.3) 20-O- [N ^α - (4-Hydroxy-phenylamino-thiocarbonyl) -Iysyl-phenylalanylj-camptothecin

Starting material: 20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-phenylalanyl] - camptothecin, trifluoroacetate 98/15571 40 -

Yield: 75% over 2 steps [TLC (acetonitrile / water / glacial acetic acid 5: 1: 0.2): R _f =

0.33]

4.4) 20-O- [N ^α - (4-hydroxy-phenylamino-thiocarbonyl) -IysyI-valyl] - camptothecin, hydrochloride reactant: 20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-valyl] -camptothecin,

Trifluoroacetate salt-free precursor: 68% over 2 stages [TLC (acetonitrile / water / glacial acetic acid

5: 1: 0.2): R _f = 0.35; FAB-MS: m / z - 727 (M + H) ⁺ ] hydrochloride: The compound is dissolved in dioxane / water and converted into the hydrochloride with an equivalent of 0.1 N hydrochloric acid. The resulting solution is then lyophilized.

4.5) 20-O- [N ^α - (4-carboxy-phenylamino-thiocarbonyl) -lysyl-valyl] - camptothecin, hydrochloride reactant: 20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-valyl] -camptothecin,

Trifluoroacetate salt-free precursor: 79% over 2 stages [TLC (acetonitrile / water / glacial acetic acid

5: 1: 0.2): R _f = 0.46] hydrochloride: The compound is dissolved in dioxane / water and converted into the hydrochloride with an equivalent of 0.1 N hydrochloric acid. The resulting solution is then lyophilized.

4.6) 20-O- [N - (4-chloro-phenylamino-thiocarbonyl) -lysyl-valyl] - ca ptothecin, hydrochloride reactant: 20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-valyl] -camptothecin,

Trifluoroacetate salt-free precursor: 86% over 2 stages [TLC (acetonitrile / water / glacial acetic acid

10: 1: 0.1): R _f = 0.24] hydrochloride: The compound is dissolved in dioxane / water and converted into the hydrochloride with an equivalent of 0.1 N hydrochloric acid. The resulting solution is then lyophilized. 98/15571 41

4.7) 20 ~ O- [N ^α - (phenylamino-thiocarbonyl) -Iysyl-valyI] -camptothecin,

Hydrochloride

Educt: 20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-valyl] -camptothecin,

Trifluoroacetate salt-free precursor: 67% over 2 stages [TLC (acetonitrile / water / glacial acetic acid

5: 1: 0.2): R _f = 0.5] hydrochloride: The compound is dissolved in dioxane / water and converted into the hydrochloride with an equivalent of 0.1 N hydrochloric acid. The resulting solution is then lyophilized.

4.8) 20-O- [N ^α - (phenyl-methyl-aminothiocarbonyI) -lysyl-valyl] - camptothecin, hydrochloride

Trifluoroacetate salt-free precursor: 55% over 2 stages [TLC (acetonitrile / water / glacial acetic acid

4.9) 20-O- [N ^α , N ^ε -bis- (4-hydroxy-phenylamino-thiocarbonyI) -IysyI-alanyl] -camptothecin

Starting material: 20-O- (lysyl-alanyl) camptothecin, di-trifluoroacetate

Yield: 64% [TLC (acetonitrile / water 10: 1): R _f = 0.72]

4.10) 20-O- [N ^α , N ^ε -bis- (4-hydroxy-phenylamino-thiocarbonyI) -lysyI-D-alanylj-camptothecin

Starting material: 20-O- (lysyl-D-alanyl) camptothecin, di-trifluoroacetate

Yield: 77% [TLC (acetonitrile / water 20: 1): R _f = 0.40; FAB-MS: m / z =

850 (M + H) ⁺ ]

4.11) 20-O- [N ^α , N ^ε -bis- (4-hydroxy-phenylamino-thiocarbonyl) -iysyl-phenylalanyl] -camptothecin

Starting material: 20-O- (lysylphenylalanyl) camptothecin, di-trifluoroacetate

Yield: 84% [TLC (acetonitrile / water 20: 1): R _f = 0.6] 4.12) 20-O- [N ^α - (3-Hydroxy-phenylamino-thiocarbonyl) -lysyI-valyI] - camptothecin, hydrochloride

Trifluoroacetate salt-free precursor: 58% over 2 stages [TLC (acetonitrile / ethyl acetate 1: 1): R _f =

0.03; Mp = 195 ° C (dec.); FAB-MS: m / z = 727 (M + H) ⁺ ] hydrochloride: water is added to the compound and the suspension is acidified to pH 2 with 1N hydrochloric acid. The resulting solution is filtered through Celite and then lyophilized.

4.13) 20-O- [N ^α - (2-HydroxyphenylIamino-thiocarbonyl) -Iysyl-vaIyl] - camptothecin, hydrochloride

Trifluoroacetate salt-free precursor: 36% over 2 steps [TLC (acetonitrile / ethyl acetate 1: 1): R _f =

0.03; Mp = 192 ° C (dec.); FAB-MS: m / z = 727 (M + H) ⁺ ] hydrochloride: water is added to the compound and the suspension is acidified to pH 2 with 1N hydrochloric acid. The resulting solution is filtered through Celite and then lyophilized.

4.14) 20-O- [N ^α - (4-methoxyphenylamino thiocarbonyl) lysyl valyl] camptothecin, hydrochloride

Trifluoroacetate salt-free precursor: 54% over 2 stages [TLC (acetonitrile / ethyl acetate 1: 1): R _f =

0.06; Mp = 195 ° C (dec.); FAB-MS: m / z = 741 (M + H) ⁺ ] hydrochloride: water is added to the compound and the suspension is acidified to pH 2 with 1N hydrochloric acid. The resulting solution is filtered through Celite and then lyophilized.

4.15) 20-O- [N - (3-methoxy-phenylamino-thiocarbonyl) -lysyl-valyI] - camptothecin, hydrochloride

Trifluoroacetate 98/15571

- 43

Salt-free precursor: 65% over 2 stages [TLC (acetonitrile / ethyl acetate 1: 1): R _f =

0.08; Mp = 197 ° C (dec.); FAB-MS: m / z = 741 (M + H) ⁺ ] hydrochloride: water is added to the compound and the suspension is acidified to pH 2 with 1N hydrochloric acid. The resulting solution is filtered through Celite and then lyophilized.

4.16) 20-O- [N ^α - (4-nitro-phenylamino-thiocarbonyI) -Iysyl-valyl] - camptothecin, hydrochloride

Educt: 20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-valyl] -camptothecin, trifluoroacetate salt-free precursor: 86% over 2 steps [TLC: (acetonitrile / water / glacial acetic acid

5: 1: 0.2): R _f = 0.5]. Hydrochloride: The compound is dissolved with dioxane / water and converted into the hydrochloride with one equivalent of 0.1N hydrochloric acid. The resulting solution is then lyophilized.

4.17) 20-O- [N ^α - (3-nitro-phenylamino-thiocarbonyI) -lysyl-valyI] - camptothecin, hydrochloride

Educt: 20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-valyl] -camptothecin, trifluoroacetate salt-free precursor: 46% over 2 stages. The purification on the Fmoc-protected intermediate stage is carried out by flash chromatography on silica gel with dichloromethane / methanol 50: 1). It is then deblocked with piperidine. [TLC: (acetonitrile / water glacial acetic acid 5: 1: 0.2): R _f = 0.45;

Hydrochloride: The compound is dissolved in dioxane / water and converted into the hydrochloride with one equivalent of 0.1 N hydrochloric acid. The resulting solution is then lyophilized. [FAB-MS: m / z = 756 (M + H) ⁺ ].

4.18) 20-O- [N - (4-Amino-phenylamino-thiocarbonyI) -lysyl-valyl] - camptothecin, hydrochloride

Trifluoroacetate mono-Fmoc-protected p-phenylene diamine: 8/15571

44

This is made from phenylenediamine with 0.5 Aq Fmoc-Cl without further base addition. It is then transferred to the Senfol according to standard conditions. Salt-free preliminary stage 46% over two stages Dichloromethane / methanol 50 1). This is followed by deblocking with piperidine. The purification is then carried out again by flash chromatography on silica gel with dichloromethane / methanol / ammonia 1 7% 1 5: 1 .0, 1) [TLC (acetonitrile / water / glacial acetic acid 5 1: 0.2) R _f = 0.45]

Hydrochloride: The compound is dissolved in dioxane / water and converted into the hydrochloride with an equivalent of 0.1 N hydrochloric acid. The resulting solution is then lyophilized [FAB-MS "m / z = 726 (M + H) ⁺ ]

4.19) 20-O- [N ^α - (4-Hydroxy-phenylamino-thiocarbonyI) -histidyl-valyl] - camptothecin, hydrochloride

Educt 20-O- [histidylvalyl] camptothecin, trifluoroacetate salt-free precursor 81% [DC. (Acetonitrile / water 10 1 R _f = 0.4] hydrochloride The compound is dissolved in dioxane / water and converted into the hydrochloride with an equivalent of 0.1 N hydrochloric acid. The resulting solution is then lyophilized

4.20) 20-O- [N ^α - (4-hydroxyphenylamino-thiocarbonyl) lysyl-leucyl] camptothecin, hydrochloride

Educt 20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-valyl] -camptothecin,

Trifluoroacetate salt-free precursor 71% over 2 stages [TLC (acetonitrile / water / glacial acetic acid

5 1 0.2) R _f = 0.45] hydrochloride The compound is dissolved in dioxane / water and transferred with an equivalent of 0.1 N hydrochloric acid to hydrochloride. The resulting solution is then lyophilized 98/15571 45

4.21) 20-Oi [N ^α -N ^e -bis- (4-hydroxy-phenylamino-thiocarbonyI)] - lysyl-valylJ-camptothecin, hydrochloride

Educt: 20-O- ιLysyI- [N ^ε - (Fluorenyl-9-methoxycarbonyl) -lysyl] -valylJ- camptothecin, bis-trifluoroacetate salt-free precursor: 79% over 2 steps.

[TLC: (acetonitrile / water / glacial acetic acid 5: 1: 0.2) R _f = 0.46]; [FAB-MS: m / z = 1006 = (M + H) ⁺ ]. Hydrochloride: The compound is dissolved in dioxane / water and converted into the hydrochloride with one equivalent of 0.1 N hydrochloric acid. The resulting solution is then lyophilized.

4.22) 20-O- [N ^α , N ^ε -bis- (4-hydroxy-phenylamino-thiocarbonyl) -lysyl-aspartyI] -camptothecin, sodium salt

Educt: 20-O- (lysyl-aspartyl) -camptothecin, di-hydrobromide salt-free precursor: 50% - the cleaning is carried out by repeatedly falling over from dichloromethane / methanol 1: 1 with diethyl ether [TLC (acetonitrile / water 5: 1): R _f = 0.58; Mp = 192 ° C (dec.); FAB-MS: m / z = 894 (M + H) ⁺ ]. Sodium salt: The compound is suspended in water and one equivalent of a 0.1 N sodium hydroxide solution is added. The resulting solution is then lyophilized.

4.23) 20-O- [N ^α , N ^ε -bis- (4-hydroxy-phenylamino-thiocarbonyI) -lysyl-seryl] -camptothecin

Educt: 20-O- (lysylseryl) camptothecin, di-hydrobromide

Yield: 36% - the purification is carried out by flash chromatography [petroleum ether / ethyl acetate 2: 1 → 1: 2 → ethyl acetate] [TLC (acetonitrile): R _f = 0.70; Mp = 183 ° C (dec.); FAB-MS: m / z = 866 (M + H) ⁺ ]. 98/15571 - 46

Example 5

Conjugates of 20 (S) -7-ethyl camptothecin; general formula:

5.1) 7-EthyI-20-O- [N ^α , N ^ε -bis- (4-hydroxy-phenylamino-thiocarbonyl) - Iysyl-alanyl] -camptothecin

Educt: 7-ethyl-20-O- (lysyl-alanyl) camptothecin, di-trifluoroacetate

Yield: 27% [TLC (acetonitrile): R _f = 0.68; Mp - 122 ° C (dec.);

FAB-MS: m / z = 879 (M + H) ⁺ ].

5.2) 7-EthyI-20-O- [N - (4-hydroxy-phenylamino-thiocarbonyl) -lysyl-vaIyI] - camptothecin, hydrochloride

Educt: 7-ethyl-20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) -lysyl-valyl] - camptothecin, trifluoroacetate salt-free precursor: 61% over 2 steps [beige crystals; TLC (acetonitrile / ethyl acetate 1: 1): R _f = 0.02; Mp = 220 ° C (dec.); FAB-MS: m / z = 755 (M + H) ⁺ ].

Hydrochloride: water is added to the compound and the suspension is acidified to pH 2 with 1N hydrochloric acid. The resulting solution is filtered through Celite and then lyophilized. 98/15571

- 47

Example 6

Conjugates of 10, ll- (methylcndioxy) camptothecin; general formula:

6) 10.1 l- (Methylenedioxy) -20-O- [N ^α - (4-Hydroxy-phenylaminothio-carbonyI) -lysyl-leucyI] -camptothecin, hydrochloride

Educt: 10, l l- (methylenedioxy) -20-O- [N ^e - (fluorenyl-9-methoxy-carbonyl) - lysyl-leucyl] -camptothecin, trifluoroacetate salt-free precursor: 90% over 2 stages [DC: (acetonitrile / Water / glacial acetic acid

5: 1: 0.2) R _f = 0.43] hydrochloride: The compound is dissolved in dioxane / water and converted into the hydrochloride with an equivalent of 0.1 N hydrochloric acid. The resulting solution is then lyophilized.

Example 7

Conjugates of 7-hydroxymethyl camptothecin; general formula

7) 7- [N - (4-Hydroxy-phenylamino-thiocarbonyl) lysyl-valyloxymethyl] camptothecin, hydrochloride

Educt: 7- [N ^ε - (Fluorenyl-9-methoxy-carbonyl) -lysyl-valyloxymethyl] - camptothecin, trifluoroacetate salt-free precursor: 60% over 2 stages. The purification on the Fmoc-protected intermediate stage is carried out by flash chromatography on silica gel with dichloromethane / methanol 20: 1). It is then deblocked with piperidine. [TLC: (acetonitrile / water / glacial acetic acid 5: 1: 0.2) R _f = 0.54]

Example 8

Conjugates of 20 (S) -7-ethyl-10-hydroxy camptothecin; general formula:

8) 7-Ethyl-10-hydroxy-20-O- [N ^α - (4-hydroxy-phenylamino-thiocarbonyl) -

Iysyl-valyl] -camptothecin, hydrochloride

Educt: 7-ethyl-20-O- [N ^ε - (fluorenyl-9-methoxycarbonyl) lysyl-valyl] -10-hydroxy-camptothecin, trifluoroacetate salt-free precursor: 69% over 2 steps [beige crystals; TLC (acetonitrile / ethyl acetate 1: 1): R _f - 0.03; Mp = 225 ° C (dec.); FAB-MS: m / z = 771 (M + H) ⁺ ] hydrochloride: water is added to the compound and the suspension is acidified to pH 2 with 1N hydrochloric acid. The resulting solution is filtered through Celite and then lyophilized.

Claims

98/15571 _ _5Q

claims

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

S

II

(Ar-X-NH-C-) _n MC

wherein

S

II

^ ' ⁿ for 1 to n' identical or different groups

S

II pations Ar- ^λ -NH-C, where n is a number 1 to n 'and n'

corresponds to the maximum number of possible connecting points of M,

wherein

Ar stands for an aryl radical with up to 10 carbon atoms which, in addition to X, can optionally be mono- or polysubstituted by alkyl with up to 6 carbon atoms, alkoxy with up to 6 carbon atoms, alkoxycarbonyl with up to 6 carbon atoms, hydroxy, carboxy, carboxylalkyl with up to 6 carbon atoms, cyano, nitro, isocyanato, isothiocyanato, halogen, sulfonyl and / or sulfonamide,

X for a direct single bond or for alkylene with up to 6

Carbon atoms,

M stands for a mono-, di-, tri- or tetrapeptide, which via the α-

Amino group and / or via amino and / or hydroxy groups of the side chains with the n same or different

S

II

Groupings Ar- ^Λ -NH-C is linked, with further 98/15571 _{5 1}

functional groups of the peptide can optionally carry protective groups, and C for a residue of a cytostatic or a cytostatic

Is a derivative which is linked to M via an amino function or via an oxygen atom,

and their stereoisomers, stereoisomer mixtures and salts.

2. Compounds according to claim 1, characterized in that

Ar represents a phenyl radical which can also carry hydroxy, carboxy, isothiocyanato or halogen para to X,

and their stereoisomers, stereoisomer mixtures and salts.

3. Compounds according to one of claims 1 or 2, characterized in that

X represents a single bond or methylene,

and their stereoisomers, stereoisomer mixtures and salts.

4. Compounds according to one of claims 1, 2 or 3, characterized in that

M stands for a mono-, di or tripeptide which, via the α-amino group and / or amino and / or hydroxy groups of the side chains with the 1 to n identical or different groups S

II Ar-X-NH-C is linked, with further functional groups

of the peptide can optionally carry protective groups.

and their stereoisomers, stereoisomer mixtures and salts.

5. Compounds according to any one of claims 1 to 4, characterized in that the peptides M consist of amino acid residues which are ab- lead from alanine, aspartic acid, glutamic acid, glycine, leusine, htstidine, lysine, arginine, ornithine, senn, tyrosine, valine or diaminopropionic acid, with several amino acid residues both via the α-amino group and optionally via the side chain amino functions and also via both functions can be linked peptically,

and their stereoisomers, stereoisomer mixtures and salts

Compounds according to any one of claims 1 to 5, characterized in that C for a batracylin, methotrexate, quinolone a, etoposide, melphalan, taxol, camptothecin residue, a mode in the A ring or B ring - Defected Camptothecin-Deπvat, a daunomycin or doxorubicin residue is available, where C is linked to M via an amino or hydroxy function

and their stereoisomers, stereoisomer mixtures and salts

Process for the preparation of compounds of the general formula (I) according to Claim 1, characterized in that compounds of the general formula (II)

M'-C (II),

wherein C has the meaning given in claim 1 and M ^{1 stands} for a radical M defined in claim 1 which carries hydrogen atoms at the desired linking sites and the remaining potential linking sites are blocked by protective groups,

with compounds of the general formula (III)

Ar - X- N = C = S (III)

S

I I (Ia)

Ar-X-NH-CI implements

and in the case of the introduction of further groups

S

II

Ar-X H, which differs from the one (s) first introduced

differ, the corresponding protective groups optionally selectively split off from the compounds of formula (Ia), these in the manner indicated above with other compounds of general formula (III), which differ from those initially introduced, and optionally this reaction sequence for the introduction of further the

S

II repeated residues of various residues Ar-X-NH-C,

and that any remaining protective groups are split off,

that the stereoisomers are furthermore optionally separated by customary methods and that the compounds are optionally converted into their salts.

8. Use of the compounds of general formula (I) according to claim 1 for the manufacture of medicaments.

9. Medicament containing compounds of general formula (I) according to claim 1.