WO2003064424A2 - Water-soluble porphyrin platinum compounds with high tumor selectivity and their use for the treatment of benign and malignant tumor diseases - Google Patents

Water-soluble porphyrin platinum compounds with high tumor selectivity and their use for the treatment of benign and malignant tumor diseases Download PDF

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WO2003064424A2
WO2003064424A2 PCT/EP2003/000874 EP0300874W WO03064424A2 WO 2003064424 A2 WO2003064424 A2 WO 2003064424A2 EP 0300874 W EP0300874 W EP 0300874W WO 03064424 A2 WO03064424 A2 WO 03064424A2
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alkyl
cycloalkyl
hetarylalkyl
hetaryl
aralkyl
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PCT/EP2003/000874
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WO2003064424A3 (en
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Karl-Christian Bart
Günther BERNHARDT
Henri Brunner
Christian Lottner
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Zentaris Gmbh
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Priority to KR10-2004-7011734A priority Critical patent/KR20040083098A/en
Priority to JP2003564047A priority patent/JP2005522429A/en
Priority to NZ534541A priority patent/NZ534541A/en
Priority to EP03734604A priority patent/EP1470139A2/en
Priority to UA20040806544A priority patent/UA78007C2/en
Priority to BR0307400-5A priority patent/BR0307400A/en
Publication of WO2003064424A2 publication Critical patent/WO2003064424A2/en
Publication of WO2003064424A3 publication Critical patent/WO2003064424A3/en
Priority to HRP20040788 priority patent/HRP20040788A2/en
Priority to NO20043650A priority patent/NO20043650L/en
Priority to HK05110413A priority patent/HK1078585A1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
    • C07F15/00—Compounds containing elements of Groups 8, 9, 10 or 18 of the Periodic System
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
    • C07F15/00—Compounds containing elements of Groups 8, 9, 10 or 18 of the Periodic System
    • C07F15/0006—Compounds containing elements of Groups 8, 9, 10 or 18 of the Periodic System compounds of the platinum group
    • C07F15/0086—Platinum compounds
    • C07F15/0093—Platinum compounds without a metal-carbon linkage
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/28—Compounds containing heavy metals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the invention relates to novel, water-soluble porphyrin platinum compounds with high tumor selectivity and their use for the treatment of benign and malignant tumor diseases.
  • the inventive compounds are suitable for photodynamic anti- tumor therapy in man and mammals.
  • novel porphyrin platinum derivatives are described, which have cytotoxic properties. Surprisingly, the compounds have good water solubility and a high selectivity.
  • the compounds can be used for the treatment of cancer and, in particular, for the photodynamic treatment of tumors.
  • R2/R3 H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
  • R4 H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
  • R5 H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst. Amino
  • R6 H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst. Amino Formula II
  • R1/R2/R3/R4 H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
  • R5 H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl R6: H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst.
  • Amino R7 H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst.
  • Amino R8 H, Alkyl
  • R2/R3 H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
  • R4 H, Alkyl, Cycloalkyl
  • R5 H, Alkyl, Cycloalkyl
  • R6 H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
  • inventive compounds have at least one center of asymmetry, they can be in the form of their racemates, their pure enantiomers and/or their diastereoisomers or in the form of mixtures of these enantiomers or diastereoisomers.
  • the inventive compounds exhibit cytotoxic activity in selected tumor cell lines.
  • the antitumor activity is intensified by irradiating with electromagnetic radiation having a wavelength of 600 to 730 nm.
  • the invention accordingly relates to the chemical combination of the cytotoxic principle of the platinum compounds of the cis platinum type with a photodynamically active molecule of the porphyrin derivative type, in such a manner, that compounds of good water solubility and high selectivity are obtained.
  • inventive compounds can be administered intraaterially, intracerebrally, intramuscularly, intraperitoneally, intrathecally, intravenously, orally, parenterally, intranasally, rectally, subcutaneously and/or topically in the form of tablets, film- coated tablets, capsules, coated tablets, powders, granulates, drops, syrups, ointments, powders for inhalation, infusion solutions, drinking solutions or in some other suitable form.
  • the medicaments comprise one or more compounds in addition to customary physiologically tolerable carriers and/or diluents or auxiliaries.
  • the process for the production of the medicament is characerized in that one or more compounds are processed to give pharmaceutical preparations or brought into a therapeutically administrable form using customary pharmaceutical carriers and/or diluents or other auxiliaries.
  • Ethyl DL-2,3-diaminopropionate dihydrochloride, ethyl L-2,4-diaminobutanoate dihydrochloride and diethyl meso-4,5-diaminosuberate dihydrochloride were synthesized according to literature procedures and used as ligands for the preparation of the corresponding diiodoplatinum(ll) complexes.
  • the diamine(dichloro)platinum(ll) fragments were activated by conversion into diamine(dihydroxy)platinum(ll) species, which were reacted with an equimolar amount of the respective porphyrin malonic acid in a mixture of CH 2 CI 2 , ethanol and water or, in the case of the water-soluble ligand, in pure water.
  • the complexes precipitated.
  • CH 2 CI 2 was added to remove neutral impurities, before the aqueous phase was evaporated to obtain the product.
  • the etherified hematoporphyrin esters were purified by column chromatography.
  • the carboxylic acids which were required for coordination to the platinum(ll) moieties, were prepared by hydrolysis of the esters with 20 % methanolic KOH solution.
  • 1 ,2-Diaminoethane, 1 ,3-diaminopropane, trans-1 -2-diaminocyclohexane and 2,2'-bi- pyridine were commercially available and used as ligands to prepare the corresponding dichloroplatinum(ll) complexes according to literature procedures.
  • Ethyl DL-2,3-diaminopropionate dihydrochloride, ethyl L-2,4-diaminobutanoate dihydrochloride and diethyl meso-4,5-diaminosuberate dihydrochloride were synthesized according to literature procedures and used as ligands for the preparation of the corresponding diiodoplatinum(ll) complexes.
  • the diamine(dichloro)platinum(ll) precursors were activated by conversion into diamine(dihydroxy)platinum(ll) species, which were reacted with an equimolar amount of the respective porphyrin carboxylic acid in a mixture of ethanol and water or, in the case of the water-soluble ligands, in pure water.
  • the complexes precipitated.
  • CH 2 CI 2 was added to remove neutral impurities and the aqueous phase was evaporated to obtain the product.
  • inventive compounds are tetraarylporphyrin platinum derivatives, covered by way of example by examples 1 and 2, and hematoporphyrin platinum derivatives, covered by way of example by examples 3, 4 and 5.
  • Diammine[2-(4- ⁇ 10,15,20-tris[4-(1 ,4,7-trioxaoctyl)phenyl]porphyrin-5- yl ⁇ phenoxy)malonato]platinum(ll) (No. 21 in Figure 1)
  • the compound 2-(4- ⁇ 10,15,20-Tris[4-(1 ,4,7-trioxaoctyl)phenyl]porphyrin-5- yl ⁇ phenoxy)malonic acid (109 mg, 0.100 mmol) was dissolved in 10 ml of CHCI 3 and
  • Diammine ⁇ 7 12-bis[1-(1 ,4,7-trioxaoctyl)ethyl]-3,8,13 I 17-tetramethylporphyrin-2,18- dipropionato ⁇ platinum(ll) (No. 21 in Figure 2).
  • cytotoxic effect was obtained, for instance, on the human tumor cell lines TCC-SUP and J82.
  • the effect of the compounds was investigated in the dark and under irradiation with light at a wavelength of 600 - 730 nm. Selected compounds are clearly more activ cytotoxically under irradiation. There is a synergism between the cytotoxic effect of the platinum component and the photodynamic principle.
  • both the dark- and phototoxicity of the porphyrin-platinum conjugates are influenced by the type of the non-leaving group.
  • the platinum complexes with 2,2'-bipyridyl (40, 41), ethyl DL-2,3-diaminopropionate (42-46), ethyl DL-2,3-diaminobutanoate (47-51), diethyl meso-4,5-diaminosuberate (52-55) ligands were inactive at a concentration of 1 ⁇ M, both in the dark and after irradiation.
  • the compounds bearing 1 ,2-diaminoethane (27-30) and 1 ,2-diaminopropane (31-34) non-leaving groups were also inactive against TCC-SUP cells.
  • the most interesting porphyrin-platinum conjugates were those with the diammine (21-26) and the (+)- trans- ,2-diaminocyclohexane (35-39) ligands.
  • the water-soluble complexes 26 and 39 were most active with T/C ⁇ rr of around 30 %
  • the reference cisplatin had a T/C ⁇ rr value of approximately 2 %. At this dosage there was no statistically significant enhancement of the cytotoxicity by irradiation of the bladder cancer cells.
  • both the dark- and phototoxicity of the tetraarylporphyrin-platinum conjugates 21-38 were highly influenced by the type of the non-leaving group the results agreeing with those of the hematoporphyrin- platinum complexes discussed above.
  • 23, 29 and 30 were the most active tetraarylporphyrin-platinum conjugates with T/C ⁇ rr values of around 37 %, 57 % and 63 %, respectively, at 1 ⁇ M concentration.

Abstract

The invention relates to novel, water-soluble porphyrin platinuim compounds of the tetraarylporphyrin platinum derivatives type or of the hematoporphyrin platinum derivatives type with high tumor selectivity and their use for the treatment of benign and malignant tumor diseases. In particular, the compounds are suitable for photodynamic anti-tumor therapy.

Description

Novel, water-soluble porphyrin platinum compounds with high tumor selectivity and their use for the treatment of benign and malignant tumor diseases
Introduction
The invention relates to novel, water-soluble porphyrin platinum compounds with high tumor selectivity and their use for the treatment of benign and malignant tumor diseases. In particular, the inventive compounds are suitable for photodynamic anti- tumor therapy in man and mammals.
Prior art
Platinum(ll) complexes with porphyrin ligands and their application as potent cytostatic and phototoxic antitumor agents have already been described in the following publications.
W.M. Sharman, CM. Allen and J.E. van Lier, DDT 4, (11) 507-517 (1999). Photodynamic therapeutics: basic principles and clinical applications
T. Okunaka and H. Kato, Rev. Contemp. Pharmacother., 10, 59-68 (1999). Potential Applications of Photodynamic Therapy.
H. Brunner, H. Obermeier and R.-M. Szeimies, Chem. Ber., 1995, 128, 173-181. Platinum(ll) complexes with porphyrin ligands: synthesis and synergism during photodynamic therapy.
H. Brunner, K.-H. Schellerer and B. Treittinger, Inorg. Chim. Acta 1997, 264, 67-69. Synthesis and in vitro testing of hematoporphyrin type ligands in platinum(ll) complexes as potent cytostatic and phototoxic antitumor agents. Description of the invention
In the invention, novel porphyrin platinum derivatives are described, which have cytotoxic properties. Surprisingly, the compounds have good water solubility and a high selectivity. The compounds can be used for the treatment of cancer and, in particular, for the photodynamic treatment of tumors.
The general formulas of the claimed compounds of the tetraarylporphyrin platinum derivatives type are: Formula I
Figure imgf000003_0001
n: 1-20
X: O, S, NH, N-Alkyl
R2/R3: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
R4: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
R5: H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst. Amino
R6: H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst. Amino Formula II
Figure imgf000004_0001
n: 1-20
X: O, S, NH, N-Alkyl
R1/R2/R3/R4: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
Oder R2-Z-R3, mit Z: (CH2)n, n=0-6
R1/R4: H, -(CH2)n-COOR8, n=0-6
R5: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl R6: H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst. Amino R7: H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst. Amino R8: H, Alkyl
The general formulas of the claimed compounds of the hematoporphyrin platinum derivatives type are:
Formula III
Figure imgf000005_0001
R2/R3: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
R4: H, Alkyl, Cycloalkyl
R5: H, Alkyl, Cycloalkyl
R6: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
Formula IV
Figure imgf000006_0001
R1/R2/R3/R4: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl oder R2-2-R4, mit 2: (CH2)n, n=0-6
Oder R1/R3: H, -(CH2)n-COOR6, n=0-6 R4/R5: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl R6: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl R7: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
If the inventive compounds have at least one center of asymmetry, they can be in the form of their racemates, their pure enantiomers and/or their diastereoisomers or in the form of mixtures of these enantiomers or diastereoisomers.
The inventive compounds exhibit cytotoxic activity in selected tumor cell lines. The antitumor activity is intensified by irradiating with electromagnetic radiation having a wavelength of 600 to 730 nm. The invention accordingly relates to the chemical combination of the cytotoxic principle of the platinum compounds of the cis platinum type with a photodynamically active molecule of the porphyrin derivative type, in such a manner, that compounds of good water solubility and high selectivity are obtained. The inventive compounds can be administered intraaterially, intracerebrally, intramuscularly, intraperitoneally, intrathecally, intravenously, orally, parenterally, intranasally, rectally, subcutaneously and/or topically in the form of tablets, film- coated tablets, capsules, coated tablets, powders, granulates, drops, syrups, ointments, powders for inhalation, infusion solutions, drinking solutions or in some other suitable form.
The medicaments comprise one or more compounds in addition to customary physiologically tolerable carriers and/or diluents or auxiliaries.
The process for the production of the medicament is characerized in that one or more compounds are processed to give pharmaceutical preparations or brought into a therapeutically administrable form using customary pharmaceutical carriers and/or diluents or other auxiliaries.
The synthesis of the inventive compounds is described.
Tetraarylporphyrin platinum derivatives
Synthesis of the substituted benzaldehydes. For the reaction with 4-hydroxy- benzaldehyde the respective oligo- and polyethyleneglycol monomethylethers had to be activated at their alcohol terminus with tosyl chloride according to a literature procedure. The etherification was performed by refluxing the tosylated alcohols and
4-hydroxybenzaldehyde together with KjCO., in DMF. The substituted benzaldehydes were separated by filtration and purified by column chromatography.
For platinum coordination to the tetraarylporphyrins to be synthesized it is necessary to introduce two adjacent carboxylic acid groups in one of the substituted benzaldehydes. Therefore, 4-hydroxybenzaldehyde was etherified with diethyl bromomalonate under alkaline conmditions. The diethyl 2-(4- formylphenoxy)malonate was used together with the substituted benzaldehydes for the synthesis of asymmetric tetraarylporphyrins.
Synthesis of the porphyrin ligands. The synthesis of the asymmetric tetraarylporphyrins was performed using the Lindsey method. Pyrrol and the respective benzaldehydes were reacted under Lewis acid catalysis to porphyrinogens, which were oxidized with p-chloranil to the corresponding porphyrins. The tetraarylpoφhyrin esters were purified by several column chromatographies. The carboxylic acids, which were required for coordination to the platinum(ll) fragments, were prepared by hydrolysis of the esters with a mixture of
CHCI3 and 20 % methanolic KOH solution or pure 20 % methanolic KOH solution only.
Synthesis of the platinum fragments. 1 ,2-Diaminoethane, 1 ,3-diaminopropane, trans-1 ,2-diaminocyclohexane and 2,2'-bipyridine were commercially available and used as ligands to prepare the corresponding dichloroplatinum(ll) complexes according to literature procedures. Ethyl DL-2,3-diaminopropionate dihydrochloride, ethyl L-2,4-diaminobutanoate dihydrochloride and diethyl meso-4,5-diaminosuberate dihydrochloride were synthesized according to literature procedures and used as ligands for the preparation of the corresponding diiodoplatinum(ll) complexes.
Synthesis of the platinum complexes. For the reaction with the porphyrincarboxylic acids cisplatin had to be activated by conversion into diammine(diaqua)platinum(ll) hydroxide. It was reacted with an equimolar amount of the porphyrin ligand in a mixture of CHCI3, ethanol and water or, in the case of the water-soluble ligand, in pure water. The resulting diammine(malonato)platinum(ll) complexes precipitated. To the reaction mixture of the water-soluble complex CH2CI2 was added to remove neutral impurities. The aqueous hphase was evaporated to obtain the product. The diamine(dichloro)platinum(ll) fragments were activated by conversion into diamine(dihydroxy)platinum(ll) species, which were reacted with an equimolar amount of the respective porphyrin malonic acid in a mixture of CH2CI2, ethanol and water or, in the case of the water-soluble ligand, in pure water. The complexes precipitated. To the water-soluble complex CH2CI2 was added to remove neutral impurities, before the aqueous phase was evaporated to obtain the product.
For the reaction with the porphyrinmalonic acids it was necessary to activate the diamine(diiodo)platinum(ll) complexes by conversion into diamine(dinitrato) platinum(ll) species, which are water-soluble. In this form they were reacted with an equimolar amount of the porphyrin ligands, in a mixture of CH2CI2, ethanol and water. The water-insoluble complexes precipitated after concentrating the solutions.
Hematoporphyrin platinum derivatives type
Synthesis of the porphyrin ligands and the platinum precursors. Hemin was transferred to protoporphyrin dimethylester, from which all the subsequent reactions started. First, protoporphyrin dimethylester was treated with 30 % hydrobromic acid in acetic acid to give the labile Markownikoff adduct of HBr to the two vinyl double bonds, which was reacted with different types of alcohols to replace bromide by the corresponding alkoxides. As alcohols we chose hydrophilic oligo- and polyethyleneglycol monomethylethers. During the etherification the HBr formed catalyzed the transesterification of the methylesters into the esters of the corresponding alcohols. The etherified hematoporphyrin esters were purified by column chromatography. The carboxylic acids, which were required for coordination to the platinum(ll) moieties, were prepared by hydrolysis of the esters with 20 % methanolic KOH solution. 1 ,2-Diaminoethane, 1 ,3-diaminopropane, trans-1 -2-diaminocyclohexane and 2,2'-bi- pyridine were commercially available and used as ligands to prepare the corresponding dichloroplatinum(ll) complexes according to literature procedures. Ethyl DL-2,3-diaminopropionate dihydrochloride, ethyl L-2,4-diaminobutanoate dihydrochloride and diethyl meso-4,5-diaminosuberate dihydrochloride were synthesized according to literature procedures and used as ligands for the preparation of the corresponding diiodoplatinum(ll) complexes.
Synthesis of the platinum complexes. Reaction of the poφhyrin carboxylic acids with cisplatin did not result in the desired complexes. Therefore, cisplatin had to be activated by conversion into diammine(diaqua)platinum(ll) hydroxide, which was reacted with an equimolar amount of the porphyrin ligand in a mixture of ethanol and water or, in the case of the water-soluble ligands, in pure water. The resulting diammine(dicarboxylato)platinum(ll) complexes precipitated. To the reaction mixtures of the water-soluble complexes CH2CI2 was added to remove neutral impurities before the aqueous phase was evaporated to obtain the products.
The diamine(dichloro)platinum(ll) precursors were activated by conversion into diamine(dihydroxy)platinum(ll) species, which were reacted with an equimolar amount of the respective porphyrin carboxylic acid in a mixture of ethanol and water or, in the case of the water-soluble ligands, in pure water. The complexes precipitated. To the water-soluble complex CH2CI2 was added to remove neutral impurities and the aqueous phase was evaporated to obtain the product.
For the reaction with the porphyrincarboxylic acids it is necessary to activate the diamine(diiodo)platinum(ll) complexes by conversion into diamine(dinitrato) platinum(ll) species, which are water-soluble. In this form they were reacted with an equimolar amount of the porphyrin ligand in a mixture of ethanol and water or, in the case of the water-soluble ligand, in pure water. The water-insoluble complexes precipitated after concentrating the solution. The water-soluble complexes were isolated by chromatography on silica. Exemplary embodiments
The following examples are intended to explain the invention in more detail. The inventive compounds are tetraarylporphyrin platinum derivatives, covered by way of example by examples 1 and 2, and hematoporphyrin platinum derivatives, covered by way of example by examples 3, 4 and 5.
Examples: Example 1
Figure imgf000011_0001
n: 2
Diammine[2-(4-{10,15,20-tris[4-(1 ,4,7-trioxaoctyl)phenyl]porphyrin-5- yl}phenoxy)malonato]platinum(ll) (No. 21 in Figure 1) The compound 2-(4-{10,15,20-Tris[4-(1 ,4,7-trioxaoctyl)phenyl]porphyrin-5- yl}phenoxy)malonic acid (109 mg, 0.100 mmol) was dissolved in 10 ml of CHCI3 and
20 ml of EtOH, combined with 0.100 mmol of the aqueous diammine(diaqua)platinum(ll) hydroxide solution and stirred for 20 h. Yield: 81.0 mg (54.2 μmol, 54 %) purple powder, mp 213-214°C. Anal. (C62H66N6014Pt • 10 H20, 1494,5) C: calcd. 49,83; found. 49,19. H, N: calcd.
5,62; found 6.09. Example 2
Figure imgf000012_0001
n: 3
(+)-frans-1 ,2-Diaminocyclohexane[2-(4-{10,15,20-tris[4-(1 , 4,7,10- tetraoxaundecyl)phenyl]porphyrin-5-yl}phenoxy)malonato]platinum(ll) (No. 29 in Figure 1). 122 mg (0.100 mmol) Of the compound 2-(4-{10,15,20-Tris[4-(1 , 4,7,10- tetraoxaundecyl)phenyl]porphyrin-5-yl}phenoxy)malonic acid in 10 ml of CH2CI2 and
20 ml of EtOH were reacted with 0.100 mmol of activated (+)-frans-1 ,2- diaminocyclohexane(dichloro)platinum(ll). Yield: 113 mg (73.9 μmol, 74 %) purple solid, mp 208°C. Anal. (C74H86N6O17Pt, 1526.6) C, H, N.
Example 3
Figure imgf000013_0001
Diammine{7)12-bis[1-(1 ,4,7-trioxaoctyl)ethyl]-3,8,13I17-tetramethylporphyrin-2,18- dipropionato}platinum(ll) (No. 21 in Figure 2).
The compound 7,12-Bis[1-(1 ^J-trioxaocty ethylj-S.δ.lS.^-tetramethylporphyrin- 2,18-dipropionic acid (80.3 mg, 0.100 mmol) was dissolved in 6 ml EtOH, combined with 0.100 mmol of the aqueous diammine(diaqua)platinum(ll) hydroxide solution and stirred for 20 h. Yield: 23.0 mg (22.3 μmol, 22 %) dark brown powder, mp >
250°C. Anal. (C44H62N6O10Pt, 1030.1). C: calcd. 51.30; found. 50.75. H: calcd. 6.07; found. 5.49. N
Example 4
Figure imgf000014_0001
(+)-frans-1 ,2-Diaminocyclohexane{7,12-bis[1-(1 ,4,7,10,13I16- hexaoxaheptadecyl)ethyl]-3,8,13,17-tetramethylporphyrin-2,18- dipropionato}platinum(ll) (No. 38 in Figure 2).
The compound 7,12-Bis[1 ,4,7,10,13,16-hexaoxaheptadecyl)ethyl]-3,8,13,17- tetramethylporphyrin-2,18-dipropionic acid (107 mg, 0.100 mmol) in 10 ml of EtOH were reated with 0.100 mmol of activated (+)-frans-1 ,2-
Diaminocyclohexane(dichloro)platinum(ll).
Yield: 25.5 mg (17.2 μmol, 17 %) reddish brown powder; mp 245°C. Anal.
(C62H94N6O16Pt • 6 H20, 1482,6). C: calcd. 50.23; found. 49.02. H: calcd. 7.21 ; found. 6.33. N: calcd. 5,67; found. 6.41. Example 5
Figure imgf000015_0001
2,2'-Bipyridyl{7,12-bis[1-(1 ,4,7-trioxaoctyl)ethyl]-3,8)13,17-tetramethylporphyrin-2I18- dipropionato}platinum(ll) (No. 40a in Figure 2). 42.2 mg (0.100 mmol) of the compound 2,2'-Bipyridyl(dichloro)platinum(ll) were suspended in 15 ml of H2O. After 10 min ultrasonic treatment 34.0 mg (0.200 mmol)
of AgN03 were added and the mixture was stirred for 4 h in the dark at room temperature. The precipitated AgCI was filtered off and washed with water. The filtrate containing the activated platinum(ll) complex was evaporated. The residue was dissolved in 5 ml of H20 and combined with a solution of 80.3 mg (0.100 mmol)
7,12-Bis[1-(1 ,4,7-trioxaoctyl)ethyl]-3,8,13,17-tetramethylporphyrin-2,18-dipropionic acid in 10 ml of EtOH. After stirring for 20 h at 50°C and cooling to room temperature the precipitated solid was filtered, washed with water and EtOH and dried in vacuo. Yield: 64.0 mg (55.5 μmol, 55 %) dark purple powder, mp > 250°C. Anal. (C^NβO^Pt, 1152.2) 0, H, N. Biological data.
Data of the cytotoxic effect was obtained, for instance, on the human tumor cell lines TCC-SUP and J82. The effect of the compounds was investigated in the dark and under irradiation with light at a wavelength of 600 - 730 nm. Selected compounds are clearly more activ cytotoxically under irradiation. There is a synergism between the cytotoxic effect of the platinum component and the photodynamic principle.
Cell lines and general procedures. To determine the antiproliferative activity of the new porphyrin ligands and the corresponding platinum complexes with different amine non-leaving groups two bladder cancer cell lines TCC-SUP and J82 were selected as in vitro models.
To discriminate between the cytotoxic and phototoxic effects all experiments were carried out in duplicate. The cells were seeded into microplates and the test compounds were added after 48 h. One batch of the microplates was kept in the dark until the end of the experiment, whereas the other microplates were irradiated
48 h after addition of the substances for 10 min with a light dose of 24 J-cm , before the plates were reincubated in the dark.
End-point chemosensitivitv assay.
Hematoporphyrin platinum derivatives type.
At a dosage of 1 μM, both the dark- and phototoxicity of the porphyrin-platinum conjugates are influenced by the type of the non-leaving group. The platinum complexes with 2,2'-bipyridyl (40, 41), ethyl DL-2,3-diaminopropionate (42-46), ethyl DL-2,3-diaminobutanoate (47-51), diethyl meso-4,5-diaminosuberate (52-55) ligands were inactive at a concentration of 1 μM, both in the dark and after irradiation. The compounds bearing 1 ,2-diaminoethane (27-30) and 1 ,2-diaminopropane (31-34) non-leaving groups were also inactive against TCC-SUP cells. The most interesting porphyrin-platinum conjugates were those with the diammine (21-26) and the (+)- trans- ,2-diaminocyclohexane (35-39) ligands. Within these series of compounds the water-soluble complexes 26 and 39 were most active with T/C∞rr of around 30 %
and 15 %, respectively. At 1 μM concentration the reference cisplatin had a T/C∞rr value of approximately 2 %. At this dosage there was no statistically significant enhancement of the cytotoxicity by irradiation of the bladder cancer cells.
An increase in the concentration of complexes 40-55 to 5 μM resulted in no or only marginal augmentation of the dark toxicity (Figure 2). For most of these complexes the phototoxicity is not much higher than the cytotoxicity observed without irradiation. However, for 42, 45, 47, 49, 50 and 53 there is a distinct effect and for 40 and 44 a very strong effect on the proliferativation of the TCC-SUP cells upon irradiation is observed (Figure 2). The highest synergism was found for compound 52 resulting in the lysis of the tumor cells.
Apart from cisplatin, the highest antitumor activities were measured within the series of porphyrin-platinum conjugates bearing diammine (21-26) and (+)-frans-1 ,2- diaminocyclohexane (35-39) non-leaving groups. The differences between dark and light -induced toxicities were best for the water-soluble porphyrin-platinum complexes 26 and 39 with a side chain length of n = 17 in position 7 and 12 of the porphyrin leaving group. All the ethylenediamine and propylenediamine complexes 27-34 showed a remarkable light-induced toxicity (Figure 2).
Tetraarylporphyrin platinum derivatives type.
At a dosage of 1 μM and 5 μM, both the dark- and phototoxicity of the tetraarylporphyrin-platinum conjugates 21-38 were highly influenced by the type of the non-leaving group the results agreeing with those of the hematoporphyrin- platinum complexes discussed above. 23, 29 and 30 were the most active tetraarylporphyrin-platinum conjugates with T/C∞rr values of around 37 %, 57 % and 63 %, respectively, at 1 μM concentration. This is analogous to the hematoporphyrin- platinum complexes, the most active of which were those with the diammine or the (+)-tøat7S-1 ,2-diaminocyclohexane non-leaving groups. At 1 μM concentration there was only a slight enhancement of the cytotoxicity of the tetraarylpoφhyrin-platinum conjugates with the side chain length n = 2 and n = 3 upon irradiation. On the average the light-induced T/C∞rτ values were by approximately 20 % lower than the dark-only cytotoxicities (data not shown).
An increase in the concentration of the complexes to 5 μM enhanced the dark effects and the phototoxicities as shown in Figure 1. Apart from cisplatin, the highest antitumor activities were measured for the tetraarylporphyrin-platinum conjugates bearing diammine (21-23) and (±)-trans-1 ,2-diaminocyclohexane (28-30) non-leaving groups. The differences between dark and light-induced toxicities were best for the tetraarylporphyrin-platinum complexes 24, 27, 32-34, 36 and 38 with a side chain length of n = 2 or n = 3 (Figure 1 ).

Claims

Patent claims
1. Porphyrin platinum derivatives of the (a) tetraarylporphyrin platinum derivatives type according to formula I
Figure imgf000019_0001
n: 1-20
X: O, S, NH, N-Alkyl
R2/R3: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
R4: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
R5: H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst. Amino
R6: H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst. Amino
and according to formula II
Figure imgf000020_0001
n: 1-20
X: O, S, NH, N-Alkyl
R1/R2/R3/R4: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
Oder R2-Z-R3, mit Z: (CH2)n, n=0-6
R1/R4: H, -(CH2)n-COOR8, n=0-6
R5: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
R6: H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst. Amino
R7: H, Alkyl, O-Alkyl, S-Alkyl, Halogen, Nitro, Cyano, Amino, subst. Amino
R8: H, Alkyl
and
(b) hematoporphyrin platinum derivatives type
according to formula III
Figure imgf000021_0001
R2/R3: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
R4: H, Alkyl, Cycloalkyl
R5: H, Alkyl, Cycloalkyl
R6: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
and according to formula IV
Figure imgf000022_0001
R1/R2/R3/R4: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl oder R2-Z-R4, mit Z: (CH2)n, n=0-6 Oder R1/R3: H, -(CH2)n-COOR6, n=0-6
R4/R5: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl R6: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl R7: H, Alkyl, Aryl, Aralkyl, Hetaryl, Hetarylalkyl, Cycloalkyl
2. Compounds as shown in formulas I, II, III and IV according to claim 1 , having one or more asymmetric carbon atoms, in the form of their racemates, their pure enantiomers and/or their diastereoisomers or in the form of mixtures of these enantiomers or diastereoisomers.
3. Use of the compounds as shown in formulas I, II, III and IV according to claims 1 and 2 as therapeutic active compounds for the production of medicaments for the treatment of cancer.
4. Use of the compounds as shown in formulas I, II, III and IV according to claims 1 and 2 as therapeutic active compounds for the production of medicaments for the photodynamic treatment of tumors.
5. Particularly preferred use of the compounds as shown in formulas I, II, III and IV according to claims 1 , 2 and 4 as therapeutic active compounds for the production of medicaments for the photodynamic treatment of tumors by irradiating with electromagnetic radiation having a wavelenght of 600 to 730 nm.
6. Medicaments comprising one or more compounds according to claims 1 and 2 in addition to customary physiologically tolerable carriers and/or diluents or auxiliaries.
7. Process for the production of a medicament according to claim 6, characterized in that one or more compounds according to claims 1 and 2 are processed to give pharmaceutical preparations or brought into a therapeutically administrable form using a customary pharmaceutical carriers and/or diluents or other auxiliaries.
8. Use of compounds of the formulas I, II, III or IV according to claims 1 and 2 and/or pharmaceutical preparations acording to claims 6 and 7 on their own or in combination with one another or in combination with carriers and/or diluents or other auxiliaries.
PCT/EP2003/000874 2002-02-01 2003-01-29 Water-soluble porphyrin platinum compounds with high tumor selectivity and their use for the treatment of benign and malignant tumor diseases WO2003064424A2 (en)

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JP2003564047A JP2005522429A (en) 2002-02-01 2003-01-29 Novel water-soluble porphyrin platinum compounds with high tumor selectivity and their use to treat benign and malignant tumor diseases
NZ534541A NZ534541A (en) 2002-02-01 2003-01-29 Water-soluble porphyrin platinum compounds with high tumor selectivity and their use for the treatment of benign and malignant tumor diseases
EP03734604A EP1470139A2 (en) 2002-02-01 2003-01-29 NOVEL, WATER−SOLUBLE PORPHYRIN PLATINUM COMPOUNDS WITH HIGH TUMOR SELECTIVITY AND THEIR USE FOR THE TREATMENT OF BENIGN AND MALIGNANT TUMOR DISEASES
UA20040806544A UA78007C2 (en) 2002-02-01 2003-01-29 Water-soluble porphyrin platinuim compounds with high tumor selectivity and their use for the treatment of benign and malignant tumor diseases
BR0307400-5A BR0307400A (en) 2002-02-01 2003-01-29 Water soluble porphyrin platinum compounds with high tumor selectivity and their use for the treatment of benign and malignant tumor diseases
HRP20040788 HRP20040788A2 (en) 2002-02-01 2004-08-30 Novel, water-soluble porphyrin platinum compounds with high tumor selectivity and their use for the treatment of benign and malignant tumor diseases
NO20043650A NO20043650L (en) 2002-02-01 2004-08-31 New, water-soluble porphyrin-platinum compounds with high tumor selectivity and their use in the treatment of benign and malignant tumor diseases
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US6995260B2 (en) * 2004-05-20 2006-02-07 Brookhaven Science Associates, Llc Carboranylporphyrins and uses thereof
US8287839B2 (en) * 2006-12-04 2012-10-16 Brookhaven Science Associates, Llc Carboranylporphyrins and uses thereof
EP1950217A1 (en) * 2007-01-26 2008-07-30 Université de Neuchâtel Organometallic compounds for the treatment of cancer
US8444953B2 (en) * 2007-03-22 2013-05-21 Brookhaven Science Associates, Llc Symmetric and asymmetric halogen-containing metallocarboranylporphyrins and uses thereof
US20080279781A1 (en) * 2007-05-10 2008-11-13 Brookhaven Science Associates, Llc Glycosylated Carboranylporphyrins and Uses Thereof
JP5823413B2 (en) * 2010-01-22 2015-11-25 カウンシル オブ サイエンティフィック アンド インダストリアル リサーチ Process for the preparation of novel porphyrin derivatives and their use as PDT agents and fluorescent probes
CN102408452B (en) * 2011-12-13 2014-09-03 中山大学 Tetrapyridylporphine bridged crossed tetra-palladium complexes, and preparation method and antitumor activity thereof
BR112015023568B8 (en) 2013-03-15 2024-01-02 Ann Mcfarland Sherri Metal-based compound as photodynamic compound
US11661433B2 (en) 2018-05-09 2023-05-30 Virginia Commonwealth University Near-IR activatable fluorescent small molecules with dual modes of cytotoxicity
CN109251206B (en) * 2018-09-21 2022-02-08 上海大学 Water-soluble platinum-porphyrin complex and preparation method thereof
CN110028513B (en) * 2019-04-30 2020-12-11 北京太阳升高科医药研究股份有限公司 Porphyrin derivatives and sonosensitizers
CN110590852B (en) * 2019-08-29 2022-12-09 合肥学院 Platinum complex with antitumor activity and preparation method thereof

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0193083A1 (en) * 1985-02-23 1986-09-03 ASTA Pharma AG Anti-tumoral(1-arylmethyl-ethylene diamine)-platinum(II)-complexes
WO1999043317A1 (en) * 1998-02-25 1999-09-02 Schering Aktiengesellschaft Necrosis-affine compounds and the utilization thereof in order to produce preparations for pharmacotherapy

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH0615545B2 (en) * 1984-10-01 1994-03-02 東洋薄荷工業株式会社 Metal pheophobide derivatives and metal porphyrin derivatives
ATE84036T1 (en) * 1985-10-23 1993-01-15 Nihon Mediphysics Co Ltd PORPHYRIN DERIVATIVES, THEIR PRODUCTION AND USE.

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0193083A1 (en) * 1985-02-23 1986-09-03 ASTA Pharma AG Anti-tumoral(1-arylmethyl-ethylene diamine)-platinum(II)-complexes
WO1999043317A1 (en) * 1998-02-25 1999-09-02 Schering Aktiengesellschaft Necrosis-affine compounds and the utilization thereof in order to produce preparations for pharmacotherapy

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
BRUNNER ET AL: "Synthesis and antitumor activity of new porphyrin-platinum(II) complexes with the cytostatic platinum fragment attached to the porphyrin side chains" CHEMISCHE BERICHTE, VERLAG CHEMIE GMBH. WEINHEIM, DE, vol. 127, no. 11, 1994, pages 2141-2149, XP002108805 ISSN: 0009-2940 *
BRUNNER, H. ET AL: "Synthesis and in vitro testing of hematoporphyrin type ligands in platinum(II) complexes as potent cytostatic and phototoxic antitumor agents" INORGANICA CHIMICA ACTA (1997), 264(1-2), 67-79 , XP002245477 cited in the application *
BRUNNER, HENRI ET AL: "Platinum(II) complexes with porphyrin ligands - additive cytotoxic and photodynamic effect" ANGEWANDTE CHEMIE (1994), 106(21), 2305-6 (SEE ALSO ANGEW. CHEM., INT. ED. ENGL., 1994, 33(21), 2214-15) , XP002245479 *
BRUNNER, HENRI ET AL: "Platinum(II) complexes with porphyrin ligands: synthesis and synergisms in the photodynamic tumor therapy" CHEMISCHE BERICHTE (1995), 128(2), 173-81 , XP002245478 cited in the application *

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