Title of Invention

"RUTHENIUM ANTICANCER COMPLEXES"

Abstract Ruthenium (II) compounds of formula (I) are useful in the treatment and/or prevention of cancer, wherein formula (I) is:
Full Text RUTHENIUM ANTICANCER COMPLEXES
This invention relates to ruthenium(II) compounds, to their use in medicine, particularly for the treatment and/or prevention of cancer, and to a process for their preparation.
Certain ruthenium(II) complexes have been proposed for use in treating cancer. For example, US 4980473 discloses 1,10-phenanthroline complexes of ruthenium(II) and cobalt(III) which are said to be useful for the treatment of tumour cells in a subject.
Some other ruthenium(II) and ruthenium(III) complexes which have been shown to exhibit antitumour activity are mentioned in Guo et al, Inorganica Chimica Acta, 273 (1998), 1-7, specifically trans-[RuCl2(DMSO)4], trans-[RuCl4(imidazole)2]" and trans-[RuCl4(mdazole)2], Clarke et al have reviewed the anticancer, and in particular the antimetastatic, activity of ruthenium complexes: Chem. Rev., 1999, 99, 2511-2533. Also, Sava has reviewed the antimetastatic activity in "Metal Compounds in Cancer Therapy" Ed by S P Flicker, Chapman and Hall, London 1994, p. 65-91.
Dale et al, Anti-Cancer Drug Design, (1992), 7, 3-14, describes a metronidazole complex of ruthenium(II) ie, [(η6-C6H6)RuCl2(metronidazole)] and its effect on DNA and on E. coli growth rates. Metronidazole sensitises hypoxic tumour cells to radiation and appears to be an essential element of the complexes of Dale et al. There is no indication in Dale et al that the complexes would be at all effective in the absence of the metronidazole ligand.
Kramer et al, Chem Eur J., 1996, 2, No. 12, p. 1518-1526 discloses half sandwich complexes of ruthenium with amino esters.
Bennett et al, Canadian Journal of Chemistry, (2001), 79, 655-669 discloses certain ruthcnium(II) complexes with acetylacetonate ligands.
Oro el al, J Chem Soc, Dalton Trans, (1990), 1463 describes ruthenium(II) complexes containing η6-p-cymene and acetylacetonate ligands.
WO 01/30790 discloses ruthenium(II) compounds and their use as anticancer agents. The compounds have neutral N-donor ligands and the resulting ruthenium complex is generally positively charged.
WO 02/02572 also discloses ruthenium(II) compounds that have activity against cancer cell lines. Again, the complexes are generally positively-charged. Complexes are disclosed containing a bidentate ligand which is a neutral diamine ligand.
Chen et al, J. Am. Chem. Soc, volume 124, no 12, 3064, (2002), describes the mechanism of binding of ruthenium complexes to guanine bases. The binding mode] requires NH bonds from a diamino ligand to be present in the complex for hydrogen bonding to the guanine base. Similarly, Morris et al, J. Med. Chem., volume 44, 3616-3621, (2001), describes the selectivity of ruthenium(II) complexes for binding to guanine bases.
There exists a need for novel anti-cancer compounds which can be used as alternatives to the compounds which are currently available.
Furthermore, there exists a need for compounds which are capable of binding to different DNA bases. Binding to different DNA bases can provide a compound that exhibits increased activity to drug-resistant tumour cells.
The present invention provides a novel class of ruthenium(II) complexes having anti-tumour activity.
According to the present invention, there is provided a ruthenium(II) compound of formula (I):
(Formula Removed)
wherein: R1, R2, R3, R4, R5 and R6 independently represent H, (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, halo, CO2R7, CONR8R9, COR10, SO3G, SO2N R11R12, aryloxy, (C1-C6)alkoxy, (C1-C6)alkylthio, -N=N-R13, NR14R15, aryl or aralkyl, which latter two groups are optionally substituted on the aromatic ring by one or more groups independently selected from (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, aryl, aralkyl, halo, CO2R7a, CONR8aR9a, COR,10a, SO3G', SO2NR11aR12a, aryloxy, (C1-C6)alkoxy, (C1-C6)alkylthio, -N=N-R,13a, NR,14aR15a , or R1 and R2 together with the ring to which they are bound represent a saturated or unsaturated carbocyclic or heterocyclic group containing up to three 3- to 8- membered carbocyclic or heterocyclic rings, wherein each carbocyclic or heterocyclic ring may be fused to one or more other carbocyclic or heterocyclic rings, and wherein each of the rings may be optionally substituted by one or more groups independently selected from (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-
C6)alkyl, amino(C1-C6)alkyl, aryl, aralkyl, halo, COR7b, CONR8bRyb, COR10b,
SO3H, SO2NRllbR12b, aryloxy, (C1-C6)alkylthio, -N=N-R13b, NR14bR,15b or (C1-
C6)alkoxy;
R7, R8, R9, R10, R11, R12, R13, R14, R15, R7a, R8a, R9a, R,10a, Rlla, R,12a, R,13a, R,14a,
R15a, R7b, R8b, R9b, R,10b, R11b, R,12b, R,13b, R14b and R,15bare independently
selected from H, (C1-C6)alkyl, aryl or aralkyl;
X is a neutral or negatively charged 0-, N- or S- donor ligand or halo;
G and G' are independently selected from alkali metals (eg, sodium or
potassium), aryl, aralkyl and (C1-C6) alkyl;
Y-L-Y' is a bidentate ligand bearing an overall negative charge with a
proportion of the charge on both Y and Y, Y and Y' are independently
selected from O, S or NR16, wherein R16 is H, (C1-C6)alkyl, aryl or aralkyl, and
L is a group linking Y and Y' and comprises one or more groups selected from
(C)-C6)alkylene, (C1-C6)alkenylene, (C1-C6)alkynylene, arylene, aralkylene,
alkarylene, each of said latter six groups being optionally substituted,
ferrocenylene, Se, Se-Se, S-S, N=N and C=O;
m is -1, 0 or +1 and the compound comprises a counterion when m is -1 or +1;
the compound of formula (I) optionally being in the form of a dimer in which
two L groups are linked either directly or through a group comprising one or
more of (C1-C6)alkylene, (C1-C6)alkenylene, arylene, aralkylene, alkarylene,
Se, Se-Se, S-S, N=N and C=O or in which L bears two Y groups and two Y'
groups;
with the proviso that:
when Y-L-Y' is (CH3C(O)CHC(O)CH3)-, X is halo or an N-donor ligand, R1,
R2, R3, R4, R5 and R6 together with the ring to which they are bound do not
represent 4-isopropyl-l-methylbenzene;
when Y-L-Y' is (CH3C(O)CHC(O)CH3)- and X is chloro, (CH3)2SO, CH3CN,
pyridine or (CH3C(O)CHC(O)CH3)-: R1, R2, R3, R4, R5 and R6 are not all H or
all methyl; R1, R3 and R5 are not all H when R2, R4 and R6 are all methyl; and
R2, R4 and R6 are not all H when R1, R3 and R5 are all methyl; and
when Y-L-Y' is (CF3C(O)CHC(O)CF3)- and X is chloro, R1, R2, R3, R4, R5 and R6 are not all H or all methyl; R1, R3 and R5 are not all H when R2, R4 and R6 are all methyl; and R2, R4 and R6 are not all H when R1, R3 and R5 are all
methyl.
The compounds of the invention may be in the form of pharmaceutically acceptable salts, solvates and/or prodrugs. Prodrugs are variants of the compounds of the invention which can be converted to compounds of formula (I) in vivo.
The compounds of formula (I) may have one or more chiral centres. When the compounds of formula (I) have one or more chiral centres, they may be in the form of one enantiomer, may be enriched in one enantiomer or may be a racemic mixture.
The term "alkyl" as used herein includes C1 to C6 alkyl groups which may be branched or unbranched and may be open chain or, when they are C3 to C6 groups, cyclic. Unbranched open chain alkyl groups include, for example, methyl, ethyl, propyl, butyl, pentyl and hexyl. Branched open chain alkyl groups include, for example, 2-propyl, 2-butyl and 2-(2-methyl)propyl. Cyclic groups include cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl. The alkyl groups in the compounds of the invention may optionally be substituted. Substiruents include one or more further unsubstituted alkyl groups and/or one or more further substiruents, such, as, for example, cyano, nitro, -CO2(C1-C6)alkyl, halo, thiol (SH), thioether (eg, S-(C1-C6)alkyl) and sulfonate. The term "alkoxy" means -O-alkyl. The term "alkylthio" means -S-alkyl.
The terms "hydroxy(C1-C6)alkyl" and "amino(C1-C6)allcyl" refer to alkyl groups, as defined above, substituted with one or more hydroxyl (OH) or amino (NH2) groups, respectively.
The terms "alkenyl" and "alkynyl" are defined similarly to the term "alkyl" but refer to groups that contain from 2 to 6 carbon atoms and include one or more carbon-carbon double bonds or one or more carbon-carbon triple bonds, respectively. Alkenyl and alkynyl groups may be optionally substituted in the same way as alkyl groups. Examples of alkenyl groups are ethenyl, 1-propenyl, 2-propenyl, 1-butenyl, 2-butenyl, 1,4- butadienyl, cyclohexenyl and cyclohexadienyl.
The term "alkylene" is defined similarly to the definition of the term "alkyl" but includes C2 to C6 groups and represents a divalent species with radicals separated by two or more (eg, from two to six) carbon atoms linked in a chain. Preferably, the alkylene groups are straight chain groups. Alkylene groups are optionally substituted in the alkylene chain, preferably with one or more phenylene (eg, 1-4-phenylene) and/or -CONR1x- groups and/or -NR2x- groups, where Rlx and R2x independently represent H, alkyl, aryl or aralkyl. Preferably, Rlx and R2x are H or C1 to C3 alkyl. The terms "alkenylene" and "alkynylene" are defined similarly and refer to divalent radicals containing one or more carbon-carbon double bonds or one or more carbon-carbon triple bonds, respectively.
The term "aryl" as used herein includes aromatic carbocyclic rings such as phenyl, naphthyl and anthracenyl and heterocyclic rings such as pyridyl, imidazolyl, pyrrolyl and furanyl. Aryl groups may optionally be substituted with one or more substituents including, for example, (C1-C6)alkyl, cyano, nitro, hydroxyl, halo(C1-C6)alkyl, -CO2(C1-C6)alkyl, halo, thiol (SH), thioether (eg, S-(C1-C6)alkyl) and sulfonate (SO3H). The term "aryloxy" means -O-aryl.
The term "heterocyclic ring" refers to a 3-, 4-, 5-, 6-, -7, or 8- (preferably 5-, 6-or 7-) membered saturated or unsaturated ring, which may be aromatic or non-aromatic, containing from one to three heteroatoms independently selected from N.,O and S, eg, indole.
The term "arylene" refers to a divalent radical comprising an aromatic carbocyclic or heterocyclic ring in which the radicals are present at different positions on the ring. An example of an arylene group is 1,4-phenylene.
The term "aralkyl" means alkyl substituted with aryl eg, benzyl. The term "alkaryl" means aryl substituted with alkyl eg, methylpheny].
The term "aralkylene" refers to a divalent radical that can be derived from an aralkyl group eg, l-methylene-4-phenyl. Each of the two radicals may be present on the aryl ring or on the alkyl group or one of the radicals may be present on the alkyl group and the other radical present on the aryl ring. The term "alkarylene" is defined similarly.
The term ferrocenylene refers to a diradical derived from ferrocene (FeCp2). Each radical may be present on the same ring or on different rings.
The term "halo" means a halogen radical selected from fluoro, chloro, bromo and iodo. Chloro is particularly preferred. When X is halo in formula (I), it will be appreciated that X may be thought of as having at least some of the character of a negatively charged ion rather than being covalently bonded to the ruthenium atom. Indeed, all ligands X may have some ionic as well as some covalent character.
The term "haloalkyl" means alkyl substituted with one or more halo groups eg, trifluoromethyl.
In the compounds of the invention, R1 and R2 together with the ring to which they are bound in compounds of formula (I) may represent an ortho- or peri-fused carbocyclic or heterocyclic ring system. R1 and R2 together with the ring to which they are bound may, for example, represent a wholly carbocyclic fused ring system such as a ring system containing 2 or 3 fused carbocyclic rings eg, optionally substituted, optionally hydrogenated naphthalene or anthracene. Thus, R1 and R2 together with the ring to which they are bound in compounds of formula (I) may represent a fused tricyclic ring such as anthracene or a mono, di, tri, tetra or higher hydrogenated derivative of anthracene. For example, R1 and R2 together with the ring to which they are bound in formula (I) may represent anthracene, 1, 4-dihydroanthracene or 1, 4, 9, 10-tetrahydroanthracene. Examples of R1 and R2 together with the ring to which they are bound in compounds of formula (I) representing heterocyclic ring systems include those compounds in which R1 and R2 together with the ring to which they are bound represent 2,3-benzofuran, indole or benzo[b]thiophene.
Preferably, R1, R2, R3, R4, R5 and R6 are independently selected from H, (C1-C6)alkyl and phenyl or R1 and R2 together with the ring to which they are bound represent anthracene or a hydrogenated derivative of anthracene, said phenyl and anthracene or a hydrogenated derivative of anthracene group being optionally substituted by one or more groups independently selected from (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, phenyl, benzyl, halo, carboxyl, CO2(C1-C6)alkyl, CONH2, COH, CO(C1-C6)alkyl, SO3H, SO2NH2, phenoxy, (C1-C6)alkylthio, NH2 or (C1-C6)alkoxy. Most preferably, one of R1, R2, R3, R4, R5 and R6 is phenyl and the other groups are all H, or one or two of R1, R2, R3, R4, R5 and R6 is or are (C1-
C6)alkyl and the other groups are H, or R1 and R2 together with the ring to which they are bound represent anthracene or a hydrogenated derivative of anthracene.
It can be desirable for the compounds of the invention to be uncharged since this may assist the transport of the compounds in in vivo systems and thus their anticancer cytotoxic activity. Therefore, in formula (I), it is preferred that m is 0. When m is +1 or -1, the compounds of formula (I) comprise a counterion. Suitable counterions include non-nucleophilic ions such as, for example, PF6 and BF4-.
In compounds of formula (I), X is a neutral or negatively charged O-, N- or S-donor ligand or halo. Suitable ligands include, for example, H2O, di((C1-C6)alkyl)S(O), (C1-C6)alkylCO2- or di((C1-C6)alkyl)C=O. Other ligands include, for example, N-donor nitrile ligands (eg, compounds of formula (C1-C6)alkylCN) and N-donor pyridine ligands, optionally substituted at one or more of the carbon rings of the pyridine ring eg, by (C1-C6)alkyl or halo. Other suitable ligands are (C1-C6)alkyl primary amines such as methylamine and ethylamine. Preferably, X is halo or CH3CN, most preferably, X is chloro.
Y-L-Y' is a bidentate ligand. The ligand has an overall negative charge ie, the ligand would bear a negative charge when not present in the complexes of the invention. The ligand may bear a single negative charge or may have more than one negative charge eg, by being a dianion. Preferably, Y-L-Y' has a single or double negative charge. The charge may be distributed throughout the ligand but a proportion of the charge is present on both Y and Y', The charge may be due to the formation of an anion by deprotonation at Y and/or Y' or by delocalisation of the charge from Y to Y' or vice versa. Preferably, there is delocalisation of the charge from Y to Y' or from Y' to Y by conjugation of electrons through L. L may comprise one or more groups, for
example L can comprise an alkylene group having a CO group at each end of the alkylene chain. Preferably, Y-L-Y' is selected from ligands of formulae
(Formula Removed)
wherein T and T' are independently selected from 0 and S, Rlg and R3g are independently H, (C1-C6)alkyl, aryl or aralkyl,
R1c to R5f (i.e., R1c, R2c, R3C, R4C, R5c, R6c R7c R8c, R9c, R1d, R2d, R3d, R4d, R5d, R6d, R7d, R8d, R9d, Rl0d, R1e, R2e, R3e; R4e, R5e, R6e, R7e, R8e, R9e, Rlf, R2f, R3f, R4f
and R5f) and R2g are independently H, (C1-C6alkyl, aryl, aralkyl, wherein the latter two groups are optionally substituted by one or more groups independently selected from (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, aryl, aralkyl, halo, carboxyl, CO2R , CONR8bR9b, COR10b, SO3H, SO2N R11bR12b, aryloxy, (C1-C6)alkylthio, -N=N-R13b, NR,4bR15b and (C1-C6)alkoxy, wherein R713, R8b, R9b, R10b, R,11b, R12b, R13b, R14b and R15bare independently selected from H, (C1-C6)alkyl, aryl or aralkyl.
The compounds of formula (I) may be in the form of dimers, which may also be termed dinuclear complexes — such complexes contain two ruthenium atoms. Dinuclear complexes can be provided by employing a ligand which comprises two linked ligands Y-L-Y' so as to bridge between two ruthenium centres. Therefore, in another embodiment of the invention, Y-L-Y' may be selected from ligands of formulae (XI) to (XV):
(Formula Removed)
wherein T, T', T" and T'" are independently selected from O and S,
A comprises one or more groups selected from (C1-C6)alkylene, (C1-C6)alkenylene, (C1-C6)alkynylene, arylene, aralkylene, alkarylene, ferrocenylene, Se, Se-Se, S-S, N^N and C^O,
and R1h to R6j (i.e., R1h, R2h, R3h, R4h, R1i, R2i, R3i, R4i, R5i, R6i, R7i, R8i, R9i, R10i R11i, R12i, R13i, R14i, R1j, R2j, R3j. R4j, R5j, and R6j) are independently H, (C1-C6)alkyl, aryl, aralkyl, wherein the latter two groups are optionally substituted by one or more groups independently selected from (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, aryl, aralkyl, halo, carboxyl, CO2R7b, CONR8bR9b, COR10b, SO3H, SO2N R11bR,12b, aryloxy, (C1-C6)alkylthio, -N=N-R13b, NR,4bR15b and (C1-C6)alkoxy, wherein R7b, R8b, R9b, R10b, Rllb, R12b, R,3b, R,4b and R15bare independently selected from H, (C1-C6alkyl, aryl or aralkyl.
Particularly preferred compounds of formula (I) are those in which Y-L-Y' is:
(Formula Removed)
wherein T and T are independently O and S, and
R, R1c and R3c are independently H, (C1-C6)alkyl, aryl, aralkyl, wherein the latter two groups are optionally substituted by one or more groups independently selected from (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, aryl, aralkyl, halo, carboxyl, CO2R7b, CONR8bR9b, COR10b, SO3H, SO2N R,1lbR12b, aryloxy, (C1-C6)alkylthio, -N=N-R,13b, NR14bR15b and (C1-C6)alkoxy, wherein R7b, R8b, R9b, R10b, R11b, R12b, R13b, R,14b and R15b are as defined above. Even more preferably, T and T' are both O and R, Rlc and R3c are independently (C1-C6)alkyl or phenyl, said phenyl optionally substituted by (C1-C6)alkyl, hydroxy(C1-C6)alkyl, amino(C1-
C6)alkyl, halo, carboxyl, CO2(C1-C6)alkyl, CONH2, COH, CO(C1-C6)alkyl, SO3H, SO2NH2, phenoxy, (C1-C6)alkylthio, NH2 or (C1-C6)alkoxy. Most preferably, R is H or (C1-C6)alkyl and R1c and R3c are independently unsubstituted (C1-C6)alkyl or optionally substituted phenyl.
In the compounds of the invention, it is preferred that Y and Y' (including T, T', T" and T'" in the above formulae) are all 0.
Compounds of formula (I) and ligands of formula Y-L-Y' may exist in one or more tautomeric forms, all of which are covered by the present invention. For example, ligands of formula:
(Formula Removed)
may have the following tautomeric forms:
(Formula Removed)
The existence and nature of the tautomers will depend on the structure of Y-L-Y',
A particularly preferred group of compounds of formula (I) are those in which: X is halo (preferably chloro);
R1, R2, R3, R4, R5 and R6 are independently selected from H, (C1-C6)alkyl and phenyl or R1 and R2 together with the ring to which they are bound represent anthracene or a hydrogenated derivative of anthracene, said phenyl and
anthracene or a hydrogenated derivative of anthracene group being optionally substituted by one or more groups independently selected from (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, phenyl, benzyl, halo, carboxyl, CO2(C1-C6)alky], CONH2, COH, CO(C1-C6)alkyl, SO3H, SO2NH2, phenoxy, (C1-C6)alkylthio, NH2 or (C1-C6)alkoxy; Y-L-Y' is:
(Formula Removed)
wherein T and T' are both O, R is H or (C1-C6)alkyl and R1c and R3c are independently (C1-C6)alkyl or phenyl, said phenyl optionally substituted by from one to three groups selected from (C1-C6)alkyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, halo, carboxyl, CO2(C1-C6)alkyl, CONH2, COH, CO(C1-C6)alkyl, SO3H, SO2NH2, phenoxy, (C1-C6)alkylthio, NH2 or (C1-C6)alkoxy.
A further preferred group of compounds of formula (I) is that in which: R1, R2, R3, R4, R5 and R6 are all H, or one of R1, R2, R3, R4, R5 and R6 is phenyl and the other groups are all H, or one or two of R1, R2, R3, R4, R5 and R6 is or are (C1-C6)alkyl and the other groups are all H, or R1 and R2 together with the ring to which they are bound represent anthracene or a hydrogenated derivative of anthracene; X is chloro; and Y-L-Y' is:
wherein T and T are both O, R is H and Rlc a
(Formula Removed)
nd R3c are independently (C1-C6)alkyl or R1c and R3c are independently unsubstituted (C1-C6)alkyl or optionally substituted phenyl.
The compounds of the invention have been found to exhibit cytotoxic activity against cancer cell lines and can therefore be expected to show anticancer activity. The compounds may be used to kill cells in vivo or ex vivo.
In another embodiment, therefore, the present invention provides a compound of formula (I) as defined above without the provisos for use in medicine. The invention also contemplates the provision of a compound of formula (I) as defined above without the provisos for use in the treatment and/or prevention of cancer and the use of a compound of formula (I) as defined above without the provisos in the treatment and/or prevention of cancer.
Also provided by the invention is the use of a compound of formula (I) as defined above without the provisos in the manufacture of a medicament for the treatment and/or prevention of cancer.
Further provided by the invention is a pharmaceutical composition comprising a compound of formula (I) according as defined above without the provisos together with one or more pharmaceutically acceptable excipients.
Yet another aspect of the invention is a method of treating and/or preventing cancer which comprises administering to a subject a therapeutically effective amount of a compound of formula (I) as defined above without the provisos or a composition of the invention.
The compounds of the invention may be used directly against a tumour. Alternatively or additionally, the compounds may be used to prevent or inhibit metastasis and/or to kill secondary tumours. It will be understood that the prevention or inhibition of metastasis is encompassed by the term "preventing cancer", as used herein.
The term tumour includes all forms of neoplastic cell growth, including tumours of the lung, liver, blood cells, skin, pancreas, stomach, colon, prostate, uterus, breast, lymph glands and bladder. Ovarian tumours may be especially suitable for treatment according to the invention.
Compounds of the invention may be effective in treating and/or preventing tumours caused by cells that are resistant to other cytotoxic drugs, such as cis-platin, for example.
Certain compounds of the invention have the surprising advantage that they do not bind selectively to guanine bases but show roughly equal affinity for binding to guanine and adenine. This effect was unexpected, given that the presence of an N-H group available for hydrogen bonding was thought to be a requirement for bonding to guanine. Furthermore, binding to guanine and adenine gives the compounds a potentially greater ability to be less susceptible to drug resistance in tumour cells. In one embodiment, the compounds can be used in a method of binding non-selectively to guanine bases, preferably a method of binding to guanine and adenine bases with roughly equal affinity. This method may be, for example, a method of killing cells in vivo or ex vivo,
or another method such as the separation of guanine and adenine bases from a mixture.
The invention also provides a pharmaceutical composition comprising one or more compounds of the invention together with one or more phannaceutically acceptable excipients. Suitable excipients include diluents and/or carriers.
The compounds of the invention may be administered by a number of routes including, for example, orally, parenterally (eg, by injection intramuscularly, intravenously or subcutaneously), topically, nasally or via slow releasing microcarriers. Thus, suitable excipients for use in the pharmaceutical compositions of the invention include saline, sterile water, creams, ointments, solutions, gels, pastes, emulsions, lotions, oils, solid carriers and aerosols.
The compositions of the invention may be formulated in unit or sub-unit dosage form including, for example, tablets, capsules and lozenges and containers containing the composition in a form suitable for parenteral administration. Preferably, the compositions are in a form that is suitable for injection.
The specific dosage level of the compounds and compositions of the invention will depend upon a number of factors, including the biological activity of the specific compound used and the age, body weight and sex of the subject. It will be appreciated that the subject may be a human or a mammalian animal.
The compounds and compositions of the invention can be administered alone or in combination with other compounds. The other compounds may have a biological activity which complements the activity of the compounds of the invention eg, by enhancing its effect in killing tumours or by reducing any side-effects associated with the compounds of the invention.
In another embodiment, the present invention provides a process for preparing the compound of formula (I) which comprises the reaction of a compound of formula [(η6-C6(R1)(R2)(R3)(R4)(R5)(R6))RuX2], optionally in the form of a dimer, with Y-L-Y', in a suitable solvent for the reaction, wherein R1R2, R3, R4, R5, R6, X, Y,Y' and L are as defined for formula (I) above. Preferably, the process comprises the reaction of a compound of formula [(η6-C6(R1)(R2)(R3)(R4)(R5)(R6))RuX2], optionally in the form of a dimer, with a salt comprising Y-L-Y' as an anion (eg, a salt of an alkali metal such as sodium) at a temperature of from 0°C to 100°C (eg, 10°C to 50 °C) in a polar solvent such as a di(C1-C6)alkyl ketone (eg, acetone) or water or mixtures thereof. The compound of formula (I) can be separated from the reaction mixture, for example by extraction into a less polar solvent (eg, dichloromethane) and, optionally, purified (eg, by recrystallisation from a suitable solvent or mixture of two or more different solvents).
Suitable compounds of formula [(Η6-C6(R1)(R2)(R3)(R4)(R5)(R6))RUX2] for use as starting materials (starting ruthenium complexes) in the process of the invention can be produced as described in WO 01/30790 and WO 02/02572. Compounds of formula Y-L-Y' are either commercially available or can be synthesised by routes well known to those skilled in the art.
The invention will now be described with reference to the following non-limiting examples.
Examples
A. Synthesis
Example 1
I[(η6 p-cymene)RuClfH3CCOCHCOCH3 - 0,0)]
The compound of Example 1 was prepared using the method described by D. Carmona et al, J. Chem. Soc, Dalton Trans., 1990, 1463 - 1476.
l(yf-p-cymme)RuCl2]2 (250 mg, 0.41 mmol) and sodium acetylacetonate monohydrate (150 mg, 1.07 mmol) were stirred in acetone (25 ml) at room temperature (RT) for 50 min. The extraction step then involved removal of the solvent in vacuo on a rotary evaporator, extraction of the product with dichloromethane, filtration using Whatman No. 541 filter paper, followed by removal of the solvent again on a rotary evaporator. The final product was recrystallised from acetone/diethyl ether and left in a freezer at 253 K overnight. The red crystals (180 mg, 0.48 mmol, 59% yield) were collected and dried in vacuo. 1H NMR (CDC13): δ 5.43 (d, 2H), 5.18 (d, 2H), 5.13 (s, 1H), 2.85 (sp, 1H), 2.24 (s, 3H), 1.97 (s, 6H), 1.30 (d, 6H).
Example 2
[(η6-p-cymenemuCl(F3CCOCHCOCF2 - 0.0)]
[(η6-p-cymene)RuCl2]2 (250 mg, 0.41 mmol) and sodium hexafluoroacetylacetonate (260 mg, 1.13 mmol) were stirred in acetone (25 ml) at RT for 40 min. After the extraction step using dichloromethane (see Example 1), the final product was dissolved in diethyl ether, the solution was concentrated on a rotary evaporator, and hexane was added. The vessel was left in a freezer at 253K overnight. The crystals (277 mg, 0.58 mmol, 70.7% yield) were collected and dried in vacuo. 1H NMR (CDC13): δ 5.86 (s, 1H), 5.63 (d, 2H), 5.36 (d, 2H), 2.88 (sp, 1H), 2.24 (s, 3H), 1.33 (d, 6H).
Example 3 [(η6p-cymene)RuCl(C6H5COCHCO6H5- 0.0)1
l,3-diphenyl-l,3-propanedione (277 mg, 1.24 mmol) and sodium methoxide (62 mg, 1.15 mmol) were stirred in methanol (30 ml) at room temperature (RT) for 100 min. The solvent was removed in vacuo and the product was washed with diethyl ether, which was then removed in vacuo on a rotary evaporator. This sodium salt (210 mg, 0.85 mmol, 73.9% yield) and [(η6-p-cymene)RuCl2]2 (240 mg, 0.39 mmol) were stirred in acetone (25 ml) at RT for 40 min. After the extraction step using dichloromethane (see Example 1), the final product was dissolved in acetone, concentrated on a rotary evaporator, and diethyl ether was added. The solution was left in a freezer at 253 K overnight. The crystals (238 mg, 0.48 mmol, 61.8% yield) were collected and dried in vacuo. 1H NMR (CDC13): δ 7.88 (m, 2H), 7.43 (m, 2H), 7.36 (m, 2H), 6.43 (s, 1H), 5.57 (d, 2H), 5.30 (d, 2H), 3.01 (sp, 1H), 2.33 (s, 3H), 1.39 (d, 6H).
Example 4
[(η6-p-cymene)RuCl((CH3)3CCOCHCOC(CH3)3- O.O)]
2,2,6,6-tetramethyl-3,5-heptanedione (299 mg, 1.62 mmol) and sodium methoxide (68 mg, 1.26 mmol) were stirred in methanol (40 ml) at RT for 5 h. The solvent and the excess liquid starting material were removed in vacuo on a rotary evaporator. The obtained sodium salt (233 mg, 1.13 mmol, 89.6% yield) and [(η6-p-cymene)RuCl2]2 (258 mg, 0.42 mmol) were stirred in acetone (25 ml) at RT for 40 min. After the extraction step using dichloromethane (see Example 1), the residue was dissolved in diethyl ether, concentrated on a rotary evaporator, and hexane was added. The solution was left in a freezer at 253 K overnight. The crystals (215 mg, 0.47 mmol, 56.0 % yield) were collected and
dried in vacuo. 1H NMR (CDC13): δ 5.38 (d, 2H), 5.37 (s, 1H), 5.10 (d, 2H), 2.88 (sp, 1H), 2.21 (s, 3H), 1.33 (d, 6H), 1.10 (s, 18H).
Example 5
[(η6-C6C6H5ORuCl(H3CCOCHCOCH2 - 0.0)]
[(η6-C6H5C6H5)RuCl2]2 (250 mg, 0.38 mmol) was refluxed in water (25 ml) for 2 h. The heat was reduced below boiling point and sodium acetylacetonate monohydrate (141 mg, 1.01 mmol) was added. The solution was stirred for 30 min and hot filtered. The solvent was removed in vacuo on a rotary evaporator. After the extraction step using dichloromethane (see Example 1), the residue was dissolved in little acetone, and diethyl ether was added until precipitation occurred. The vessel was left in a freezer at 253 K for 6 d. The product (65 mg, 0.16 mmol, 21.3% yield) was collected and dried in vacuo. 1H NMR (CDC13): δ 7.67 (m, 2H), 7.45 (m, 3H), 5.83 (m, 2H), 5.75 (m, 3H), 5.10 (s, 1H), 1.83 (s, 6H).
Example 6
[η6-p-cymene)Ru(CH,CN)(H3CCOCHCOCH3 - 0.0))BF4
The compound of Example 1 (144 mg, 0.39 mmol) and AgBF4 (72 mg, 0.37 mmol) were stirred in acetonitrile (20 ml) overnight. The resulting solution was filtered through glass wool and the solvent was removed in vacuo on a rotary evaporator. The residue was dissolved in a minimum amount of acetone, and diethyl ether was added until precipitation occurred. The flask was placed in a freezer at 253 K for 4 d. The crystals (149 mg, 0.32 mmol, 87.1% yield) were collected and dried in vacuo. 1H NMR (CDC13): δ 5.74 (d, 2H), 5.48 (d, 2H), 5.16 (s, 1H), 2.85 (sp, 1H), 2.43 (s, 3H), 2.21 (s, 3H), 1.98 (s, 6H), 1.33 (d, 6H).
Example 7
[η6-p-cymene)RuCl(H3CCOCCOCH3, - 0.0)]2
(Formula Removed)
The preparation of the tetradentate ligand tetraacetylethane has been reported by Robert. G. Charles in Organic Syntheses (1959), 39, 61. Tetraacetylethane (208 mg, 1.05 mmol) was reacted in water with 21 ml of a 0.1 M solution of KOH in methanol for 2 h. The solvents were evaporated and the salt was dissolved in acetone. [(p-Cymene)RuCl2]2 (643 mg, 1.05 mmol) was added to the solution and the mixture was reacted for 3 h. The solvent was evaporated and the residue was extracted with CH2Cl2. Filtration and evaporation of the solvent gave the pure dinuclear complex, which was recrystallized from acetone (550 mg, 71%).
B. Biological Data
1. Protocol for testing Ru compounds
The compounds are tested on 24-well trays. Cells growing in a flask are harvested just before they become confluent, counted using a haemocytometer and diluted down with media to a concentration of lxl04 cells per ml. The cells are then seeded in the 24-well trays at a density of lxl04 cells per well (i.e. 1ml of the diluted cell suspension is added to each well). The cells are then left to
plate down and grow for 72 hours before adding the compounds of the
invention.
The Ru complexes are weighed out and made up to a concentration of lmg/ml with deionised water and 0.5 % w/w DMSO. The appropriate volume of the Ru solution is added to 5ml of media to make it up to a concentration of 100 µM for each drug. This 100 µM solution is then serially diluted to make up the 10 µM, 1 uM and 0.1 µM solutions.
The media is removed from the cells and replaced with 1ml of the media dosed with drug. Each concentration is done in duplicate. A set of control wells are left on each plate, containing media and 0.5 % w/w DMSO without drug.
The cells are left exposed to the drugs for 24 hours and then washed with phosphate buffered saline before fresh media is added.
They are allowed to grow on for a further 3 days before being counted using a Coulter counter.
Preparing cells for counting:
Media is removed and 1ml of PBS is added to the cells.
250 µl of trypsin is added and cells left in incubator for a few minutes to allow
the monolayers to detach.
Once trypsinised, 250µl of media is added to each well to neutralise the trypsin.
200µl of this suspension is added to 10ml of NaCl for counting.
2. Results
Using the above protocol, a number of compounds of the invention were tested on A2780 ovarian cancer cell line. Compounds of the invention have an IC50 of less than 150 µM, preferably less than 100 µM, more preferably less than 50 µM when tested according to this protocol. The results are as follows:

(Table Removed)
C. NMRData
The ability of the compounds of the invention to bind to DNA bases was studied by nmr and the results are illustrated in the accompanying figures in which:
Figure 1 is a 500 MHz 1H NMR spectrum of a mixture of guanosine : adenosine : compound of Example 1 in the mole ratio 1.3 : 1 : 1, respectively, inl0%D2O/90%H2O.
Figure 2 shows expansions of the regions 5.1-5.4 ppm and 7.9-8.6 ppm of the 1H NMR spectrum shown in Fig. 1.
In Figure 2, the region from 5.1 to 5.4 ppm (right) corresponds to the central CH on the acac ligand. Peaks a and b correspond to the adenosine adducts,
peak c to the guanosine adduct. In the region 7.9- 8.6 ppm (left), peaks for the H2 and H8 protons of adenosine and H8 protons of guanosine appear. It can be seen that both adenosine and guanosine form adducts with the compound of Example 1 (bound labels). The atom labelling scheme for the nucleobase part (A and G) of the nucleosides is shown.




WE CLAIM:
1 Ruthenium(II) compound of formula(I) :
(Formula Removed)
wherein: R1, R2, R3, R4, R5 and R6 independently represent H, (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, halo, CO2R7, CONR8R9, COR10, SO3H, SO2NR11R12, aryloxy, (C1-C6)alkoxy, (C1-C6)alkylthio, -N=N-R13, NR14R15, aryl or aralkyl. which latter two groups are optionally substituted on the aromatic ring by one or more groups independently selected from (C1-C6)alkyL (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, aryl, aralkyl, halo, CO2R7a, CONR8aR9a, COR10a, SO3G, SO2NRllaR12a, aryloxy, (C1-C6)alkoxy, (C1-C6)alkylthio, -N=N-R13a, NR14aR15a, or R1 and R2 together with the ring to which they are bound represent a saturated or unsaturated carbocyclic or heterocyclic group containing up to three 3-to 8-membered carbocyclic or heterocyclic rings, wherein each carbocyclic or heterocyclic ring may be fused to one or more other carbocyclic or heterocyclic rings, and wherein each of the rings may be optionally substituted by one or more groups independently selected from (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(Cr C6)alkyl, amino(C1-C6)alkyL aryl, aralkyl, halo, CO2, CONR8bR9b, COR10b, SO3G', SO2NRllbR12b, aryloxy, (C1-C6)alkylthio, -N=N-R13b, NR14bR15b or (C1-C6)alkoxy;
R7. R8 R9. R10, R11, R12, R13 R14, R15, R7a, R8a. R9a, Rl0a, Rlla. Rl2a, R13a. R14a. Rl5a.
R7b. R8b. R9b„ R10b, Rllb. Rl2b, R,13b, R14b. and R15b are independently selected from
H, (C1-C6) alkyl, aryl or aralkyl;
X is a neutral or negatively charged O-, N- or S-donor ligand or halo;
G and G' are independently selected from alkali metals, aryl. aralkyl and (C1-C6)
alkyl;
Y-L-Y" is a bidentate ligand bearing a negative charge with a proportion of the
charge on both Y and Y", Y and Y' are independently selected from O, S or NR16,
wherein R16 is H, (C1-C6) alkyl, aryl or aralkyl and L is a group linking Y and Y"
and comprises one or more groups selected from (C1-C6) alkylene, (C1-C6)
alkenylene, (C1-C6) alkynylene, arylene, arylene, aralkylene. alkarylene, each of
said latter six groups being optionally substituted, ferrocenylene, Se, Se-Se. S-S,
N=N and CO;
m is -1 or +1 and the compound comprises a counterion such as herein described
when m is -1 or +1;
the compound of formula (I) optionally being in the form of a dimer in which two L
groups are linked either directly or through a group comprising one or more of (C1-
C6) alkylene, (C1-C6) alkenylene, arylene, aralkylene, alkarylene, Se, Se-Se, S-S,
N==N and C=O or in which L bears two Y groups and two Y' groups;
with the provisos that:
when Y-L-Y' is (CH3C(O)CHC(O)(CH3)\ X is halo or an N-donor ligand, R1. R2,
R3, R , R and R together with the ring to which they are bound do not represent 4-
isopropy 1-1 -methylbenzene:
when Y-L-Y' is (CH3C(O)CH(O)CH3)" and X is chloro. (CH3)2SO, CH3CN,
pyridine or (CH3C(O)CHC(O)CH3)": R1, R2, R3, R4, R5 and R6 are not all H or all
methyl; R1, R3 and R5 are not all H when R2, R4 and R6 are all methyl; and R2, R4
and R6 are not all H when R1, R3 and R5 are all methyl;
when Y-L-Y' is (CF3C(O)CHC(O)CHF3)" and X is chloro, R1, R2, R3, R4, Rs and
R6 are not all H or all methyl; R1, R3 and R5 are not all H when R2, R4 and
R6 are all methyl; and R2, R4 and R6 are not all H when R1, R3 and R5 are all
methyl;
when Y-L-Y' is (CF3C(O)CHC(OPEt) and X is chloro, R1, R2, R3, R4, R5
and R together with the ring to which they are bound do not represent 4-
isopropyl-1 -methylbenzene;
when Y-L-Y' is ((ferrocenylene)C(O)CHC(O)(ferrocenylene))" and X is
chloro, R1, R2, R3, R4, R5 and R6 together with the ring to which they are
bound do not represent 4-isopropyl-l-methylbenzene;
when R1, R2, R3, R4, R5 and R6 together with the ring to which they are bound
represent 4-isopropyl-l-methylbenzene and X is chloro, Y-L-Y' is neither (3-
OCH3-4-OH-PhCHCHC(O)CHC(O)CHCH-3-OCH3-4-OH-Ph); (3-OCH3-4-
OCH3-PhCHCHC(O)C(CH3)C(O)CHCH-3-OCH3-4-OCH3-Ph)", (3-OCH3-4-
COOCH3-PhCHCHC(O)CHC(O)CHCH-3-OCH3-4-COOCH3-Ph)", (4-OH-
PhCHCHC(O)CHC(O)CHCH-4-OH-Ph)~ nor (3-OCH3-4-OCH3-
PhCHCHC(O)CHC(O)CHCH-3-OCH3-4-OCH3-Ph):
2. Compound as claimed in Claim 1, wherein R1, R2, R3, R4, R5 and R6 are independently selected from H (C1-C6) alkyl and phenyl or R1 and R2 together with the ring to which they are bound represent anthracene or a hydrogenated derivative of anthracene, said phenyl and anthracene or a hydrogenated derivative of anthracene group being optionally substituted by one or more groups independently selected from (C1-C6)alkyl, (C2-C6) alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, phenyl, benzyl, halo, carboxyl, CO2(C1-C6)alkyl, CONH2, COH, CO(C1-C6)alkyl, SO3H, SO2NH2, phenoxy. (C1-C6)alkylthio, NH2 or (C1-C6) alkoxy.
3. Compound as claimed in Claim 1 or Claim 2, wherein m is 0.
4. Compound as claimed in any one of Claims 1 to 3, wherein X is halo or
CH3CN.
5. Compound as claimed in any one of Claims 1 to 4, wherein Y-L-Y' is selected from ligands of formulae (II) to (X):
(Formula Removed)
wherein T and T' are independently selected from O and S.
R1g and R3g are independently H, (C1-C6) alkyl, aryl or aralkyl,
Ric to R5f and R2g are independently H, (C1-C6)alkyl, aryl, aralkyl, wherein the
latter two groups and the corresponding groups for R1g and R3g are optionally
substituted by one or more groups independently selected from (C1-C6alkyL
(C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, aryl,
aralkyl, halo, CO2R7b, CONR8bR9b, COR10b, SO3G', SO2NRllbR12b, aryloxy,
(C1-C6)alkylthio, -N=N-R13b, NR,4bR15b or (C1-C6)alkoxy, wherein R7b, R8b, R9b,R10b, R11b, R12b, R13b, R14b, and R15b are defned in Claim 1
6. Compound as claimed in any one of Claims 1 to 4, wherein Y-L-Y1 is selected from:
(Formula Removed)
wherein T, T, T" and T"' are independently selected from O and S,
A comprises one or more groups selected from (C1-C6)alkylene, (C1-
C6)alkenylene, (C1-C6) alkynylene, arylene, aralkylene, alkarylene,
ferrocenylene, Se, Se-Se, S-S, N=N and C=O
and R1h to R6j are independently H, (C1-C6)alkyl, aryl, aralkyl, wherein the
latter two groups are optionally substituted by one or more groups
independently selected from (C1-C6)alkyL (C2-C6)alkenyl, (C2-C6)alkynyl,
hydroxy(C1-C6)alkyl, amino(C1-C6)alkyL aryl, aralkyl, halo, CO2R ,
CONR8bR9b, COR,10b, SO3G', SO2NRllbR12b, aryloxy, (C1-C6)alkylthio, -N=N-
R13b, NR14bR15b or (C1- C6)alkoxy, wherein R7b, R8b, R9b, R,10b, R,11,b, R12b, R13b,
R14b, and R15b are as defined in Claim 1.
7. Compound as claimed in any one of Claims 1 to 4, wherein Y-L-Y1 is:
(Formula Removed)
wherein T and T are independently O and S, and R, R1c, and R3c are independently H, (C1-C6)alkyl, aryl, aralkyl, wherein the latter two groups are optionally substituted by one or more groups independently selected from (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, hydroxy(C1-C6)alkyl, amino(C1-C6)alkyl, aryl, aralkyl, halo, CO2R7b, CONR8bR9b, COR10b, SO3G', SO2NRllbR12b, aryloxy, (C1-C6)alkylthio, -N=N-RBb, NR,4bR15b or (C1-C6)alkoxy, wherein R7b, R8b, R9b, R,10b, R11b, R12b, R,13b, R,14b, and R15b are as defined in Claim 1.
8. Compound as claimed in claim 7, wherein one ofR1,R2,R3,R4,R5and R is phenyl and the other groups are H.
9. Compound as claimed in Claim 7 or Claim 8, wherein T and T are both O, R is H or (C1-C6) alkyl and Rlc and R3c are independently (C1-C6)alkyl or phenyl, said phenyl optionally substituted by (C1-C6)alkyl, hydroxy(C1-C6)alkyl, amino(C1-C6) alkyl, halo, carboxyl, CO2(C1-C6)alkyl, CONH2, COH, CO(C1-C6)alkyl, SO3H, SO2NH2, phenoxy, (C1-C6) alkylthio, NH2 or (C1-C6)alkoxy.
10. Compound as claimed in claim 9, wherein R1c and R3c are independently phenyl, said phenyl optionally substituted by (C1-C6)alkyl, hydroxy(C1-C6)alkyl, amino(C1-C6) alkyl, halo, carboxyl, CO2(C1-C6)alkyl, CONH2, COH, CO(C1-C6)alkyl, SO3H, SO2NH2, phenoxy, (C1-C6) alkylthio, NH2 or (C1-C6)alkoxy.
1 1. Compound as claimed in claim 9. wherein R is H and Rlc and R3c are independently (C1-C6) alkyl or phenyl.
12. Compound as claimed in any one of claims 1 to 9, wherein Y and Y' are both O.
13. Compound as claimed in any of claims 1 to 12, as and when used as a medicament for the treatment and/or prevention of cancer.

Documents:

276-delnp-2005-abstract.pdf

276-DELNP-2005-Claims.pdf

276-delnp-2005-complete specification(as filed).pdf

276-delnp-2005-complete specification(granted).pdf

276-delnp-2005-correspondence-others.pdf

276-delnp-2005-correspondence-po.pdf

276-DELNP-2005-Description (Complete).pdf

276-DELNP-2005-Drawings.pdf

276-delnp-2005-form-1.pdf

276-delnp-2005-form-18.pdf

276-DELNP-2005-Form-2.pdf

276-delnp-2005-form-3.pdf

276-delnp-2005-form-5.pdf

276-delnp-2005-gpa.pdf

276-delnp-2005-pct-210.pdf

276-delnp-2005-pct-304.pdf

276-delnp-2005-pct-409.pdf

276-delnp-2005-pct-416.pdf

abstract.jpg


Patent Number 245353
Indian Patent Application Number 276/DELNP/2005
PG Journal Number 03/2011
Publication Date 21-Jan-2011
Grant Date 14-Jan-2011
Date of Filing 25-Jan-2005
Name of Patentee THE UNIVERSITY COURT,THE UNIVERSITY OF EDINBURGH
Applicant Address OLD COLLEGE,SOUTH BRIDGE,EDINBURGH EH8 9YL,U.K
Inventors:
# Inventor's Name Inventor's Address
1 MELCHART,MICHAEL 96/11 NICHOLSON STREET,EDINBURGH EH8 9EW,U.K
2 SADLER,PETER JOHN MILL LADE COTTAGE,4A VALLEYFIELD ROAD,PENICUIK EH26 8LW,U.K
3 FERNANDEZ LAINEZ,RAFAEL 7 LASSWADE GROVE,EDINBURGH EH17 8HX,U.K
4 HABTEMARIAM,ABRABA 7/6 CHESSELS COURT,240 CANONGATE,EDINBURGH EH8 8AD,U.K
5 JODRELL DUNCAN IAN 34 BRUNBRAE,EDINBURGH EH12 8UB,U.K
PCT International Classification Number C07F 15/00
PCT International Application Number PCT/GB 2003/002879
PCT International Filing date 2003-07-04
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 0215526.5 2002-07-05 U.K.