INDOLOQUINONE DERIVATIVES AS BIOREDUCTIVE AGENTS The present invention relates to the use of a group of indoloquinones as therapeutic agents, particularly as bioreductively activated anti-tumour agents, to novel members of this group and compositions containing them and to methods of using these to treat tumours. Fused cyclopropamitosenes and closely related indoloquinones have recently been evaluated as novel bioreductive anticancer agents (references 1-3 below). These compounds were originally designed as analogues of the archetypal quinone bioreductive alkylating agent mitomycin C (Figure 1 herein; MMC, compound 1) having much reduced electophilicity at C-l due to the inertness of the 1,2-cyclopropane compared to the aziridine in MMC. Certain indoloquinones (e.g. Figure 1; compound 2) in this series have been found to be highly potent cytotoxins as compared with MMC and in some cases have substantially higher hypoxic cytotoxicity ratios (HCR) (see references 2-3). The cyclopropamitosenes have been found to be more rapidly reduced by DT-diaphorase, an important activator of mitosenes, compared to MMC, but this could not explain fully the higher potency of the compounds compared to the current lead clinical agent of this type EO9 (3), which is reduced two orders of magnitude more rapidly than compound 2 by DT-diaphorase (see WO 87/06227 ≡ US 5097257).

The presence of particular groups as the 5-substituent, in particular aziridines and substituted aziridines, and as the leaving group on the 3-methylene substituent was therefore identified as an important feature, both in terms of oxic and hypoxic potency and rate of reduction, i.e. ability to act as substrates for reductase enzymes. This does not vary greatly among 5-aziridine and 5-methoxy drivatives (see references 2 and 6 below), but it should be noted that the relative rates of reduction of this type of compound by one-electron reductases, which will be important under hypoxic conditions, is unknown, and there is very little data on one electron reduction potentials of mitosenes.

Ionic ring-opening of the fused cyclopropane is considered unlikely but radical ring opening of the cyclopropane to give a reactive H-atom abstractor has been suggested as a possible explanation for hypoxic potency in particular. There has however been no direct evidence for this mechanism.

The present inventors have now designed and synthesized analoguesOf compound 2 wherein inter alia (i) cyclopropane rings are included not fused to the indoloquinone ring system, (ii) lower alkyl eg. isopropyl analogues of these that are structurally very closely related to 2 but unable to undergo radical ring-opening reactions and (iii) analogues containing no substituent, ie. where Rr is hydrogen. These derivatives have been designed as closer analogues of EO9 with the aim of targeting and exploiting DT-diaphorase activity. Particularly desirable is the optimization of hypoxic-cytotoxicity by such modifications and this novel series of compounds offers many compounds of relatively high HCR.

Some of the compounds used by the invention are disclosed in US patents US 3226397, US 3226398, US 3226399, US 3265698, US 3267117 and US 3226385; all corresponding to the single UK patent application GB 1087325. The UK documents lists three hundred and fifteen examples of which twenty seven are specified as having anti-microbial activity.

The compounds for the use of the present invention form a selection from the general formula of GB 1087325 and are those of the general class referred to in US 5097257 but distinguished therefrom by, inter alia, particular groups at R ¹ and R ² of the formula I below. The formula I, and particularly preferred members of that formula such as those of formula II, offer compounds with advantages of selective hypoxic and anoxic potency. Thus in a first aspect of the present invention there is provided the use of a compound of general formula I

or a salt thereof wherein

R and R ⁴ are independently selected from hydrogen, halogen and C _j _g alkyl or haloalkyl,

C2.6 alkenyl or haloalkenyl, C _g alkoxy, phenoxy, C _j .g alkylthio, phenylthio, primary and secondary amino or hydroxy groups and

R? is hydrogen, hydroxy, a C _j _g alkyl or haloalkyl, C2_g alkenyl or haloalkenyl or C _] .5 alkoxy or haloalkoxy group for the manufacture of a medicament for the treatment of neoplasms, particularly tumours and more particularly cancer tumours, characterised in that

R* is selected from a C .g alkyl or haloalkyl group, -CO2R , where R^ is hydrogen or a

C _j _g alkyl or haloalkyl group, or a group -CH2-X, where X is selected from groups of

where R" is hydrogen or a leaving group the acid HR ⁶ of which has a pKa of 10 or less and R' and R° are the same or different and are selected from C _j .g alkyl or haloalkyl or together with the interjacent nitrogen form a heterocyclic ring of 5 to 7 atoms optionally substituted by a C _j _^ alkyl group or haloalkyl and

R^ is selected from hydrogen, C _j _4 alkyl and haloalkyl and -(CH2) _n CHR ⁹ R ¹⁰ of more than four carbons where n is an integer of 0 to 2 and R? and R* " are independently selected from a C _j _4 alkyl or haloalkyl group, or R ⁹ and R*" together with the interjacent carbon atom form a 03.7 cycloalkyl or cycloalkenyl ring optionally substituted with one or more Cι_ alkyl or haloalkyl, or C2.4 alkenyl or haloalkenyl groups.

In a preferred use of the invention R is a C _j _g alkoxy, haloalkoxy, C2_4 alkenoxy or haloalkenoxy, phenoxy, C _j .g alkylthio, phenylthio, primary or secondary amino or hydroxy group; R^ is preferably hydrogen, hydroxy, a C _j _g alkyl or haloalkyl, C2_g alkenyl or haloalkenyl or C _j _g alkoxy group and R ⁴ is preferably hydrogen, halogen or a C _j .g alkyl or haloalkyl, C2_4 alkenyl or haloalkenyl group.

Preferred salts are suitable for direct adminstration to patients and thus are pharmaceutically or 'physiologically' acceptable salts. Such salts may be made with a compound described above and a physiologically acceptable acid or, where the compound contains a suitable negatively chargeable group, base. Thus salts may be formed with various inorganic and organic acids. Examples of these acids are phosphonic, nitric,

sulphuric, hydrohalic, citric, oxalic, fumaric, maleic, lactic, succinic, malic, tartaric and methane sulphonic acids. Preferred halide salts are those of hydrochloric, hydrobromic or hydroiodic acid. Examples of bases are alkali metal hydroxides and quaternary ammonium hydroxides eg.2-arnino-2-hydroxymethyl propane- 1,3-diol (Tris) salts. R is preferably selected from C _j ^ alkoxy, C _4 haloalkoxy, C ₂ _4 alkenoxy,

C ₂ .4 haloalkenoxy and aziridin-1-yl groups, the aziridin-lyl groups being optionally substituted with C _4 alkyl, or C _j _ haloalkyl or C ₂ _4 alkenyl or C ₂ _4 haloalkenyl.

R ² is preferably selected from C _ n-alkyl or -haloalkyl groups or groups of formula -(CH ₂ ) _rι CHR ⁹ R ¹⁰ ' where n is an integer from 0 to 2, where R ⁹ and R ¹⁰ are independently selected from C _j _ _Δ alkyl and C _j _^ haloalkyl or together with the interjacent carbon atom form a C^. cycloalkyl or cycloalkenyl group optionally substituted with

C _j _ ₄ alkyl, C _j _ haloalkyl, C ₂ _4 alkenyl or C2.4 haloalkenyl.

Preferably Rr is a C _j _4 n-, iso- or cyclo-alkyl group, particularly a methyl or cyclopropyl group ^s - R^ is more preferably selected from C _j _4 alkyl or haloalkyl and C2_4 alkenyl and haloalkenyl groups.

R^ is more preferably selected from hydrogen, C .4 alkyl and haloalkyl groups and most preferably is hydrogen.

Particularly preferred compounds are those where R is aziridin-1-yl optionally substituted with C _j .4 alkyl or C _j .4 haloalkyl or where R is C _j _ _A alkoxy.

One group of preferred compounds are those where the group R ¹ is selected from -CH ₂ -O-R ⁶ and -CH ₂ -S-R ⁶ where R ⁶ is hydrogen or where R ⁶ is an optionally substituted phenyl or benzyl group or a group -C(O)-Rl 1 where R' ¹ is an optionally substituted phenyl, benzyl or amino group; examples of such groups R* including alkoxy, aryloxycarbonyloxy, aryloxycarbonyloxy and carbamoyloyloxy. Optional substituents include halo, C _j .g alkyl or haloalkyl, nitro and sulpho groups. Conveniently R ¹ is hydroxy methyl.

Still more preferred compounds are those wherein R ¹ is a group -CH2X where X

R ⁷ is -N^ wherein R' and R , together with the interjacent nitrogen form a 5 to 7 membered ^X R8

heterocyclic ring containing nitrogen and carbon with optional oxygen or sulphur members, e.g. piperazine, morpholine, thiomoφholine rings. More preferably the heterocyclic ring is a piperazinyl ring optionally substituted by one or more C _j .4 alkyl or haloalkyl groups. It is particularly preferred that X is a 4-alkylpiperazin-l-yl group, eg. a 4-methylpiperazine group.

R? is most preferably a C _j ^ alkyl group and R ⁴ is most preferably hydrogen.

Particularly preferred compounds include those where R is aziridin-1-yl optionally substituted with C _j _4 alkyl, eg. 2-methylaziridin-l-yl, R ² is cyclopropyl or isopropyl or C _ n-alkyl, R ³ is methyl and R ⁴ is hydrogen. The most preferred compounds for the use of the invention are those which have a

C< _j θ(Air)μM/C5o(N2)μM ratio of 40 or more, still more preferably of 100 or more. No particular upper limit applies to preferred compounds as a relatively selective action against hypoxic or anoxic cells is desired. It is also preferred that the compounds have a hypoxic potency such that their value is less than lOμM, more preferably less than 1 μM and most preferably less than 0.01 μM. Typically this is in the range 0.005 to 1 μM.

Most preferred compounds are described herein as compounds 21, 54 and 84 of the Examples described below; the numbers being in brackets at the end of the example title.

In a second aspect of the invention there are provided novel compounds falling within formula I above. Particular novel compounds of the invention are those of formula la below. Thus this second aspect of the present invention provides a novel compound of formula la

or a salt thereof characterised in that

-

R is C _4 alkoxy or haloalkoxy or aziridin-1-yl optionally substituted with one or more

C _4 alkyl or haloalkyl groups,

Rr is hydrogen or a C _ alkyl or haloalkyl group,

R ³ is hydrogen, hydroxy, C _j _ ₆ alkyl or haloalkyl, C ₂ _g alkenyl or haloalkenyl or C _g alkoxy or haloalkoxy,

R ¹ is -CO ₂ R where R ⁵ is a hydrogen or C _} _ ₆ alkyl or haloalkyl group, or is a group

-CH2X where X is selected from -S-R ⁶ , -O-R ⁶ and -NR ⁷ R ⁸ where R ⁶ is hydrogen or a leaving group the acid HR ⁶ of which has a pKa of 10 or less and R? and R are the same or different and are selected from C _j .g alkyl or haloalkyl or together with the interjacent nitrogen form a heterocyclic ring of 5 to 7 atoms optionally substituted by a C _j .

4 alkyl or haloalkyl group and

R ² is hydrogen or a Cμ alkyl group or a group -(CH2) _n CHR ⁹ R ¹⁰ of more than four carbons where n is an integer of 0 to 2 and R ⁹ and R^" are independently selected from

C .4 alkyl and haloalkyl or R9 and Rl 0 together with the interjacent carbon atom form a 03.7 cycloalkyl or cycloalkenyl ring optionally substituted with one or more C _4 alkyl or haloalkyl or C2_ alkyl or haloalkyl groups; with the provisos that

(i) when R is methoxy, R ¹ is hydroxymethyl or a carbamate or C _j _g aliphatic ester thereof, Rr is methyl or ethyl and R ³ is methyl, optionally 2-substituted ethyl, propyl or butyl, then R ² is not hydrogen, methyl, fluoromethyl, chloromethyl or ethyl,

(ii) when R is methoxy, R is hydrogen, R is hydroxymethyl or the propylcarbamate thereof and R ³ is ethyl, then R ² is not methyl,

(iii) when R is methoxy, R ² is methyl, R ³ is ethyl and Rr is methyl, then R ¹ is not hydroxymethyl cyclohexylcarboxylate, benzoate, furanyl-2-carboxylate, 3-(2-dimethyl- aminoethyl)-piperazine-l-carboxylate, morphalinocarbamate, 4-(3-hydroxypropyl)- piperazinecarbamate or 4-(3-dimethylaminopropyl)-piperazinecarbamate, and

(iv) when R is aziridinyl or ethoxy, R ¹ is hydroxymethyl or a carbamate or C _j _g aliphatic ester thereof, R ⁴ is methyl or bromo and R ³ is ethyl, then R ² is not methyl.

Preferred novel compounds of the invention are those where R ¹ is a hydroxymethyl group or a group -CH2-X as defined above. Most preferred novel compounds are those where R is aziridin-1-yl or 2-alkyl-aziridin-l-yl, such as 2-methyl-aziridin-l-yl.

R ³ is preferably C _j _4 alkyl or haloalkyl and R ² is preferably n-C _j ^ alkyl, isopropyl or C3.7 cycloalkyl. R ⁴ is preferably methyl or hydrogen, most preferably hydrogen.

A third aspect of the present invention provides a novel compound of the second aspect of the invention or a physiologically acceptable salt thereof for use as a therapeutic agent. A preferred use as therapeutic agent as provided by this invention is for the treatment of rumour cells, and most preferably the use is that which adminsters the compound in treatment also comprising radiation treatment, e.g. with therapeutic radiation such as X-rays, γ-rays and electrons from an accelerator, e.g. linear accelerator. The use of the compound with radiation is particularly effective for achieving cytostasis and/or death of tumour cells, particularly of cancer cells. Most preferably the use is aimed at providing cytotoxicity of anoxic tumour cells, ie. those relatively poorly supplied with blood.

Particularly the use is aimed at providing cytotoxicity towards hypoxic and anoxic tumour cells which, at the time of treatment, exist at an oxygen tension of less than 7.6 mm Hg of O ₂ (1%). A fourth aspect of the present invention provides compositions comprising a novel compound of the second aspect of the invention or a pharmaceutically acceptable salt thereof together with a pharmaceutically acceptable carrier. Compositions will preferably be provided as discrete dosage units and packs thereof, e.g. in a form suitable for parenteral or oral adπainstration. Other suitable dosage unit forms will occur to those skilled in the art A fifth aspect of the present invention provides a method of treating a human or animal body for the purposes of killing, and/or preventing growth of, tumour cells comprising aά'ministering a medicament manufactured with the compound of the first aspect of the invention, or a pharmaceutically acceptable salt thereof, to the body in a dose sufficient to kill or prevent growth of some or all of the tumour cells.

Preferred methods of the fifth aspect are for treating hypoxic and anoxic tumour cells, particularly cancer cells. It will be realised that the treatment will usually include killing of oxic cells, but that in preferred forms the ratio of such cells killed as compared with anoxic or hypoxic cells will be relatively low. A still more preferred method of the fifth aspect comprises administering the medicament manufactured with the compound or salt of the first aspect to the body within a set period of treatment of the body with doses of radiation, the doses of radiation being sufficient to kill or prevent growth of some or all of any oxic tumour cells eg. those relatively well supplied with blood. This combination treatment allows targeted radiation to be used to damage and/or kill the oxic tumour cells while the systemically adminstered compound of the invention is used to damage or kill the hypoxic and anoxic cells selectively, thus causing reduced peripheral damage to healthy non-tumour cells.

The compounds of the present invention are of the indoloquinone class and as such will have application in treatment of tumours such as those vulnerable to mitomycins and the compounds disclosed in US 5079257. These include all solid tumour types, for example cervical, breast, pancreatic and prostatic adenocarcinoma, colon, bladder, gastro-intestinal cancers and melanomas. Such tumours may exist in an area of hypoxia and contain the reductive enzymes necessary to activate the agents for the use of this invention. A sixth aspect of the invention provides novel compounds that have utility as intermediates for use in the preparation of a compound of the first aspect Of the invention, these intermediates being novel compounds as described in the Figures 2 to 5 and the Examples 1 to 49.

Particularly preferred novel intermediates are those of formula II below

Formula II

wherein R is C _j _4 alkoxy, R is C _4 alkoxycarbonyl and R , R ³ and R ⁴ are as for formula la. It will be realised that some of these compounds will also be compounds of the first aspect of the invention but, where these have an HCR of less than 40, and particularly have an HCR of less than 10, their main interest will be as intermediates for use in synthesis of the more preferred compounds. In this respect the 5-alkoxy compounds will usually have a less favourable efficacy and HCR ratio than the corresponding aziridinyl compounds and thus their prime use will be as intermediates. Preferred intermediates are those where R is methoxy, R ¹ is methoxycarbonyl and R ³ is methyl.

A seventh aspect of the present invention provides a process for preparing compounds of formula la and II .

In a first preferred process of this aspect there is provided a process for preparing an optionally substituted aziridin-l-yl compound of formula II where R^ is alkoxycarbonyl comprising reacting a corresponding 2-substituted-3-alkoxycarbonyl-5-alkoxy-l-alkyl- indole-4,7-dione with an optionally substituted aziridine. In a second preferred process for preparing a compound of formula la where R ¹ is a hydroxymethyl group a corresponding 2-substituted-3-alkoxycarbonyl-5-alkoxy-l-alkyl- indole-4,7-dione is reacted with an oxidising reagent, such as Na2S2θ4, then the resultant hydroquinone is reduced, e.g. with DIBA -H, to the product.

For all aspects of the present invention halo is preferably fluoro or chloro. The compounds, compositions, use, methods and intermediates of the invention will now be described by way of illustration only by reference to the following non-limiting Figures and Examples. Further embodiments of the invention will occur to those skilled in the art in the light of these. Figures. Figure 1 : Shows the chemical structure of some known compounds referred to in the specific description above as compounds 1 to 5.

Figure 2: Shows the outline of the synthetic route used to prepare compounds 10, 11, 12 and 27, 28, 29 referred to herein.

Figure 3: Shows the outline of the synthetic route used to prepare compounds 13 to 42 referred to herein.

Figure 4: Shows the outline of the synthetic route used to prepare compounds 47 to 57 referred to herein.

Figure 5: Shows the outline of the synthetic route used to prepare compounds 58 to 66 referred to herein.

Figure 6: Is a graph plotting mean relative surviving RIF-1 tumour cell fraction against a a varying dose of compound 21 alone and against varying dose of compound 21 adminstered after 15 GY of radiation. The surviving fraction after 15 GY radiation alone is shown by a flat line.

Figure 7: Is a graph plotting mean relative surviving RIF-1 tumour cell fraction under conditions as set out for Figure 6 but using compound 54 as test compound.

Figure 8: Is a graph plotting mean relative surviving KHT tumour cell fraction against a varying dose of compound 21 alone and against a varying dose of compound 21 adminstered after 10 GY of radiation. The surviving fraction after 10 GY radiation alone is shown by a flat line.

Figure 9: Is a graph plotting mean relative surviving KHT tumour cell fraction under conditions as set out for Figure 8 but using compound 54 as test compound.

Definitions of the groups used in the Figures can be found in the text of Examples below and in the Figures themselves.

EXAMPLES

General methods for preparation of compounds 18 to 84:

2- Alkyl and cycloalkyl substituted indoles were synthesized in 14 steps from the common precursor 3-chlorophenol as shown in Figure 2 and 3. The crucial step was the 1 ,5-electrocyclisation of the imine (e.g. compound 8) to the 2,3-dihydroindole derivatives, which was successfully carried out using isobutyraldehyde, cyclohexane carboxaldehyde or cyclopropane carboxaldeyde, but not with acrolein in a proposed alternative route, using zinc acetate in methanol as has been employed in the synthesis of EO9 via a 2-acrylate derivative in WO 87/06227. Nitration at the desired 4-position could be achieved only subsequent to the N-methylation step in order to avoid increasing yields of the 6-nitro isomer. The subsequent 6 steps, including nitration, oxidations (DDQ and Fremy's salt) and reductions (Sn/HCl, LiAli^, Na ₂ S2θ4 and DIBAL-H) left the 2-cyclopropyl moiety intact and the desired indoloquinones were obtained. Substitution of the 5-methoxy substituent was successful in high yielding reactions with aziridine and 2-methylaziridine.

Comparable 1,2-dimethyl analogues 52-56 were synthesised from commercially available 2-methyl-5-methoxyindole in 7 steps (Figure 4 ). Substitution of the 5-position with aziridine and 2-methylaziridine was again succcessful in high yield, as was the czs-2,3-dimethylaziridine. However 2,2-dimethylaziridine reacted with difficulty and the resulting 5-(2,2-dimethylaziridinyl) analogue was unstable in aqueous solution and on silica gel, ring-opening via an S^ 1 mechanism to give 57. The 2-unsubstituted analogues 62-66 were obtained in 6 steps from 5-methoxyindole-3- carboxaldehyde (Figure 5).

NMR spectra were obtained at 90 MHz with a JEOL FX90Q spectrometer using SiMβ4 as internal standard. Elemental analyses were determined by

Butterworth Laboratories Ltd., Teddington, Middlesex, U.K. Solutions in organic solvents were dried by treatment with MgSO4 or Na2SO4 and filtration. Dichloromethane (CH2CI2) was dried over calcium chloride and passed through neutral alumina prior to use. Dimethylformamide (DMF), toluene and tetrahydrofuran (THF) were anhydrous commercial grades. Silica gel for flash column chromatography was

Merck grade (230-400 mesh). Melting points were determined on a Thomas Hoover melting point apparatus and on a Thermogallen Microscope and Hot Stage Apparatus and are uncorrected. Stereochemically pure cw-2,3-dimethylaziridine and 2,2-dimethyl- aziridine were synthesized from the appropriately substituted 2-aminoethanois by O-sulfation and elimination with KOH according to the methods of Dickey et al. (see reference 19). Fused cyclopropamitosenes 4 and 5 were synthesized as described by Cotterill et al. (see reference 1) and O' Sullivan (see reference 20). Starting materials 2-chlorophenol, 5-methoxy-2-methylindole and 5-methoxyindole-3-carboxaldehyde were all purchased from Sigma-Aldrich Chemical Company.

Example 1: Preparation of 2-Cyclopropyl-3-methoxycarbonyl-5-methoxy-l- methylindole-4,7-dione (18) by reactions (v) to (xvii) of Figure 2. (a) Diethyl 5-Methoxy-2-nitrophenylmalonate (6)

A solution of 17 g (0.064 mol) of ethyl 5-methoxy-2-nitrophenylcyanoacetate, prepared in 4 steps from 3-chlorophenyl as described by Speckamp and Oostveen (WO 87/06227), in EtOH ( 100 mL), was saturated with HCl gas on cooling in an ice/salt bath. The solution was stirred for 2 days and then ice-water (100 mL) added and the solution stirred for a further 24 hours at 4°C before the crystalline solid formed was collected by filtration, dried and recrystallized from EtOH to give 17 g (85%) of 6 as a white solid: mp 92-93 °C (Lit. (Reference 4 ) 92-93 °C).

(b) 2-Cyclopropyl-3 -diethoxycarbonyl-2 -dihydro-5-methoxyindole (9).

Compound 6 (5.0 g, 16.1 mmol) was dissolved in a mixture of toluene (62.5 mL) and EtOH (3.75 mL) and reduced with H2 at atmospheric pressure over Ptθ2 (75 mg) catalyst. After 7 hours at room temperature the solution was filtered through Celite and evaporated to dryness BELOW 30 °C, and the resulting pale green oil (7) dissolved immediately in MeOH (75 mL). To this methanolic solution was added cyclopropane carboxaldehyde (1.3 g, 18.6 mmol) dissolved in MeOH (10 mL) and the solution stirred for 15 minutes. The resulting solution of imine 8 was treated in situ with Zn(OAc)2.2H ₂ O (1.13 g, 5.15 mmol) and stirred for 18 hours at room temperature.

The solution was evaporated to dryness at 30 °C, HCl (2.0 M, 50 mL) added and extracted with CH ₂ C1 ₂ (2 x 150 mL), washed with sat. NaHCO ₃ (aq., 150 mL), sat. NaCl (aq., 100 mL), dried and evaporated. The residue was purified on silica, eluting with hexane/EtOAc (1:1, Rf=0.75) to give 9 (3.3 g, 68%) as a yellow oil: ^! H-NMR (CDC1 ₃ ) δ 0.48-0.52 (m, 4H, 2 x cyclopropyl-CH ₂ ), 1.03-1.08 (m, 1Η, cyclopropyl-H), 1-2 (t, 6Η, J=7.2 Hz, CH ₂ CH ₅ ), 1.3 (t, 6Η, J=7.2 Hz, CH ₂ CH ₃ ), 3.75 (s, 3H, CH ₃ O-), 3.85 (d, IH, J=2.7 Hz, 2-H), 4.1-4.35 (m, 4H, CH ₂ CH ₃ ), 6.63 (s, IH, Ar-4H), 6.71 (d, 1Η, J=1.5 Ηz, Ar-6H) and 7.03 (d, 1Η, J=1.5 Ηz, Ar-7H).

(c) l-AcetyI-2-cyclopropyl-3 -diethoxycarbonyl-2 -dihydro-5-methoxyindole (10).

Compound 9 (3.0 g, 10 mmol) was dissolved in Ac ₂ O (5 mL) and stirred for 3 hours at room temperature. The anhydride was then evaporated in vacuo and the residue purified on silica, eluting with hexane/EtOAc (2:1, Rf=0.4) to give a pale yellow oil which was redissoived in Et2θ and evaporated. Trituration of the resulting oil with Et2θ gave a white solid which was collected by filtration and washed with cold Et2θ to give 10 (3.0 g, 87%) as a white solid: mp 104-105°C; iΗ-NMR (CDC1 ₃ ) δ 0.5-0.57 (m, 4Η, 2 x cyclopropyl-CH ₂ ), 1.0-1.15 (m, 1Η, cyclopropyl-H), 1.2 (t, 3Η, J=7.2 Hz, CH ₂ CH ₃ ), 1.3 (t, 3Η, J=7.2 Hz, CH ₂ CH ₃ ), 3.81 (s, 3Η, CH3O-), 4.05-4.3 (m, 4Η, CH ₂ CH ₃ ), 4.5 (d, IH, J=2.7 Hz, 2-H) and 6.9-7.2 (m, 3Η, Ar-4,6,7H).

(d) l-AcetyJ-3-carboxy-2-cyclopropyl-2 -dihydro-5-methoxyindole (11).

The acetylindole 10 (1.0 g, 2.9 mmol) was dissolved in EtOΗ (10 mL) and cooled to 0 ^* C in an ice-salt bath, and a cold (0 ^* C) solution of KOΗ (aq., 10%, 10 mL) was added. The solution was stirred at -5°C for 4 hours and then for 18 hours at 4°C. The solution was poured onto ice/water (25 mL) and washed with Et2θ. The aqueous layer was then acidified with 2.0 M ΗC1 and extracted with CΗCI3 (6 x 50 mL), dried and evaporated to give 0.68 g (97%) of 11 as a yellow oil (Rf=0.45, Me ₂ CO), which was used in the next step without further purification; H-NMR (CDCI3) δ 0.5-0.7 (m, 4H, 2 x cyclopropyl-CH ₂ ), 1.25-1.4 (m, 1Η, cyclopropyl-H), 2.33 (s, 3Η, COCH ₃ ), 3.78 (s, 3Η, CH ₃ O-), 3.85 (br, 1Η, 3-H), 4.5 (br s, 1Η, 2-H), 6.9-7.0 (m, 2Η, Ar-4,6H), 8.05-8.15 (m, 1Η, Ar-7H) and 10.81 (s, 1Η, CO ₂ H).

(e) Methyl l-acetyl-2-cyc.opropy.-23-dihydro-5-methoxyindo.e-3-carboxyl ate (12).

To a solution of 11 (0.68 g, 2.8 mmol) in DMF (10 mL) was added K ₂ CO (0.83 g, 6 mmol) and (MeO) ₂ SO2 (2 g, 15.8 mmol) and the solution stirred at room temperature for 4 hours. The solution was then poured onto HCl (2.0 M, 20 mL) and extracted with CHCI3 (4 x 25 mL), washed with sat. NaCl, dried and evaporated. The residue was purified on silica, eluting with EtOAC/hexane (1 :2, Rf=0.7 (EtOAc)) to give a pale yellow oil of 12 (0.7g, 98%), which solidified on standing; ^-NMR (CDC1 ₃ ) δ 0.58-0.64 (m, 4H, 2 x cyclopropyl-CH ₂ ), 1.25-1.33 (m, 1Η, cyclopropyl-H), 2.33 (s, 3Η, COCH3), 3.68 (s, 3Η, CO ₂ CH ₃ ), 3.79 (s, 3Η, CH3O-), 3.85 (br, 1Η, 2-H), 4.54 (br, 1Η, 2-H), 6.78-6.95 (m, 2Η, Ar-4, 6H) and 8.2 (br s, 1Η, Ar-7H).

(f) Methyl l-acetyl-2-cyclopropyl-5-methoxyindole-3-carboxylate (13).

A solution of 12 (1.0 g, 4 mmol) was stirred under reflux with DDQ (0.96 g, 4.2 mmol) in toluene (12.5 mL) for 7 hours. The DDQΗ2 was removed by filtration and the filtrate evaporated in vacuo. The residue was purified on silica, eluting with hexane EtOAc (1 :1, Rf=0.7) to give 13 (0.88 g, 87%) as a pale yellow oil; ^! H-NMR (CDCl ₃ ) δ 0.73-0.79 (m, 2H, cyclopropyl-CH ₂ ), 1.22-1.3 (m, 2Η, cyclopropyl-CH ₂ ), 2.15-2.25 (m, 1Η, cyclopropyl-H), 2.83 (s, 3Η, COCH ₃ ), 3.86 (s, 3Η, CO ₂ CH ₃ ), 3.97 (s, 3Η, CH ₃ O-), 6.99 (dd, 1Η, J=2.7 and 9 Ηz, Ar-6H), 7.5 (d, 1Η, J=2.7 Ηz, Ar-4H) and 7.96 (d, 1Η, J=9 Ηz, Ax-IH).

(g) Methyl 2-cyclopropyl-5-methoxyindole-3-carboxylate (14).

A solution of 13 (0.88 g, 3.46 mmol) in KOΗ (4% in MeOΗ, 50 mL) was stirred at room temperature for 1.5 hours, neutralized with 6.0M ΗC1 and extracted with EtOAc (3 x 25 mL). The organic layer was washed with Η2O (50 mL), dried and evaporated to give 14 (0.51 g, 70%) as a white solid: mp 128-131 °C; *H-NMR (CDC1 ₃ ) δ 0.82-0.92 (m, 2H, cyclopropyl-CH ₂ ), 1.05-1.2 (m, 2Η, cyclopropyl-CH ₂ ), 2.2-3.0 (m, 1Η, cyclopropyl-H), 3.85 (s, 3Η, CO ₂ CH ₃ ), 3.95 (s, 3Η, CH3O-), 6.79 (dd, 1Η, J=2.7

and 9 Hz, Ar-6H), 7.22 (d, 1Η, J=6.3 Ηz, Ar-7H), 7.58 (d, 1Η, J=2.7 Ηz, Ar-4H) and

8.2 (br s, 1Η, NH).

(h) Methyl 2-cyclopropyI-5-methoxy-l-methylindoIe-3-carboxylate (15).

Compound 14 (9.0 g, 42.6 mmol) was added under argon to a stirred suspension of NaΗ (6.0 g, 0.13 mol) in DMF (150 mL). The solution was heated at 45°C for

0.5 hours, cooled to 0-10°C and Mel (33 mL, 0.23 mol) added. The solution was then gradually heated to 60 °C and stirred at this temperature for 1 hour, cooled and poured onto cold (0°C) NaΗSO ₄ (aq., 10%, 500 mL) and extracted with EtOAc (5 x 75 mL). The organic extracts were washed with sat. NaCl (150 mL), dried and evaporated. The residue was purified on silica, eluting with EtOAc hexane ( 1 :2, Rf=0.45) to give 15 (8.3 g, 86%) as a pale yellow waxy solid; *H-NMR (CDC1 ₃ ) δ 0.77-0.84 (m, 2H, cyclopropyl-CH ₂ ), 1.21-1.29 (m, 2Η, cyclopropyl-CH ₂ ), 1.85-2.05 (m, 1Η, cyclopropyl-H), 3.8 (s, 3Η, CH ₃ N-), 3.86 (s, 3Η, CO ₂ CH ₃ ), 3.9 (s, 3Η, CH ₃ O-), 6.86 (dd, 1Η, J=2.7 and 9 Ηz, Ar-6H), 7.23 (d, 1Η, J=9 Ηz, Ar-7H) and 7.62 (d, 1Η, J=2.7 Ηz, Ar-4H).

(i) Methyl l-methyl-2-cyclopropyl-5-methoxy-4-nitroindole-3-carboxylate (16).

To a solution of 15 (8.0 g, 34.66 mmol) in AcOΗ (150 mL) cooled to 0°C, was added dropwise a cold (0°C) mixture of f.ΗNO ₃ (27 mL) in AcOH (100 mL). The solution was stirred for 3 hours while allowing to reach room temperature, and then poured onto 300 g of crushed ice and after 15 minutes stirring the resulting yellow solid collected by suction filtration. The dried residue was purified on silica, eluting with EtOAc/hexane (1:1, Rf=0.45) to give 7.5 g (71%) of 16 as a yellow solid, recrystallised from EtOAc: mp 129-131 °C; ¹ H-NMR (CDCl ₃ ) δ 0.74-0.81 (m, 2H, cyclopropyl-CH ₂ ) _> 1-22-1.3 (m, 2H, cyclopropyl-CH2) _> 1.85-2.05 (m, IH, cyclopropyl-H), 3.8 (s, 3Η, CH ₃ N-), 3.81 (s, 3Η, CO ₂ CH ₃ ), 3.89 (s, 3Η, CH ₃ O-), 6.94 (d, 1Η, J=9 Ηz, Ar-7H) and 7.32 (d, 1Η, J=9 Ηz, Ar-6H).

(j) Methyl 4-amino-2-cyclopropyl-5-methoxy-l-methylindole-3-carboxylate (17). To a suspension of 2.5 g (9.25 mmol) 16 in EtOH (180 mL) were added tin powder (5.25 g, 44.2 mmol) and HCl (3.0 M, 70 mL) and the solution stirred at room temperature for 1 hour. The solution was then decanted from the excess tin and neutralized with sat. NaHCO ₃ (aq.) and the resulting red suspension added to an equal volume of H ₂ O and extracted with CHC1 ₃ (5 x 50 mL) and then EtOAc (5 x 50 mL) and the combined extracts evaporated. The residue was purified on silica, eluting with EtOAc/hexane (1:1, Rf=0.5) to give 17 (2.2 g, 87%) as a white solid which was used immediately in the next step: mp 49-50 °C; ¹ H-NMR (CDCl ₃ ) δ 0.65-0.72 (m, 2H, cyc.opropyl-CH ₂ ), 1.22-1.3 (m, 2Η, cyclopropyl-CH ₂ ), 1.8-1.9 (m, 1Η, cyclopropyl-H), 3.73 (s, 3Η, CH ₃ N-), 3.85 (s, 3Η, CO ₂ CH ₃ ), 3.9 (s, 3Η, CH ₃ O-), 5.6 (br s, 1Η, NH ₂ ), 6.5 (d, 1Η, J=9 Ηz, Ar-7H) and 6.9 (d, 1Η, J=9 Ηz, Ar-6H).

(k) 2-Cyclopropyl-3-methoxycarbonyl-5-methoxy-l-methylindole-4,7 -dione (18).

To a solution of 17 (2.0 g, 7.2 mmol) in Me ₂ CO (250 mL) was added a solution of potassium nitrosodisulfonate ((KSO ₃ ) ₂ NO, Fremy's salt, 9.7 g, 36.2 mmol)) in

NaΗ ₂ PO ₄ /Na ₂ ΗPO ₄ buffer (250 mL, 0.3 M, pH 6.0) and the solution stirred at room temperature for 1 hour. The Me CO was removed in vacuo and the resulting orange precipitate collected by suction filtration, washed with H ₂ O and dried in a vacuum oven at 45 °C to afford 18 as an orange solid, recrystallized from EtOAc: mp 169-170 °C; ^l H-NMR (CDC1 ₃ ) δ 0.69-0.77 (m, 2H, cyclopropyl-CH ₂ ), 1.1 - 1.2 (m, 2Η, cyclopropyl-CH ₂ ), 1-48-1.58 (m, IH, cyclopropyl-H), 3.80 (s, 3Η, CH ₃ N-), 3.90 (s, 3Η, CO ₂ CH ₃ , 4.01 (s, 3Η, CH ₃ O-) and 5.64 (s, IH, 6-H). Anal. C; 62.06, Η; 5.20, N; 4.71%, QJ _C (C ₁₅ Η ₁₅ NO ₅ ) C; 62.28, H; 5.19, N; 4.84%.

Example 2: 5-rAziridin-l-vn-2-cvclθDroDvl-3-methoxvcarbonvl-l-methvlin dole- 4.7-dione (\9 hv reaction (χyύ' ϊ\ of Figure 2.

Compound 18 (0.29 g, 1 mmol) was dissolved and stirred in freshly redistilled l(H)-aziridine (3 mL, ca.70 mmol, CAUTION!) for 1 hour, evaporated in vacuo and the residue redissoived in EtOAc. The solvent was then partially evaporated until a red precipitate appeared. The red solid was then collected by suction filtration and

recrystallized from EtOAc to afford 19 (0.25 g, 83%) as a red solid: mp 138-140°C; ^! H-NMR (CDC1 ₃ ) δ 0.65-0.72 (m, 2H, cyclopropyl-CH ₂ ), 1.2-1.35 (m, 2Η, cyclopropyl-CH ₂ ), 1.7-1.8 (m, 1Η, cyclopropyl-H), 2.18 (s, 4Η, 2 x aar-CH ₂ ), 3.92 (s, 3Η, CH ₃ N-), 3.99 (s, 3Η, CO ₂ CH ₃ and 5.77 (s, IH, 6-H). Anal. C; 63.60, Η; 5.41, N; 9.33%, Calc. (C ₁₆ Η ₁₆ N ₂ θ4) C; 64.00, H; 5.33, N; 9.33%.

Fxamnle 3: 2-Cvclopropvl-3-hvdroxvmethvl-5-methoxv-1-mcthvliπdnle-4.7- dinne m hv reaction (xix\ of Figure 2.

To a solution of 18 (0.3 g,l .03 mmol) in CHC1 ₃ (30 mL) and EtOH (11 mL) was added a solution of Na2S2θ4 (2.1 g, 12 mmol) in H2O (13 mL). The solution was stirred at room temperature for 0.5 hour and the organic layer separated, washed with sat. NaCl (50 mL), dried and evaporated. The crude hydroquinone was then dissolved in anhydrous CH2C-2 (30 mL) under argon and cooled to -30°C, and DIBAL-H (5 mL of a 1.5 M solution in toluene) added dropwise such that the solution temperature remained BELOW -30°C. The solution was then allowed to reach 0°C and stirred for 2.5 hours at this temperature, and a solution of FeCl ₃ (9 mL, 1.0 M (0.1 M HCl)) added. The solution was stirred for 10 minutes at 0°C and then CHCI3 (150 mL) and H2O (150 mL) added. The aqueous layer was extracted with CHCI3 (5 x 50 mL) then EtOAc (5 x 50 mL) and the combined organic phases washed with sat. NaCl (250 mL), dried and evaporated. The residue was purified on silica, eluting with EtOAc (Rf=0.5) to give 20 as an orange solid after recrystallization from EtOAc (125 mg, 47%): mp 200-202°C; ^l H-NMR (CDCI3) δ 0.71-0.82 (m, 2H, cyclopropyl-CH ₂ ), 1.2-1.33 (m, 2Η, cyclopropyl-CH ₂ )> 1.61-1.71 (m, IH, cyclopropyl-H), 3.81 (s, 3Η, CH3N-), 3.98 (s, 3Η, CH3O-), 4.0 (br s, 1Η, CΗ ₂ OH), 4.69 (br d, 2Η, CH ₂ OH) and 5.64 (s, IH, 6-H). Anal. C; 64.00, Η; 5.66, N; 5.12%, Calc. (C J ₄ ΗJ ₅ O ₄ ); C; 64.36, H; 5.75, N; 5.36%.

Example 4: 5-f Aziridin-1 -vl)-2-cvclopro 1-3-hvdroxvmethvM -methylindnle-4.7- dione (IV. bv reaction (xx of Figure 2.

Compound 20 (0.1 g, 0.38 mmol) was dissolved and stirred in freshly distilled (CAUTION!) l(H)-aziridine (3 mL, ca. 70 mmol) for 0.75 hour, evaporated in vacuo and the residue redissoived in EtOAc, evaporated until a red precipitate appeared and the solid collected. The red solid was recrystallized from EtOAc to give 21 ( 80 mg, 77%): mp 177-179.5°C; H-NMR (CDC1 ₃ ) δ 0.7-0.8 (m, 2H, cyclopropyl-CH ₂ ), 1.22-1.3 (m, 2H, cyclopropyl-CH ₂ ), 1.6-1.7 (m, IH, cyclopropyl-H), 2.18 (s, 4Η, 2 x azir-CH ₂ ), 3.97 (s, 3H, CH3N-), 4.73 (s, 2Η, CH ₂ OH) and 5.77 (s, IH, 6-H). Anal. C; 66.35, Η; 5.61, N; 10.26%, Calc. (C ₁₅ Η _j6 N ₂ θ3) ^{C 66 18} ' ^H ÷ ⁵ - ⁸⁸ ' N; 10.29%.

Example s: 2-Cvclopropvl-3-hvdroxvmethvl-5-f2-methvlaziridin-l-vn-l-met hvl- indole-4.7-dione (22) bv reaction (xx. of Figure 2.

Compound 20 (0.1 g, 0.38 mmol) was dissolved and stirred in freshly distilled 2-methylaziridine (3 mL, ca. 50 mmol) for 2.5 hours, he solution was evaporated in vacuo and the residue redissoived in EtOAc, evaporated until a red precipitate appeared and the solid collected. The red solid was recrystallized from EtOAc to give 22 ( 85 mg, 78%): mp 130-131 °C; *H-NMR (CDCI3) δ 0.68-0.74 (m, 2H, cyclopropyl-CH ₂ ), 1.08-1.2 (m, 2H, cyclopropyl-CH ₂ ), 1.42 (d, 3Η,_J=4.5 Hz, azu--CH ₃ ), 1.5-1.6 (m, 1Η, cyclopropyl-H), 2.01-2.15 (m, 3Η, azir-CHCH ₂ ),

3.97 (s, 3Η, CH3N-), 4.73 (s, 2H, CH ₂ OH) and 5.74 (s, IH, 6-H). Anal. C; 67.12, Η; 6.04, N; 9.57%, Calc. (CJ ₆ ΗJ ₈ N ₂ O ₃ ) C; 67.13, H; 6.29, N; 9.79%.

Example 6: 2-CvcloproPV--5-methoxv-l-methv.-3-ff(phenoxvcarbonvnoxvlmet hvH indole-4.7-dione f23ϊ bv reaction (xxϊ) of Figure 2. To a solution of 20 (0.1 g, 0.38 mmol) in anhydrous pyridine (6 mL) at 0°C, was added dropwise phenylchloroformate (0.1 g, 0.64 mmol) and the solution then allowed to reach room temperature and stirred for 2 hours. The solution was then extracted with CH ₂ C1 ₂ (25 mL) and washed with H ₂ O (25 mL) and sat. NaCl (25 mL), dried and

evaporated. The residue was purified on silica, eluting with EtOAc (Rf=0.75) to give 23 as an orange solid: mp 136-139°C; ^-NMR (CDCI3) δ 1.02-1.18 (m, 2H, cyclopropyl-CH ₂ ), 1.21-1.28 (m, 2Η, cyclopropyl-CH ₂ ), 1.78-1.88 (m, 1Η, cyclopropyl-H), 3.79 (s, 3Η, CH3N-), 4.01 (s, 3Η, CH3O-), 5.27 (s, 2Η, CH ₂ OCOPh), 5.51 (s, 1Η, 6-H) and 7.15-7.3 (m, 5Η, Ar).

Examnle 7: 2-Cvclopropvl-5-methoxv-l-methvl-3-Kcarhamoyloxy^methyll indole- 4 _T 7-dinne (2 \ bv reaction fxxiii of Figure 2.

The phenylcarbonate 23 (0.3 g, 0.78 mmol) was dissolved in anhydrous CH ₂ C1 (38 mL) and the solution cooled to -78 °C. The solution was then saturated with NH3 and stirred at 78 °C until reaction was complete (ca. 2 hours). The solution was then allowed to reach room temperature and evaporated in vacuo. The residue was redissoived in CH ₂ C1 ₂ (100 mL) and washed with H ₂ O (2 x 100 mL) and sat NaCl (50 mL), dried and evaporated, and the residue recrystallized from EtOAc to afford 220 mg (92%) of 24 as an orange solid: mp 240-242°C (dec.); -NMR (CDC1 ₃ ) δ 1.02-1.13 (m, 2H, cyclopropyl-CH ₂ )» 1-22-1.26 (m, 2H, cyclopropyl-CH ₂ ). 1.51-1.55 (m, IH, cyclopropyl-H), 3.78 (s, 3Η, CH ₃ N-), 3.99 (s, 3Η, CH3O-), 4.79 (br s, 2Η, NH ₂ ), 5.3 (s, 2Η, CH ₂ OCONH ₂ ) and 5.62 (s, IH, 6-H). Anal. C; 59.42, Η; 4.88, N; 8.86%, Calc. (C ₅ ΗJ ₆ N ₂ O ₅ ) C; 59.21, H; 4.88, N; 9.21%.

Example 8: 5-f Aziridin-l-vn-2-cvclopropvl-l-methvl-3-[(carbamovloxv^methvl l - indole-4.7-dione f25^ bv reaction (xxiiii of Figure 2.

The carbamate 24 (0.3 g,1.0 mmol) was stirred at room temperature in l(H)-aziridine (2 mL, CAUTION!) for 15 minutes, evaporated and redissoived in EtOAc (5 mL). The solution was then evaporated to 50% volume and the resulting red precipitate filtered and washed well with cold EtOAc to afford 25 (210 mg, 63%) as a red solid: mp 235-238°C(dec); ^-NMR ((CD ₃ ) ₂ SO) δ 0.65-0.73 (m, 2H, cyclopropyl-CH ₂ ), 1.05-1.15 (m, 2Η, cyclopropyl-CH ₂ ), 1.73-1.85 (m, 1Η, cyclopropyl-H), 2.18 (s, 4Η, 2 x azir-CH ₂ ), 3.93 (s, 3Η, CH3N-), 5.06 (s, 2Η, CH ₂ OCONH ₂ ), 5.78 (s, IH, 6-H) and 6.42 (br s, 2Η, NH ₂ ). Anal. C; 60.93, Η; 5.60, N; 13.38%, Calc. (C ₁₆ Η _j7 N ₃ O ₄ ) C; 60.95, H; 5.39, N; 13.33%.

Example 9: 2-Cvclopropvl-l-methvl-5-(2-methvlaziridin-l-vl V (Jcarhamovloxvlmethvll- indole-4.7-dione f26^ bv reaction (xxiH of Figure 2.

The carbamate 24 (0.05 g, 0.164 mmol) was stirred at room temperature in 2-methylaziridine (1.5 mL) for 4 hours, evaporated and redissoived in EtOAc (5 mL). The solution was then evaporated to 50% volume and the resulting red precipitate filtered and washed well with cold EtOAc, and then recrystallized from EtOAc to afford 26 (30 mg, 56%) as a red solid: mp 209-21 l °C(dec); ¹ H-NMR ((CD ₃ ) ₂ SO) δ 0.63-0.72 (m, 2H, cyclopropyl-CH ₂ ), 1.01-1.09 (m, 2Η, cyclopropyl-CH ₂ ), 1.29 (d, 3Η, J=5.4Hz, azir-CH ₃ ), 1.75-1.85 (m, 1Η, cyclopropyl-H), 1.98-2.05 (m, 3Η, azir-CHCH ₂ ), 3.93 (s, 3Η, CH ₃ N-), 5.06 (s, 2Η, CH ₂ OCONH ₂ ), 5.76 (s, IH, 6-H) and 6.41 (br s, 2Η, NH ₂ ). Anal. C; 61.76, Η; 5.41, N; 12.24%, Calc. (C ₁₇ Η ₁₉ N ₃ O ₄ ) C; 62.00, H; 5.77, N; 12.76%.

Example 10: Preparation of 2-Isopropvl-3-methoxvcarbonvl-5-methoxv-l- methvlindole-4.7-dione (35i bv reactions rix xvin of Figure 2. (a) l-Acetyl-3 -diethoxycarbonyl-2 -dihydro-2-isopropyI-5-methoryindole (27). This compound was prepared (56%) by the method described for compound 9 but using isobutyraldehyde in place of cyclopropane carboxaldehyde. The residue after work-up was dissolved in Ac ₂ O (5 mL) and stirred for 3 hours at room temperature. The anhydride was then evaporated in vacuo and the residue purified on silica, eluting with hexane/EtOAc (1:1, Rf=0.5) to give 27 (87%) as a white solid: mp 77.5-78.5°C; *H-NMR (CDC1 ₃ ) δ 0.6 (d, 3H, J=7.2 Hz, CHCH ₃ ), 0.9 (d, 3Η, J=7.2 Hz, CHCH ₃ ), 1.2 (t, 6Η, J=7.2 Hz, CH ₂ CH ₃ ), 1.3 (t, 6Η, J=7.2 Hz, CH ₂ CH ₃ ), 2.12-2.28 (m, 1Η, CH(CΗ ₃ ) ₂ ), 2.36 (s, 3H, COCH ₃ ), 3.81 (s, 3Η, CH ₃ O-), 4.1-4.33 (m, 4Η, 2 x CH ₂ CH ₃ ), 4.8 (br, IH, 2-H) and 6.9-7.8 (m, 3Η, Ar-4,6,7H).

(b) l-Acetyl-3-carboxy-2 -dihydro-2-isopropyl-5-methoxyindole (28).

The acetylindole 27 was hydrolysed as described for the preparation of 11 to give 28 (90%) as an off-white foam (Rf=0.45, Me ₂ CO), which was used in the next step without further purification; ^! Η-NMR (CDC1 ₃ ) δ 0.59 (d, 3H, J=7.2 Hz, CHCH ₃ ), 0.9 (d, 3Η,

J=7.2 Hz, CHCH ₃ ), 2.15-2.22 (m, 1Η, CH(CΗ _{3 2} ), 2.2 (s, 3H, COCH3), 3.72 (s, 3Η, CH3O-), 3.98 (br, 1Η, 3-H), 4.62 (br, 1Η, 2-H), 6.8-6.98 (m, 2Η, Ar-4,7H) and 7.78-7.85 (m, 1Η, Ar-6H).

(c) Methyl l-acetyl-2 -dihydro-2-isopropyI-5-methoxyindole-3-carboχylate (29). In a procedure identical to that carried out on 11, compound 29 was prepared from compound 28 as a pale brown oil (93%); ^-NMR (CDCI3) δ 0.71 (d, 3Η, J=7.2 Hz, CHCH ₃ ), 1.0 (d, 3H, J=7.2 Hz, CHCH3), 2.1-2.2 (m, IH, CH(CH ₃ )2), 2.32 (s, 3H, COCH3), 3.69 (s, 3H, CO ₂ CH ₃ ), 3.79 (s, 3H, CH ₃ O-), 3.8 (br , 1Η, 3-H), 4.6 (br, 1Η, 2-H), 6.8-6.98 (m, 2Η, Ar-4, IH) and 7.78-7.85 (m, IH, Ar-6H).

(d) Methyl l-acetyl-2-isopropyl-5-methoxyindoIe-3-carboxyiate (30).

A solution of 29 (1.0 g, 4.0 mmol) was stirred under reflux with DDQ (0.96 g, 4.2 mmol) in toluene (12.5 mL) for 2 days. The DDQΗ2 was removed by filtration and the filtrate evaporated in vacuo. The residue was purified on silica, eluting with hexane/Me ₂ CO (3:1, Rf=0.65) to give 30 (0.44 g, 44%) as a pale red oil; ^! H-NMR (CDC1 ₃ ) δ 1.45 (d, 6H, J-7.2 Hz, CH(CH ₃ ) ₂ ), 2.78 (s, 3Η, COCH3), 3.87 (s, 3H, CO ₂ CH ₃ ), 3.96 (s, 3Η, CH3O-), 3.88-3.98 (m, 1Η, CH(CΗ ₃ ) ₂ ), 6.89 (dd, IH, J=2.7 and 9 Hz, Ar-6H), 7.44 (d, 1Η, J=9 Ηz, Ar-7H) and 7.55 (d, 1Η, J=2.7 Ηz, Ar-4H).

(e) Methyl 2-isopropyl-5-methoxyindo.e-3-carboxy.ate (31).

This compound was prepared from compound 30 as described for 14 (70%) as a pale red oil; -NMR (CDC1 ₃ ) δ 1.43 (d, 6Η, J=7.2 Hz, CH(CH ₃ ) ₂ ), 3.87 (s, 3Η,

CO ₂ CH ₃ ), 3.92 (s, 3Η, CH ₃ O-), 4.08-4.25 (m, 1Η, CH(CΗ ₃ ) ₂ ), 6.9 (dd, 2H, J=2.7 and 9 Hz, Ar-6H), 7.1-7.6 (m, 2Η, Ar-4,7H) and 8.2 (br s, 1Η, NH).

(f) Methyl 2-isopropyl-5-methoxy-l-methylindoIe-3-carboxylate (32).

In a procedure identical to that carried out on 14, compound 32 was prepared (80%) from compound 31 and purified on silica, eluting with EtOAc/hexane (1:2, Rf=0.74) as a pale yellow solid: mp 105-106°C; *Η-NMR (CDC1 ₃ )

δ 1.45 (d, 6H, J=7.2Hz, CH(CH ₃ ) ₂ ), 3.79 (s, 3H, CH ₃ N-), 3.88 (s, 3Η, CO ₂ CH ₃ ), 3.92 (s, 3Η, CH3O-), 4.22-4.45 (m, IH, CH(CH ₃ ) ₂ ), 6.9 (dd, 2H, J=2.7 and 9Hz, Ar-6H) and 7.1-7.64 (m, 1Η, Ar-4,7H).

(g) Methyl 2-isopropyl-5-methoxy-l-methyl-4-nitroindole-3-carboxylate (33). To a solution of 32 (8.0 g, 34.66 mmol) in AcOΗ (150 mL) cooled to 0°C, was added dropwise a cold (0°C) rnixture of f.ΗNO ₃ (27 mL) in AcOH (100 mL). The solution was stirred for 3 hours while allowing to reach room temperature, and then poured onto 300 g of crushed ice and the resulting yellow solid collected by suction filtration. The dried residue was purified on silica, eluting with EtOAc/hexane (1 :2, Rf=0.26) to give 7.5 g (71%) of 33 as a yellow solid, recrystallised from EtOAc: mp 149-150.5°C; *H-NMR (CDC1 ₃ ) δ 1.44 (d, 6H, J=7.2 Hz, CH(CH ₃ ) ₂ ), 3.78 (s, 3Η, CH3N-), 3.82 (s, 3Η, CO ₂ CH ₃ ), 3.91 (s, 3Η, CH ₃ O-), 3.95-4.15 (m, 1Η, CH(CΗ ₃ ) ₂ ), 6.97 (d, IH, J=9 Hz, Ar-6H) and 7.35 (d, 1Η, J=9 Ηz, Ar-7H).

(h) 2-IsopropyI-3-methoιycarbonyl-5-methoxy-l-methylindole-4,7- dιone (35). Compound 33 was reduced as described for 17 to give 34 (85%) and the crude material oxidized with Fremy's salt as described for 18. After work-up the resulting orange precipitate was collected by suction filtration, washed with Η ₂ O and dried in a vacuum oven at 45 °C to afford 35 as an orange solid (75%), recrystallized from EtOAc: mp 192-194°C; ^-NMR (CDC1 ₃ ) δ 1.34 (d, 6H, J=7.2 Hz, CH(CH ₃ ) ₂ ), 3.12-3.28 (m, 1Η, CH(CΗ ₃ ) ₂ ), 3.81 (s, 3H, CH3N-), 3.90 (s, 3H, CO ₂ CH ₃ ), 3.97 (s, 3H, CH3O-) and 5.64 (s, 1Η, 6-Η). Anal. C; 62.31, H; 5.82, N; 4.90% Calc. (C ₁₅ H ₁₇ NO ₅ ) C; 61.86, H; 5.84, N; 4.81%.

Example 11 : 3-Hvdroxvmethvl-2-isopropvl-5-methoxy-l-methylindole-4.7-dio ne (36Ϊ bv reaction (xviifl of Figure 2. To a solution of 35 (0.5 g, 1.7 mmol) in CHCI3 (50 mL) and EtOH (18 mL) was added a solution of Na ₂ S ₂ O ₄ (3.5 g, 20 mmol) in H O (22 mL). The solution was stirred at room temperature for lhour and the organic layer separated, washed with sat. NaCl

(50 mL), dried and evaporated. The crude hydroquinone was dried over 18 hours in vacuo and then dissolved in anhydrous THF (5 mL) under argon and added to a solution of L-AIH4 (12 mL of a 1.0 M solution inTHF) dropwise at room temperature and under argon. The solution was then stirred for 1 hour at 30 °C, cooled to 0°C, and H O ( 15 mL) added dropwise, followed by a solution of FeCl ₃ ( 12 mL, 1.0 M (0.1 M HCl)) added at 0°C. The solution was stirred for 10 minutes at 0°C and then EtOAc (150 mL) and H ₂ O (150 mL) added. The aqueous layer was extracted with EtOAc (5 x 50 mL) and the organic phase washed with sat. NaCl (250 mL), dried and evaporated. The residue was purified on silica, eluting with EtOAc/hexane, (1:1, Rf=0.22) to give, after recrystallization from EtOAc, 36 as an orange solid (140 mg, 31 %): mp 160-161°C; ^-NMR (CDCI3) δ 1.37 (d, 6H, J=7.2 Hz, CH(CH ₃ ) ₂ ), 3.1-3.28 (m, 1Η, CH(CΗ _{3 2} ), 3.82 (s, 3H, CH3N-), 3.97 (s, 3Η, CH ₃ O-), 4.71 (br, 2Η, CH ₂ OH) and 5.64 (s, IH, 6-H). Anal. C; 63.42, Η; 6.50, N; 5.24% Calc. (C ₁₄ Η ₁₇ NO ₄ ) C; 63.88, H; 6.46, N; 5.32%.

Example 12: 5-f Aziridin-l-vlV-3-hvdroxvmethvl-2-isopropvl-l-methvlindole-4. 7- ipne (37) py reac ion fox) pf Figure 2,

A solution of 36 (50 mg, 0.19 mmol) in l(H)-aziridine (0.5 mL, ca. 11.7 mmol, CAUTION!) was stirred for 0.5 hour at room temperature and then evaporated to dryness and the residue purified on silica, eluting with EtOAc (Rf=0.55) to give, after recrystallization from EtAOc, 37 (42 mg, 81%)) as a red solid: mp 144-145°C;

^! H-NMR (CDC1 ₃ ) δ 1.37 (d, 6H, J=7.2 Hz, CH(CH ₃ ) ₂ ), 2.18 (s, 4Η, 2 x azir-CH ₂ , 3.1-3.28 (m, 1Η, CH(CΗ ₃ ) ), 3.95 (s, 3H, CH ₃ N-), 4.68 (br s, IH, CH ₂ OH), 4.76 (br, 2H, CH ₂ OH) and 5.76 (s, IH, 6-H). Anal. C; 65.74, Η; 6.64, N; 9.91% Calc. (C ₁₅ Η ₁₈ N ₂ O ₃ ) C; 65.69, H; 6.57, N; 10.22%.

Example 13: 3-Hvdroxvmethvl-2-isopropvl-5-(2-methvtaziridin-1 -vH-l- methvlindole-4.7-dione (38 bv reaction (xxi of Figure 2.

Compound 36 (0.1 g, 0.38 mmol) was dissolved and stirred in freshly distilled 2-methylaziridine (3 mL, ca .50 mmol) for 2.5 hours. The solution was evaporated

in vacuo and the residue redissoived in EtOAc, evaporated and purified on silica (eluting with EtOAc) to afford a red glass (38, 85 mg, 78%): ^l H-NMR (CDC1 ₃ ) δ 1.37 (d, 6H, J=7.2 Hz, CH(CH ₃ ) ₂ ), 1.42 (d, 3H, J=4.5 Hz, azir-CH ₃ ), 2.01-2.15 (m, 3Η, azir-CHCH ₂ ), 3.1-3.28 (m, 1Η, CH(CΗ ₃ ) ₂ ), 3.95 (s, 3H, CH ₃ N-), 4.68 (br s, 1Η, CΗ ₂ OH), 4.76 (br, 2Η, CH ₂ OH) and 5.76 (s, IH, 6-H). Anal. C; 67.12, Η; 6.04, N; 9.57% Calc. (C _j6 Η ₂₀ N ₂ O ₃ ) C; 66.67, H; 6.94, N; 9.72%.

Example 14: Preparation of 2-isopropvl-5-methoxv-l-methvl-3-l(carbamovloxvϊ- mgrhvl)-inrlole-4.7-dione (40 bv reactions (xxϊ\ and (xxif) of Figure 2. (a)2-Isopropyl-5-methoxy-l-methyl-3-[[(phenoxycarbony.)oxy]m ethyl] indole-4,7- dione (39).

To a solution of 36 (0.55 g, 2.1 mmol) in anhydrous pyridine (25 mL) at 0 ^C C, was added dropwise phenylchloroformate (0.5 g, 3.2 mmol) and the solution then allowed to reach room temperature and stirred for lhour and then a further 0.25 g (1.6 mmoL) of phenylchloroformate was added. After a further 1 hour, H ₂ O (200 mL) was added and the solution was extracted with EtOAc (4 x 50 mL) and washed with H O (100 mL) and sat. NaCl (100 mL), dried and evaporated. The residue was purified on silica, eluting with EtOAc (Rf ).8) to give 39 (0.6 g, 75%) as an orange solid: mp 126-127°C; !H-NMR (CDC1 ₃ ) δ 1.37 (d, 6H, J=7.2 Hz, CH(CH ₃ ) ₂ ), 3.1-3.28 (m, 1Η, CH(CΗ ₃ ) ₂ ), 3.79 (s, 3H, CH3N-), 4.01 (s, 3Η, CH3O-), 5.27 (s, 2Η, CH ₂ OCOPh), 5.51 (s, 1Η, 6-H) and 7.15-7.3 (m, 5Η, Ar).

(b) 2-Isopropyl-5-methoxy-l-methyl-3-[(carbamoyloxy)methyl] indole-4,7-dione (40).

The phenylcarbonate 39 (0.1 g, 0.26 mmol) was dissolved in anhydrous CH2C-2 (15 mL) and the solution cooled to -78 °C. The solution was then saturated with NH ₃ and stirred at -78°C until reaction was complete (ca. 2.5 hours). The solution was then allowed to reach room temperature and evaporated in vacuo. The residue was redissoived in CH ₂ C1 (50 mL) and washed with H ₂ O (2 x 50 mL) and sat. NaCl (25 mL), dried and evaporated, and the residue recrystallized from EtOAc to afford

55 mg (69%) of 40 as an orange solid: mp 244-246°C (dec); ^-NMR CCDC δ 1.37 (d, 6H, J=7.2 Hz, CH(CH ₃ ) ₂ ), 3.1-3.28 (m, IH, CH(CH ₃ ) ₂ ), 3.78 (s, 3H, CH ₃ N-), 3.99 (s, 3Η, CH _j O-), 4.79 (br s, 2Η, NH ₂ ), 5.3 (s, 2Η, CH ₂ OCONH ₂ ) and 5.62 (s, IH, 6-H). Anal. C; 58.67, Η; 5.76, N; 9.02%, Calc. (C ₁₅ Η ₁₈ N ₂ O ₅ ) C; 58.82, H; 5.88, N; 9.15%.

Example 15: 5-(Aziridin-l-vlV2-isopropvl-l-methvl-3- rcarbamoyloxylmethyl] indolβ-4.7-dione (411 bv reaction (xxiiil of Figure 2.

The carbamate 40 (0.1 g, 0.33 mmol) was stirred at room temperature in l(H)-aziridine (2 mL, CAUTION!) for 25 minutes, evaporated and redissoived in EtOAc (5 mL). The solution was then evaporated to 50% volume and the resulting red precipitate filtered and washed well with cold EtOAc to afford 41 (55 mg, 53%) as a red solid: mp 230-233 °C(dec); ^! H-NMR ((CD ₃ ) ₂ SO) δ 1.37 (d, 6H, J=7.2 Hz, CH(CH ₃ ) ₂ ), 2.18 (s, 4Η, 2 x aar-CH ₂ ), 3.1-3.28 (m, IH, CH(CH ₃ ) ₂ ), 3.93 (s, 3H, CH ₃ N-), 5.06 (s, 2Η, CH ₂ OCONH ₂ ), 5.78 (s, IH, 6-H) and 6.42 (br s, 2Η, NH ₂ ). Anal. C; 60.25, Η; 6.04, N; 13.30%, Calc. (C ₁₆ Η ₁₉ N ₃ O ₄ ) C; 60.57, H; 5.99, N; 13.25%.

Example 16: 2-Isopropvl-l-methvl-5-(2-methvlaziridin-l-vll-3- [(carbamovloxvlmethvll indole-4.7-dione (421 hv reaction (xxiiil of Figure 2.

The carbamate 40 (0.1 g, 0.32 mmol) was stirred at room temperature in 2-methylaziridine (1.5 mL) for 2.5 hours, evaporated and redissoived in EtOAc (5 mL). The solution was then evaporated to 50% volume and the resulting red precipitate filtered and washed well with cold EtOAc, and then recrystallized from EtOAc to afford 42 (55 mg, 52%) as a red solid: mp 204-205 °C(dec); ^! H-NMR ((CD ₃ ) ₂ SO) δ 1.37 (d, 6H, J=7.2 Hz, CH(CH ₃ ) ₂ ), 1.42 (d, 3Η, J=5.4 Hz, azir-CH ₃ ), 1.98-2.05 (m, 3Η, azir-CHCH ₂ ), 3.1-3.28 (m, 1Η, CH(CΗ ₃ ) ₂ ), 3.93 (s, 3H, CH ₃ N-), 5.06 (s, 2Η, CH ₂ OCONH ₂ ), 5.76 (s, IH, 6-H) and 6.41 (br s, 2Η, NH ₂ ). Anal. C; 61.52, Η; 6.36, N; 12.90%, Calc. (C _j7 Η2 _j N ₃ O ₄ ) C; 61.63, H; 6.34, N; 12.69%.

Example 17: 1.3-Dimethvl-2--8opropvl-5-methoxvindole-4.7-dione (431.

To a solution of 35 (0.5 g, 1.7 mmol) in CHC1 ₃ (50 mL) and EtOH (18 mL) was added a solution of Na2S2θ4 (3.5 g, 20 mmol) in H2O (22 mL). The solution was stirred at room temperature for lhour and the organic layer separated, washed with sat. NaCl (50 mL), dried and evaporated. The crude hydroquinone was dried over 18 hours in vacuo and then dissolved in anhydrous THF (5 mL) under argon and added to a solution of DIBAL-H (10 mL of a 1.5 M solution in toluene) dropwise at -30°C and under argon. The solution was then stirred for 18 hours at 4°C, cooled to 0°C, and H ₂ O (15 mL) added dropwise, followed by a solution of FeCl ₃ (12 mL, 1.0 M (0.1 M HCl)) added at 0°C. The solution was stirred for 10 minutes at 0°C and then EtOAc (150 mL) and H2O (150 mL) added. The aqueous layer was extracted with EtOAc (5 x 50 mL) and the organic phase washed with sat. NaCl (250 mL), dried and evaporated. The residue was purified on silica, eluting with EtOAc/hexane, (1: 2, Rf=0.44) to give, after recrystallization from EtOAc, 43 as an orange solid (48 mg, 11 %): mp 168-169°C; ^-NMR (CDCI3) δ 1.35 (d, 6H, J=7.2 Hz, CH(CH ₃ ) ₂ ), 2.38 (s, 3Η, 3-CH ₃ ), 3.1-3.28 (m, 1Η, CH(CΗ ₃ ) ₂ ), 3.78 (s, 3H, CH3N-), 3.95 (s, 3H, CH3O-) and 5.57 (s, 1Η, 6-H).

Example 18: 5-(Aziridin-l-vll-13-dimethvl-2-isopropvlindole-4.7-dione (441.

A solution of 43 (30 mg, 0.12 mmol) in l(Η)-aziridine (0.5 mL, ca. 11.7 mmol, CAUTION!) was stirred for 0.5 hour at room temperature and then evaporated to dryness and the residue purified on silica, eluting with EtOAc/hexane (1:2, Rf=0.32) to give, after recrystallization from EtAOc, 44 (5.5 mg, 18%)) as a red solid: mp 110-112°C; ^! H-NMR (CDCI3) δ 1.35 (d, 6H, J=7.2 Hz, CH(CH ₃ ) ₂ ), 2.15 (s, 4Η, 2 x azir-CH ₂ ), 2.38 (s, 3Η, 3-CH ₃ ), 3.1-3.28 (m, 1Η, CH(CΗ ₃ ) ₂ ), 3.94 (s, 3H, CH ₃ N-) and 5.72 (s, 1Η, 6-H).

Example 19: 2-Cvclohexvl-3-hvdroxvmethvl-5-methoxv-l-methvlindole-4.7-di one s

The corresponding 3-methoxycarbonyl precursor compound was prepared as described for 18 and 35 but using cyclohexylcarboxaldehyde in the in-line cyclisation step and the 5 subsequent oxidation step with DDQ required a much prolonged reaction time of 12h. The crude 4-amino compound was again oxidized with Fremy's salt as described for 18. After work-up the resulting orange precipitate was collected by suction filtration, washed with H20 and dried in a vacuum oven at 45°C to give 2-cyclohexyl-3- methoxycarbonyl-5-methoxy-l-methylindole-4,7-dione as an orange solid (75%),

10 recrystallized from EtOAc: mp 190-192°C; ^-NMR (CDC1 ₃ ) 8 1.27 (m, 6H, 6 x cylohexyl-H), 1.83 (m, 5H, 5 x cyclohexyl-H), 3.79 (s, 3H, CH ₃ N-), 3.91 (s, 3H, CO ₂ CH ₃ ), 3.96 (s, 3H, CH ₃ O-) and 5.63 (s, IH, 6-H). This compound was then reduced with LiAIH4 as described for 36 to give 45 as an orange solid (59 %): mp 214-215°C; H-NMR (CDC1 ₃ ) & 1.25 (m, 6H, 6 x cylohexyl-H), 1.78 (m, 5H,

15 5 x cyclohexyl-H), 3.81 (s, 3H, CH ₃ N-), 3.97 (s, 3H, CH ₃ O-), 4.76 (s, 2H, CH ₂ OH) and 5.63 (s, IH, 6-H). Anal. C; 67.54, H; 6.82, N; 4.67% Calc. (C ₁₇ H ₂₁ NO4) C; 67.31, H; 6.98, N; 4.62%.

Example 20: 5-(Aziridin-l-vlV2-cvclohexvl-3-hvdroxvmethvl-l-methvlindole -4.7- ione (46).

20 A solution of 45 (30mg,0. lmmol) infreshly distilled l(H)-azrridine (0.5mL, ca. 11.7mmol, CAUTION !) was stirred for 0.5h at room temperature and then evaporated to dryness and the residue purified on silica, eluting with EtOAc/hexane.

( 1 : (1:1, Rf=0.3) to a give, after recrystallization from EtAOc, 46 (22mg, 70%)) as a red solid: mp 128-130°C; *H-NMR (CDC1 ₃ ) 8 1.26 (m, 6H, 6 x cylohexyl-H),

25 1.78 (m, 5H, 5 x cyclohexyl-H), 2.18 (s, 4H, 2 x aziridine-CH ₂ ), 3.95 (s, 3Η, CH ₃ N-), 4.76 (s, 2Η, CH2OH) and 5.77 (s, IH, 6-H). Anal. C; 63.72, H; 6.97, N; 8.22% Calc. (C ₁₈ H ₂₂ N ₂ 0 ₃ .1.5H ₂ O)C; 63.34, H; 7.33, N; 8.21%.

Example 21 Preparation of 5-Methoxv-3-hvdroxvmethvl-l.2-dimethvlindole-4.7- dione (521 hv reactions (il to (vii of Figure 3.

(a) 5-Methoxy-l,2-dimethyIindole (47).

5-Methoxy-2-methylindole (10 g, 0.062 mol) was added gradually and under dry argon to a stirred suspension of NaH (2.73 g of a 60% dispersion, 0.068 mol) in DMF (150 mL). The suspension was heated at 45°C for 10 minutes, cooled to room temperature, and Mel (33 mL, 0.23 mol) added over 5 minutes. The solution was then heated at 60 °C for 1 hour, cooled and poured onto cold (0°C) NaHSO (aq., 10%, 150 mL) and extracted with EtOAc (3 x 100 mL), dried and evaporated. The residue was purified on silica, eluting with 3% EtOAc/hexane (Rf=0.5) to give 4.5 g (41%) of 45 as a pale brown solid: mp 73-74°C; -NMR CCDCty δ 2.37 (s, 3H, 2-CH ₃ ), 3.59 (s, 3Η, CH ₃ N-), 3.82 (s, 3Η, CH ₃ O-), 6.16 (s, 1Η, 3-H) and 6.9-7.28 (m, 3Η, Ar-4,6,7H).

(b) 5-Methoxy-l,2-dimethylindole-3-carboxaldehyde (48). N-methylformanilide (0.95 g, 7.04 mmol) and POCI3 (1.08 g, 7.05 mmol) were stirred at room temperature until the yellow solid chloroimmonium Vilsmeier compound formed. The yellow solid was then added to a solution of 47 (0.7 g, 4 mmol) in 1,2-dichloroethane (15 mL) and the solution heated under reflux for 1.5 hours, cooled, and NaOAc (1.0 M, 50 mL) added. The solution was extracted with EtOAc (4 x 100 mL), dried and evaporated. The residue was purified on silica, eluting with

EtOAc/hexane (1:1, Rf=0.5 (EtOAc)) to give 0.35 g (43%) of 48 as an off-white solid: mp 108-110°C; ^-NMR (CDCI3) δ 2.62 (s, 3Η, 2-CH ₃ ), 3.63 (s, 3Η, CH3N-), 3.89 (s, 3Η, CH ₃ O), 6.94-7.2 (m, 2Η, Ar-6,7H), 7.8 (d, IH, J=2 Hz, Ar-4H) and 10.1 (s, 1Η, CHO).

(c) 5-Methoxy-l,2-dimethyl-4-nitroindoIe-3-carboxaldehyde (49) by reaction (iii).

Compound 48 (6.0 g, 0.03 mol) was dissolved in AcOΗ (480 mL) and cooled to 5°C and f.ΗNO ₃ (18 mL) in AcOH (72 mL) added dropwise with stirring over 5 minutes. The solution temperature was allowed to rise to 20°C over 18 hours and the mixture

poured on to crushed ice (500 g) and the yellow precipitate collected by suction filtration and dried in vacuo at 50 °C. The yellow solid was purified on silica, eluting with EtOAc hexane (2:1, Rf=0.42 (EtOAc)) to give 4.37 g (59%) of 49 as a pale yellow solid: mp 236-238°C(dec); ^-NMR ((CD ₃ ) ₂ SO) δ 2.71 (s, 3H, 2-CH ₃ ), 3.76 (s, 3Η, CH ₃ N-), 3.89 (s, 3Η, CH ₃ O), 7.25 (d, 1Η, J=9 Ηz, Ar-7Η), 7.75 (d, IH, J=9 Hz, Ar-6H) and 9.9 (s, 1Η, CH0).

(d) 4-Amino-5-methoxy-l,2-dimethyIindo-e-3-carboxaldehyde (50) by reaction (iv).

Nitro-compound 49 (0.18 g, 0.74 mmol) was dissolved in EtOΗ (15 mL) and powdered tin (0.45 g, 3.8 mmol) added, followed by ΗC1 (3.0 M, 5.6 mL) and the solution heated under gentle reflux for 1 hour. Water (50 mL) was added and the solution neutralized with NaΗCO ₃ (aq.) and then extracted with CHC1 ₃ (3 x 100 mL), dried and evaporated. The residue was purified on silica, eluting with EtOAc/hexane (1:1, Rf=0.57 (EtOAc)) to give 0.1 g (62%) of 50 as a pale yellow solid: mp 152-153°C (dec.); ^-NMR ((CD ₃ ) ₂ SO) δ 2.61 (s, 3H, 2-CH ₃ ), 3.59 (s, 3H, CH3N-), 3.75 (s, 3Η, CH3O), 6.0 (br s, 2Η, NH ₂ ), 6.56 (d, 1Η, J=9 Ηz, Ar-7Η), 6.85 (d, IH, J=9 Hz, Ar-6H) and 9.71 (s, 1Η, CHO).

(e) 5-Methoxy-l,2-dimethyI-4,7-dioxo-ndoIe-3-carboxaIdehyde (51).

To a solution of 50 (0.074 g, 0.34 mmol) in Me^O (14 mL) was added a solution of Fremy's salt (0.45 g, 1.68 mmol) in NaΗ ₂ PO ₄ Na ₂ ΗPO ₄ buffer (14 mL, 0.3 M, pH 6.0) and the solution stirred at room temperature for 1 hour. The solution was evaporated at 30°C to remove most of the Me CO and the resulting orange precipitate collected by suction filtration and washed well with H ₂ O and cold MeOH, to give 51 (0.06 g, 75%): mp 239-242°C; ¹ H-NMR ((CD ₃ ) ₂ SO) δ 2.5 (s, 3H, 2-CH ₃ ), 3.82 (s, 3Η, CH ₃ N-), 3.88 (s, 3Η, CH ₃ O), 5.89 (s, 1Η, 6-H) and 10.37 (s, 1Η, CHO).

(f) 5-Methoxy-3-hydroxymethyl-l,2-dimethylindole-4,7-dione (52) by reaction (vi). To a suspension of 51 (0.25 g, 0.94 mmol) in MeOΗ (100 mL, degassed by boiling in under argon in vacuo) was added NaBΗ4 (0.36 g, 9.7 mmol) while maintaining a dry

argon atmosphere. The solution was stirred at room temperature for 1 hour and the solution aerated prior to the addition of H ₂ O (40 mL) and the solution extracted with CH ₂ C-2 (3 x 100 mL), dried and evaporated. The residue was purified on silica, eluting with EtOAc (Rf=0.4) to afford 52 (0.1 g, 33%) as an orange solid, recrystallized from EtOAc: mp 215-216°C; *H-NMR (CDC1 ₃ ) δ 2.21 (s, 3H, 2-CH ₃ ), 3.81 (s, 3H,

CH ₃ N-), 3.86 (s, 3Η, CH ₃ O), 4.65 (br d, 2Η, J=7.2 Hz, CH ₂ OH) and 5.61 (s, IH, 6-H)

Example 22: 5- ⁽ Aziridin-l-vll-3-hvdroxvmethvl-1.2-dimethylindole-4.7- dione (531 via reaction (viii of Figure 4.

A solution of 52 (235 mg, 1.0 mmol) in freshly redistilled l(Η)-aziridine (1.5 mL, ca. 35 mmol, CAUTION!) was stirred for 0.5 hour at room temperature and evaporated in vacuo. The residue was redissoived in EtOAc, condensed by 75%, and the precipitate collected and washed with cold EtOAc, to give 225 mg ( 91%) of 53 as a dark red solid: mp 173-174°C (dec); ^-NMR (CDC1 ₃ ) δ 2.19 (s, 4H, 2 x azrr-CH ₂ ), 2.22 (s, 3H, 2-CH ₃ ), 3.86 (s, 3Η, CH ₃ N-), 3.95 (br, 1Η, CΗ ₂ OH), 4.65 (br d, 2Η, CH ₂ OH) and 5.76 (s, IH, 6-H). Anal. C; 63.08, Η; 5.62, N; 11.07% Calc (C ₁₃ Η _j 4N ₂ O ₃ ) C; 63.41, H; 5.69, N; 11.38%.

Example 23 : 3-Hvdroxvmethvl-5-(2-methvlaziridin-1-vll-1.2-dimethvlindole -4.7- dinnβ (541 hv reaction (viii of Figure 4.

A solution of 52 (235 mg, 1.0 mmol) in 2-methylaziridine (2 mL, ca. mmol) was stirred at room temperature for 4 hours and worked up as described for 53 to give 54 (195 mg, 75%) as a dark red solid: mp 120-122°C; *H-NMR (CDCI3) δ 1.42 (d, 3H, J=5.4 Hz, azir-CH ₃ ), 2.08-2.21 (m, 3Η, azir-CHCH ₂ ), 2.22 (s, 3Η, 2-CH ₃ ), 3.89 (s, 3Η, CH ₃ N-), 4.61 (br, 1Η, CΗ ₂ OH), 4.63 (br, 2Η, CH ₂ OH) and 5.75 (s, IH, 6-H). Anal. C; 65.09, Η; 5.79, N; 10.72% Calc. (C ₁₄ Η ₁₆ N ₂ O ₃ ) C; 64.61, H; 6.15, N; 10.77%.

Example 24: 3-Hvdroxvmethvl-5-(cft-23-dimethvlaziridin-1-vll-1.2- dimethvlindole-4.7-d.one (551 bv reaction (viii of Figure 4.

A solution of 52 (100 mg, 0.43 mmol) in cis-2,3-dimethylaziridine (2 mL) was stirred for 1 hour at room temperature and then evaporated in vacuo at 30 °C. The residue was purified on silica, eluting with EtOAc (Rf=0.6), to give 55 (65 mg, 55%) after recrystallization from EtOAc: mp 132-135°C; *H-NMR (CDC1 ₃ ) δ 1.38 (d, 6H, J=5.4Hz, 2 x azir-CH ₃ ), 2.15-2.21 (m, 2Η, 2 x azir-CH), 2.22 (s, 3Η, 2-CH ₃ ), 3.85 (s, 3Η, CH ₃ N-), 4.24 (br, 1Η, CΗ ₂ OH), 4.63 (br, 2Η, CH ₂ OH) and 5.73 (s, IH, 6-H). Anal. C; 65.93, Η; 6.50, N; 10.12% Calc. (C ₅ Η _j8 N ₂ O ₃ ) C; 65.69, H; 6.57, N; 10.22%.

Example 25: 3-Hvdroxvmethvl-5-( 2.2-dimethvlaziridin-l-vll-1.2-dimethvlindole- 4.7-dione (561 bv reaction (viiil of Figure 4.

A solution of 52 (100 mg, 0.43 mmol) in 2,2-dimethylaziridine (2 mL) was stirred at 95 °C for 5 hours. After this time (reaction does not go to completion) the solution was cooled and evaporated in vacuo, and the residue redissoived in EtOAc and evaporated to precipitate 56 as a dark red solid, which was unstable on silica and in solution: mp 138-141 °C; ^! H-NMR (CDC1 ₃ ) δ 1.32 (s, 6H, 2 x azir-CH ₃ ), 2.07 (s, 2Η, azir-CH ₂ ), 2.22 (s, 3Η, 2-CH ₃ ), 3.87 (s, 3Η, CH3N-), 4.25 (br, 1Η, CΗ ₂ OH), 4.86 (br, 2Η, CH ₂ OH) and 5.68 (s, IH, 6-H). Anal. C; 62.92, Η; 6.88, N; 9.58% Calc. (C ₁₅ Η _{j 8} N2θ3.2/3Η ₂ O) C; 62.94, H; 6.76, N; 9.79%.

Example 26: 3-Hvdroxvmethvl-5-( 2.2-dimethvl-2-hvdroxvethyllaminol-1.2- dimethvlindole-4.7-dione (571 bv reaction (viiil of Figure 4.

Compound 56 (25 mg, mmol) was dissolved in 5 mL H2O with warming to 45 °C and then evaporated in vacuo at 50°C. The residue was purified on silica, eluting with EtOAc (R =0.35) to give 57, after recrystallization from EtOAc: mp 188-190°C;

^] H-NMR ((CD ₃ ) ₂ SO) δ 1.14 (s, 6H, 2 x CH ₃ ), 2.21 (s, 3Η, 2-CH ₃ ), 3.02 (d, 2Η, J=7.2 Hz, NHCH ₂ C(CH ₃ ) ₂ ), 3.84 (s, 3H, CH ₃ N-), 4.58 (br, 2Η, CH ₂ OH), 4.65 (br, IH, CH ₂ OH), 5.14 (s, 1Η, 6-H) and 6.63 (br t, 1Η, NH). Anal. C; 60.62, Η; 6.96, N; 9.34% Calc. (C ₁₅ Η ₂₀ N ₂ O ₄ .1/3Η ₂ O) C; 60.40, H; 6.94, N; 9.39%.

Example 27: Preparation of 3-Hvdroxvmethvl-5-(2-methvlaziridin-1-vll-l-methvl- indole-4.7-dione (621 bv reactions (II to (vl of Figure S.

(a) 5-Methoxy-l-methylindo.e-3-carboxaldehyde (58). 5-methoxyindole-3-carboxaldehyde (2.0 g, 11.4 mmol) was added portionwise to a suspension of NaH (0.55 g, 13.7 mmol) in DMF (50 mL) with stirring. The suspension was stirred for 0.5 hour, and Mel (1.94 g, 13.7 mmol) added and the mixture stirred for 1 hour at room temperature. The reaction mixture was then poured on to NaHCO ₃ (10%, 300 mL) and extracted wih EtOAc (4 x 75 mL), washed with NaHCO ₃ (10%, 3 x 50 mL), sat. NaCl (3 x 100 mL), dried and evaporated in vacuo to give 58 (1.70 g, 79%) as a white solid: mp 132-133 °C; ^! H-NMR (CDC1 ₃ ) δ 3.81 (s, 3H, CH ₃ N-), 3.89 (s, 3Η, CH ₃ O-), 7.08 (dd, 1Η, J=2 and 9 Ηz, Ar-6Η), 7.31 (d, IH, J=9 Hz, Ar-7H), 7.59 (s, IH, 2-H), 7.8 (d, IH, J=2 Hz, Ar-4H) and 9.93 (s, 1Η, CHO).

(b) 5-Methoxy-l-methyl-4-nitroindole-3-carboxaldehyde (59).

To a solution of 56 (1.50 g, 7.94 mmol) dissolved in AcOΗ (150 mL) was added a mixture of c.ΗNO ₃ (4.5 mL) in AcOH (25 mL) dropwise at 0°C over 3 hours. After addition, the mixture was stirred at room temperature for 16 hours, and then added to crushed ice (75 g), filtered and washed with H2O (5 x 100 mL) and dried to give 57 (1.56 g, 84%) as a pale yellow solid: mp 197-198°C; *H-NMR ((CD ₃ ) ₂ SO) δ 3.94 (s, 6H, CH ₃ O- and CH ₃ N-), 7.32 (d, 1Η, J=9 Ηz, Ar-7H), 7.72 (d, 1Η, J=9 Ηz, Ar-6H), 8.27 (s, 1Η, 2-H) and 9.75 (s, 1Η, CHO).

(c) 4-Amino-5-methoxy-l-methylindole-3-carboxaIdehyde (60).

To a suspension of 59 (1.0 g, 4.27 mmol) in EtOΗ (150 mL) was added tin (4.43 g, 37 mmol) followed by ΗCI (3.0 M, 60 mL). The mixture was stirred at room temperature for 2 hours and decanted. The solution was added portionwise to sat. NaΗCO ₃ (aq., 300 mL). and extracted with EtOAc (3 x 100 mL). The organic layer was separated, washed with sat NaHCO ₃ (aq., 2 x 175 mL), sat. NaCl (3 x 75 mL), dried and evaporated in vacuo to give 60 (0.79 g, 91%) as a dark yellow solid which was used in the next step without further purification; Rf=0.64 (EtOAc);

^! H-NMR (CDC1 ₃ ) δ 3.75 (s, 3H, CH ₃ N-), 3.88 (s, 3Η, CH ₃ O-), 5.79 (br s, 2Η, NH ₂ ), 6.56 (d, IH, J=9 Hz, Ar-7H), 7.53 (d, 1Η, J=9 Ηz, Ar-6H), 7.64 (s, 1Η, 2-H) and 9.60 (s, 1Η, CHO).

(d) 5-metho ^χ y-l-methyl-4,7-dioxoindole-3-carboxaldehyde (61). To 60 (0.75 g, 3.68 mmol) dissolved in Me CO (75 mL) was added Fremy's salt (4.0 g, 14.9 mmol) in Η ₂ O (20 mL) followed by a solution of Na ₂ HPO ₄ /NaH ₂ PO ₄ buffer (0.3 M, pH 6, 20 mL). The mixture was stirred for 0.75 hour, excess Me^O removed and the product filtered and washed with H ₂ O (50 mL), dried, and recrystallized from EtOAc to give 61 (0.61 g, 76%) as a yellow solid: mp 188-190°C; -NMR (CDC1 ₃ ) δ 3.87 (s, 3H, CH ₃ N-), 4.02 (s, 3Η, CH ₃ O-), 5.78 (s, 1Η, 6-H), 7.44 (s, 1Η, 2-H) and 10.54 (s, 1Η, CHO).

(e) 3-Ηydroxymethyl-5-methoxy-l-methylindole-4,7-dione (62).

To a solution of 61 (0.5 g, 2.28 mmol) in anhydrous MeOH (300 mL) was added NaBH4 (0.65 g, 17 mmol). The solution was degassed with argon and stirred for 2 hours under argon and then evaporated in vacuo to give a solid which was diluted with CH ₂ C1 ₂ (300 mL) and washed with H ₂ O (2 x 100 mL), sat NaCl (100 mL) and condensed to give 62 as an orange solid (0.2 g, 40%) after recrystallization from EtOAc: mp 185-186°C; *H-NMR (CDC1 ₃ ) δ 3.85 (s, 3H, CH ₃ N-), 3.94 (s, 3Η, CH ₃ O-), 4.25-4.29 (m, 2H, CH ₂ OH), 5.73 (s, IH, 6-H) and 6.88 (s, 1Η, 2-H). Anal. C; 59.84, Η; 4.79, N; 6.30% Calc. (C _j JΗJ _j NO ₄ ) C; 59.73, Η; 4.98, N; 6.33%.

Example 28: Preparation of 5-(Aziridin-1-vll-3-hvdroxvmethvl-l-methvlindole-4.7- dipqe (63) by reaction (vj) pf Figure g _f

A solution of 62 (200 mg, 0.9 mmol) in l(Η)-aziridine (1.5mL, ca. 35 mmol, CAUTION!) was stirred at room temperature for 1.5 hours. Excess aziridine was removed in vacuo, and the product was recrystallized from EtOAc to give 63 (130 mg, 62%) as an orange solid: mp 169-171 °C; ¹ H-NMR (CDCl ₃ ) δ 2.22 (s, 4H, 2 x aziridine-CH ₂ ), 3.91 (s, 3Η, CH ₃ N-), 4.64 (s, 2Η, CH ₂ OH), 5.81 (s, IH, 6-H),

and 6.69 (s, IH, 2-H). Anal. C; 61.47, Η; 5.14, N; 12.22% Calc. (C ₁₂ Η ₁₂ N ₂ O ₃ ) C; 62.07, H; 5.17, N; 12.07%.

Example 29: Preparation of 3-Hvdroxvmethvl-5-(2-methvlaziridin-1-yll-1-methyl- indole-4.7-dione (641 bv reaction (vii of Figure 5. A solution of 62 (200 mg, 0.9 mmol) in 2-methylaziridine ( 1 mL) was stirred at room temperature for 2.5 hours. Excess 2-methylaziridine was removed in vacuo, and the product was purified on silica, eluting with EtOAc (Rf=0.6) to give 64 (120 mg, 54%) as a red solid recrystallized from EtOAc: mp 89-90°C; ^-NMR (CDC1 ₃ ) δ 1.47 (d, 3H, J=4.5 Hz, azir-CH ₃ ), 2.04-2.2 (m, 3Η, azir-CHCH ₂ ), 3.91 (s, 3Η, N-CH3), 4.67 (m, 2Η, CH ₂ OH), 5.79 (s, 1 H, 6-H) and 6.70 (s, 1 Η, 2-H).

Example 30: Preparation of 5-methoxv-l-methvl-3-l(carbamovloxvlmethvll indole- 4.7-dioπe (651 bv reaction (viii of Figure 5.

To a solution of 62 (0.1 g, 0.45 mmol) in anhydrous pyridine (5 mL) at 0°C, was added dropwise phenylchloroformate (0.1 g, 0.64 mmol) and the solution then allowed to reach room temperature and stirred for 2 hours. The solution was then extracted with CΗ C1 ₂ (30 mL) and washed with H ₂ O (35 mL) and sat. NaCl (35 mL), dried and evaporated. The residue was purified on silica, eluting with EtOAc (Rf=0.8) to give an orange solid of 5-methoxy-l-methyl-3-[[(phenoxycarbonyl)oxy]methyl] indole: mp 85-86°C; *H-NMR ((CD ₃ ) ₂ SO/CDCl ₃ ) δ 3.79 (s, 3H, CH ₃ N-), 3.91 (s, 3Η, CH ₃ O-), 5.43 (s, 2Η, CH ₂ OCOPh), 5.67 (s, 1Η, 6-H), 6.89 (s, 1Η, 2-H) and 7.15-7.3 (m, 5Η, Ar). This material (0.1 g, 0.3 mmol) was dissolved in anhydrous CH ₂ C1 ₂ (18 mL) and the solution cooled to -78 °C. The solution was then saturated with NH ₃ and stirred at -78 °C until reaction was complete (0.75 hour). The solution was then allowed to reach room temperature and evaporated in vacuo. The residue was redissoived in CH ₂ C1 ₂ (100 mL) and washed with H O (2 x 100 mL) and sat. NaCl (50 mL), dried and evaporated, and the residue recrystallized from EtOAc to afford 75 mg (98%) of 65 as an orange solid: mp 231-234°C (dec); ^! H-NMR ((CD ₃ ) ₂ SO/CDCl ₃ ) δ 3.78 (s, 3H, CH ₃ N-), 3.88 (s, 3Η, CH ₃ O-),

5.04 (s, 2H, CH ₂ OCONH ₂ ), 5.77 (s, IH, 6-H), 6.42 (br s, 2Η, NH ₂ ) and 7.09 (s, 1Η, 2-H). Anal. C; 54.98, Η; 4.56, N; 10.18%, Calc. (C ₁₂ Η ₁₂ N ₂ O ₅ ) C; 54.55, H; 4.55, N; 10.61%.

Example 31: Preparation of 5-(Aziridin-l-vll-l-methvl-3-f(carbamovloxvlmethvl] indole-4.7-dione (661 bv reaction (viiil of Figure 5.

Compound 65 (50 mg, 0.2 mmol) was stirred for 0.5 hour in l(H)-aziridine (ca. 35 mmol), evaporated in vacuo and redissoived in EtOAc. The solution was evaporated again and the residue recrystallized from EtOAc to afford 35 mg (70%) of 66 as a red solid: mp 195-198°C (dec); -NMR ((CD ₃ ) ₂ SO) δ 2.18 (s, 4Η, 2 x azir- CH ₂ ), 3.90 (s, 3Η, CH ₃ N-), 5.23 (s, 2H, CH ₂ OCONH ₂ ), 5.49 (br s, 2H, NH ₂ ) 5.76 (s, 1Η, 6-H) and 6.86 (s, 1Η, 2-H). Anal. C; 56.70, Η; 4.94, N; 15.35%, Calc. (C ₁₃ Η ₁₃ N ₃ O ₄ ) C; 56.73, H; 4.73, N; 15.27%.

Further compounds were pepared in order to investigate the effect of different leaving groups at the R* position in Formula I; these compounds being divided into three formula options A, B and C and the corresponding chloromethyl compound. By keeping the groups R, R^ and R ³ methyl and R ⁴ hydrogen, as in Examples 32 to 44 and 46 to 49, bioassay could be used to evaluate the effect of different leaving groups. Example 45 describes a preferred compound where R^ is cyclopropyl.

B

Exam ^p le 32: Pre ^p aration of 3-ChIoromethvl-1.2-dimethyl-5-methoxyindole-4.7- dione (671

5-Methoxy-3-hydroxymethyl-l,2-dimethylindole-4,7-dione (52, 500mg, 1.97mmol) was stirred at room temperature with 5mL SOCI2 for 0.5h. The solution was then evaporated in vacuo, redissoived in EtOAc (25mL) and evaporated to dryness. This procedure was repeated twice and the crude orange solid (0.43 g, 85%) of 3-cUoromethyl-l,2-dimethyl-5-methoxyindole-4,7-dione (mp 204-205"C (dec)) was used in the next step without further treatment. A small sample was recrystallized from EtOAc for NMR and CHN analysis. *H-NMR (CDC1 ₃ ) d 2.28 (s, 3H, 2-CH ₃ ), 3.80 (s, 3Η, C/ 3N-), 3.89 (s, 3H, CH ₃ O-), 4.86 (s, 2Η, CH ₂ C1) and 5.62 (s, 1Η, 6-H). Anal. C; 55.71, Η; 5.41, N; 5.70, Cl; 14.00% Calc. C ₁₂ Η ₁₂ NO ₃ C1.0.33Η ₂ O C; 55.49, H; 4.88, N; 5.39, Cl; 13.68%

GF.NER Aϊ, METHOD FOR THE SYNTHESIS OF TARGET COMPOUNDS OF

5-Methoxy-3 -hydroxymethyl- 1 ,2-dimethylindole-4,7-dione (52, 500mg, 1.97mmol) was dissolved in CH C1 ₂ (50mL) together with pyridine (5mL) and the appropriate carbonyl chloride (5mmol) added. The solution was the heated under reflux for the reaction time given below, cooled and EtOAc (150mL) added followed by HCl (aq., 0.1M, 150mL). The organic layer was separated and washed again with HCl (aq., 0.1M, lOOmL), sat. NaCl (aq., lOOmL), dried and evaporated in vacuo. The residue was purified on silica, eluting with the solvent specified below and recrystallized to give the target compound.

B : GENERAL METHOD FOR THE SYNTHESIS OF TARGET COMPOUNDS OF TYPE B AND TYPE C 3-Chloromethyl-l ,2-dimethyl-5-methoxyindole-4,7-dione (67, 1 OOmg, 0.39mmol) was dissolved in EtOAc (20mL) and the appropriate alcohol, phenol or thiol (lmmol) added dropwise or in portions with stirring. Stirring was continued at room temperature for the

reaction times given below and then water (20mL) added. The organic layer was separated and washed with sat.NaHCO ₃ (aq., 20mL) and sat NaCl (aq., 20mL), dried and evaporated to dryness. The residue was purified on silica, eluting with the solvent specified below, and recrystallized to give the target compound.

Example 33: Preparation of l-2-Dimethvl-5-methoχy-3-(2-nitrobenzoyloxyl- ethvlindole-4.7-dione (681

General method A was employed (reaction time 0.75h), and the product eluted on silica with EtOAc (Rf=0.65), and recrystallized from EtOAc to give an orange solid (68%) : mp 199-200°C; -NMR (CDC1 ₃ ) d 2.34 (s, 3H, 2-CH ₃ ), 3.80 (s, 3Η, CH ₃ N), 3.91 (s, 3Η, CH ₃ O), 5.53 (s, 2Η, CH ₂ OCOAr), 5.62 (s, 1Η, 6-H), 7.50-8.07 (m, 4Η, 4 x Ar-H). Anal.

Example 34: Preparation of 1.2-Dimethvl-3-(2-fluorobenzovloxvlmethvl-5- methow indole-4.7-dione (691.

General method A was employed (reaction time 1.5h), and the product eluted on silica with EtOAc (Rf=0.6), and recrystallized from EtOAc to give a yellow solid (83%): mp 166-168°C; ^! Η-NMR (CDC1 ₃ ) d 2.35 (s, 3H, 2-CH ₃ ), 3.81 (s, 3Η, CH ₃ N), 3.90 (s, 3Η, CH ₃ O), 5.51 (s, 2Η, CH ₂ OCOAr), 5.63 (s, 1Η, 6-H), 6.98-7.52 (m, 3Η, 3 x Ar-H) and 7.83-8.00 (dd, lΗ, J=9Ηz andJ=1.5Ηz, Ar3-H). Anal. C; 62.83, Η^4.59, N; 3.76% Calc. (C ₁₉ H ₁₆ NO ₅ F.0.33H ₂ O) C; 62.81, H; 4.59, N; 3.86%

Example 35: Preparation of 1.2-Dimethvl-3-(4-fluorobenzovloxvlmethvl-5- methoxvindole-4.7-dione (701.

General method A was employed (reaction time 1.25h), and the product eluted on silica with EtOAc (Rf=0.65), and recrystallized from EtOAc to give a yellow solid (88%) : mp 209-210°C; *H-NMR (CDC1 ₃ ) d 2.35 (s, 3H, 2-CH ₃ ), 3.81 (s, 3Η, CH ₃ N), 3.91 (s, 3Η, CH ₃ O), 5.49 (s, 2Η, CH ₂ OCOAr), 5.63 (s, 1Η, 6-H), 6.95-7.27 (dd, 2Η, J=9Hz and J=lHz, Ar2,6-H) and 7.94-8.10 (dd, 2Η, J=9Hz andJ=1.5Hz, Ar3,5-H). Anal. C; 63.34, Η; 4.44, N; 3.88% Calc (C ₁₉ Η ₁₆ NO ₅ F) C; 63.86, H; 4.51, N; 3.92%.

Example 36: Preparation of 3-Benzovloxvmethvl-1.2-dimethyl-5-methoxyindole- 4,7-dione (71)

General method A was employed (reaction time 1.5h), and the product eluted on silica with EtOAc (Rf=0.75), and recrystallized from EtOAc/Me ₂ CO to give a yellow solid (69%) : mp 168-170°C; ^! H-NMR (CDC1 ₃ ) d 2.31 (s, 3H, 2-CH ₃ ), 3.76 (s, 3Η, CH ₃ N), 3.86 (s, 3Η, CH ₃ O), 5.46 (s, 2Η, CH ₂ OCOAr), 5.58 (s, 1Η, 6-H), 7.34-7.41 (m, 3Η, 3 x ArH) and 7.93-8.01 (m, 2Η, 2 x Ar-H).

Example 37: Preparation of 3-(2-Acetoxvhenzovloxvl-methvl-1.2-dimethvl-S- methoxvindole-4.7-dione (721. General method A was employed (reaction time 1.5h), and the product eluted on silica with EtOAc (Rf=0.5), and recrystallized from EtOAc to give a yellow solid (90%): mp 159-161°C; -NMR (CDC1 ₃ ) d 2.27 (s, 3Η, CH ₃ CO ₂ Ar), 2.31 (s, 3Η, 2-CH ₃ ), 3.81 (s, 3Η, CH ₃ N), 3.90 (s, 3Η, CH ₃ O), 5.47 (s, 2Η, CH ₂ OCOAr), 5.63 (s, 1Η, 6-H), 7.01-7.53 (m, 3Η, 3 x ArH) and 7.94-8.04 (dd, 1Η, J=9Ηz and J=lHz, Ar-H). Anal. C; 63.81, Η; 4.81, N; 3.71% Calc (C ₂₁ Η _j 9NO ₇ ) C; 63.47, H; 4.82, N; 3.52%

Example 38: Preparation of 3-Acetoxvmethvl-1.2-dimethvl-5-methoxvindole-4.7- dione (73)

General method A was employed (reaction time 0.5h), and the product eluted on silica with EtOAc (Rf=0.6), and recrystallized from EtOAc to give a yellow solid (90%): mp 185-186°C; ^! H-NMR (CDC1 ₃ ) d 2.04 (s, 3H, CH ₃ CO), 2.28 (s, 3Η, 2-CH ₃ ), 3.81 (s, 3Η, CH ₃ N), 3.90 (s, 3Η, CH3O), 5.24 (s, 2Η, CH ₂ OCOCH ₃ ) and 5.62 (s, IH, 6-H).

Example 39: reparation of 3-(Cvclohexvlcarbonv.oxvl-methvl-1.2-dimethvl-5- methoxvindole-4.7-dione (741. General method A was employed (reaction time 0.25h), and the product eluted on silica with EtOAc (Rf=0.75), and recrystallized from EtOAc to give a yellow solid (65%) : mp 164-165°C; *Η-NMR d 1.2-1.83 (m, 1 IH, 11 x cyclohexyl-H), 2.27 (s, 3Η, 2-CH ₃ ), 3.80 (s, 3Η, CH ₃ N), 3.89 (s, 3Η, CH ₃ O), 5.22 (s, 2Η, CH ₂ OCOR) and 5.62 (s, 1Η, 6-H).

Example 40: Preparation of 1.2-Pimethvl-3-(4-nitrophenoxyl-methyl-5- pιethoxyindole-4.7-dione (75).

General method B was employed (reaction time 0.5h), and the product eluted on silica with EtOAc/hexane (1:1, Rf=0.7), and recrystallized from EtOAc to give an orange solid (54%) : mp 215-216°C (dec); *H-NMR d 2.33 (s, 3H, 2-CH ₃ ), 3.81 (s, 3Η, CH ₃ N), 3.89 (s, 3Η, CH ₃ O), 4.99 (s, 2Η, CH ₂ OAr), 5.62 (s, 1Η, 6-H) and 6.79-6.93 (m, 4Η, 4 x Ar-H).

Example 41: Preparation of 1.2-Dimethvl-3-(4-fluorophenoxyl-methyl-5- methoxvindole-4.7-d-one (761. General method B was employed, and the product eluted on silica with EtOAc/hexane (1:1, Rf=), and recrystallized from EtOAc to give an solid (40%).

Example 42: Preparation of U-Dimethvl-3-(2-fluorophenoxvl-methvl-5- methoxvindo1e-4.7-dione (771.

General method B was employed, and the product eluted on silica with EtOAc hexane (1:1, Rf=), and recrystallized from EtOAc to give an solid (55%).

Example 43: Preparation of 3-(2-Carbomethoxvthiophenvll-methvl-1.2-dimethvl- 5-methoxvindole-4.7-dione (781.

General method B was employed (reaction time 2h), and the product eluted on silica with EtOAc/hexane (1:1, Rf=0.25), and recrystallized from EtOAc to give an orange solid (66%) : mp 214-216°C ; ^! Η-NMR (CDC1 ₃ ) d 2.21 (s, 3H, 2-CH ₃ ), 3.77 (s, 3Η, CH ₃ N), 3.86 (s, 6Η, CH ₃ O and CO ₂ CH ₃ ), 4.39 (s, 2Η, CH ₂ SAr), 5.59 (s, 1Η, 6-H), 7.28-7.49 (m, 3Η, 3 x ArH) and 7.95 (d, 1Η, J=9Ηz, Ar-H).

Example 44: Preparation of 3-Benzyloxymethyl-1.2-dimethyl-5-methoxyindole-4.7- dione (791. General method B was employed (reaction time lh) but with the addition of K ₂ CO ₃ (138mg, lmmol) to the reaction mixture, and the product eluted on silica with

EtOAc/hexane (1:1, Rf=0.6), and recrystallized from EtOAc to give an orange solid (53%): mp 132-133°C (dec); *H-NMR (CDC1 ₃ ) d 2.25 (s, 3H, 2-CH ₃ ), 3.80 (s, 3Η, CH ₃ N), 3.86 (s, 3Η, CH ₃ O), 4.59 (s, 2Η, 3-CH ₂ OCH ₂ Ar), 4.74 (s, 2Η, 3-CH ₂ OCH ₂ Ar), 5.60 (s, IH, 6-H) and 7.28-7.36 (m, 5Η, 5 x ArH). Anal. C; 67.81, Η; 5.95, N; 4.49% Calc. (C _j9 Η _j9 Nθ4.0.5Η ₂ O) C; 68.26, H; 5.99, N; 4.19%

Example 45: Preparation of 3-Benzovloxvmethvl-2-cvc.opropvl-5-methoxv-l- methvlinrlole-4.7-dione (801: 2-cvclopropane derivative.

General method A was employed (reaction time 1.5h) using compound 20, and the product eluted on silica with EtOAc/hexane (1:1, Rf=0.35), and recrystallized from EtOAc/Me ₂ CO to give a yellow solid (68%): mp 221 -223°C; ¹ H-NMR (CDC1 ₃ ) d 0.65-0.82 (m, 2H, cyclopropyl-CH ₂ ), 1.11-1.23 (ra, 2Η, cyclopropyl-CH ₂ ), 1.61-1.71 (m, 1Η, cyclopropyl-H), 3.79 (s, 3Η, CH ₃ N), 4.03 (s, 3Η, CH ₃ O), 5.56 (s, 2Η, CH ₂ OCOAr), 5.64 (s, 1Η, 6-H), 7.31-7.49 (m, 3Η, 3 x ArH) and 7.96-8.08 (m, 2Η, 2 x Ar-H). Anal. C; 69.04, Η; 5.30, N; 3.82% Calc. (C ₂₁ Η ₁₉ NO ₅ ) C; 69.03, H; 5.24, N; 3.83%.

Example 46: 5-(2-Methvlaziridin-l-vll-3-(2-nitrohenzoyloxymethyll-1.2- dimethvlindole-4.7-dione (811.

Compound 3 (O.lg, mmol) was dissolved and stirred in freshly distilled 2-methylaziridine (3mL, ca.50mmol) for 2.5h. The solution was evaporated in vacuo and the residue redissoived in EtOAc, evaporated until a red precipitate appeared and the solid collected. The red solid was recrystallized from EtOAc to give 81 (85mg, %): mp 168-170°C; *H-NMR (CDC1 ₃ ) d 1.42 (d, 3H, J=4.5Hz, azir-CH ₃ ), 2.01-2.15 (m, 3Η, azir-CHCH ₂ ), 2.33 (s, 3Η, 2-CH ₃ ), 3.90 (s, 3Η, CH ₃ N-), 5.54 (s, 2Η, CH ₂ OCOAr), 5.73 (s, 1Η, 6-H), 7.53-8.01 (m, 4Η, 4 x Ar-H).

^0-

Example 47: Preparation of 5-(2-Methvlaziridin-1-vll-3-benzoyloxymethyl-1.2- dimethylindole-4.7-dione (821.

This compound was prepared from 6 by the same method used for the synthesis of 82. The red solid was recrystallized from EtOAc to give 17 (70 %): mp 133-134°C; ^! H-NMR (CDC1 ₃ ) d 1.42 (d, 3H, J=4.5Hz, azir-CH ₃ ), 2.01-2.15 (m, 3Η, azir-CHCH ₂ ), 2.33 (s, 3Η, 2-CH ₃ ), 3.90 (s, 3Η, CH ₃ N-), 5.54 (s, 2Η, CH ₂ OCOAr), 5.73 (s, 1Η, 6-H), 7.34-7.41 (m, 3Η, 3 x ArH) and 7.93-8.01 (m, 2Η, 2 x Ar-H). Anal. C; 67.96, Η; 5.67, N; 7.23%, Calc (C _2j Η ₂₀ N ₂ O ₄ .0.5Η ₂ O) C; 67.56, H; 5.63, N; 7.51%.

Example 48: Prenration of 1.2-Dimethvl-3-(4-methvlpiperazin-l-yllmethyl-5- methoxyindole-4.7-dione (831.

General method B was employed (reaction time lh) but using N-methylpiperazine (0.65mL, 0.58mmol) in place of the alcohol or thiol, and the product eluted on silica with MeOH (Rf=0.15) to give 83 as an orange solid (38%): mp 159-162°C (dec); !H-NMR (CDC1 ₃ ) d 2.24 (s, 6H, 2-CH ₃ and N-CH ₃ ), 2.31-2.62 (m, 8Η, 4 x piperazine- CH ₂ ), 3.73 (s, 2Η, 3-CH ₂ N), 3.79 (s, 3Η, CH ₃ N), 3.88 (s, 3Η, CH ₃ O) and 5.69 (s, 1Η, 6-H).

Example 49: Preparation of 5-(Aziridin-l-vll-1.2-dimethvl-3-(4-methvlpiperazin-l- yi)methy.indple-4,7-diQiιe (84),

Compound 83 (50mg, 0.158mmol) was stirred for 0.3h with IH-aziridine (CAUTION!) at room temperature. Excess aziridine was removed in vacuo and the residue twice redissoived in EtOAc (5mL) and evaporated to dryness. The residue was recrystallised from EtOAc to give 84 as a dark red solid ( mg, 62%): mp 143-146°C (dec); *Η-NMR (CDC1 ₃ ) d 2.17 (s, 4Η, aziridine), 2.25 (s, 6H, 2-CH ₃ and N-CH ₃ ), 2.31-2.62 (m, 8Η, 4 x piperazine-CH ₂ ), 3.74 (s, 2Η, 3-CH ₂ N), 3.87 (s, 3Η, CH ₃ N), and 5.73 (s, 1Η, 6-H).

The amount of compound to be adminstered will of course vary with individual and cancer type. However, suitable doses will be typically within the range l-200mg/kg bodyweight, preferably 10-150mg/kg and more preferably 30-lOOmg/kg.

The form of adminstration will also vary, with formulations suitable for oral, rectal, topical, parenteral (including subcutaneous, intramuscular and intravenous injection) being useable. Formulations may range from pure drug and simple aqueous solutions thereof , eg. in water or saline, to those including solid and/or more complex components such as liquid or solid carriers, diluents and active agents. Particular forms available include inter alia capsules, cachets, tablets or lozenges, boluses, electuaries or pastes containing solid or liquid compound of the invention, or suspensions, syrups, emulsions etc in liquid form. Suitable formulations will occur to those skilled in the art of pharmacy and may be exemplified by those used for the current lead EO9 and the antimicrobial agents described previously.

Simple saline injections may be used for the purpose of demonstrating efficacy of the compounds.

Biological Evaluation of Compounds 18 to 84 In Vitro Example 50: MTT assay

Selective toxicity to hypoxic V79-379A cells was determined for all compounds using the MTT assay as has been described previously (see references 7 to 9). These results are presented in the Table 1 below, where C50 (air) values, the concentration required to kill 50% of the aerobic cells under the conditions of the assay, are divided by C^ (N ₂ ) values, concentrations required to kill hypoxic cells, to give hypoxic cytotoxicity ratios (HCR), which enable quantitative comparisons of bioreductive activities of drugs.

TABLE 1 Compounds 1 to 66

to

I

B

Type R R, Rr. HCR"

1 MMC 2.0 ^a

2 A CH ₂ OCONH

E09 t> 1.0 ^b

50.3

108.4±5.9 60.6±7.4 1.8 0.965±0.1 1 0.073±0.01 1 13.2

103.5+5.2 59.8±1 1.4 1.7

540±52 820±83 0.65

250±24 60±9.8 4.2

1.72±0.18 0.93±0.067 1.8

0.603±0.099 0.0058±0.0013 103.5

3.33±0.75 0.074±0.015 45.0

130±9.8 5.6±0.6 23.4

12.5±2.9 0.459±0.096 27.23

150+15 150+15 1.0 e Me

36 B MeO CH ₂ OH . e 800+80 200±20 4.0 "Me

37 B CH ₂ OH .Me 25.7+3.0 0.79±0.14 32.5 - "Me

38 B Me 127.8±13.5 13.9+1.5 9.2

40 B MeO CH ₂ OCONH ₂ „Me 20.9+1.67 0.117±0.018 178.6 Me

41 B _\ CH ₉ OCONH ₂ „Me 47.5+14.6 5.46+0.43 8.7 ""Me

4 171

42 B 20.6±2.5 0.87±0.215 23.7

43 B MeO Me Me 1.236+0.189 0.657+0.063 1.88 Me

44 B Me t> ,Me 0.333±0.014 0.188±0.011 1.77 "Me

45 B MeO CH ₂ OH Cyclohexyl 122.4±17.5 20.5±5.3 6.0

46 B o- CH ₂ OH Cyclohexyl 34.5+.6.0 0.93+0.078 37.0

52 B MeO CH ₂ OH Me 1077±44 284.8±37.6 3.78 53 B CH ₂ OH Me 0.149±0.011 0.0116±0.0008 12.8

[>-

54 B Me CH ₂ OH Me 94±7.6 0.5±0.07 188

55 B CH ₂ OH Me 202±11 14.2±1.7 14.2

I 56 B Me Me H CH ₂ OH Me 1260±126 500+120 2.5 σs HO

I

62 B MeO 220±20 240±23 0.92 63 B 0.153±0.013 0.0086±0.0009 15.4 -

64 B Me 4.42+1.17 0.0179±0.02 24.7

65 B MeO 3.1 ±0.24 0.037±0.007 83.8 66 B ι>- 0.00019±0.000032 0.00013±0.000025 1.46

*HCR = hypoxic cytotoxicity ratio (C5ø(Air)/C5ø(N ₂ )). ^a Stratford, I.J. et α/.(1990), in Selective Activation of Drugs by Redox Processes; Adams, G.E. et al, Eds., Plenum, N.Y. ^b Moody, CJ. et α/.(1994), Anti-Cancer Drugs, 5, 367-372.

TABLE 2: LEAVING GROUP ACID DISSOCIATION CONSTANTS AND HCR (MTT) OF DRUGS WITH VARYING INDOLOQUINONE 3-METHYLENE LEAVING GROUPS

RB

RB

RB

RB 21.3

RB96717N AcOH 4.75 27.3 2.45 11.1 73 1.54 ±0.18

R 63.4

RB 0.86

1.7

RB94547J 50 H ₂ O 15.74 1077.0 284.8 3.78 ±44.0 ±37.6

1. Streitwieser, A.; Heathcock, C. H. In Introduction to Organic Chemistry, Macmillan Pub. Co., Inc., New York, 1981.

2. Weast, R.C.; Astle, M. J. In Handbook of Chemistry and Physics (62nd edn.) 1981-82. 3, 4, 5 Values for C ₂ H ₅ CO ₂ H, C ₆ H ₅ SH and C ₂ H ₅ OH from ref.1.

I o Other compounds prepared include those where the leaving group is 2,4,6 trichloro phenol and 4-dimethylamino benzoac acid having pKa

I (H ⁺ ) of 6.0 and 4.92 respectively.

TABLE 3:DATA AVAILABLE ON COMPOUNDS 80-84

I (XI

Example 52; Effect of compounds on RIF-1 and KHT rumour cells when administered with single dose radiation regimen.

Compounds 21 and 54 (RB 94573 and RB 94577) were applied to RIF-1 and KHT tumour cells in separate experiments with presence and absence of single doses of radiation (15GY and 10 GY respectively for 21 and 54). Maximum tolerated doses and tumour response experiments were carried out as described in reference 9. Tumour bearing mice were given doses of radiation chosen to kill most of the oxic cells in the tumours with the measured response reflecting the survival of residual hypoxic clonogenic cells (see reference 10). Indoquinones were given immediately after X-rays such that any therapeutic response greater than that achieved by radiation alone is a reflection of residual hypoxic cell killing. These conditions were achieved by giving 10GY to KHT tumours and 25 GY to RIF-1 tumours. Each experiment also included mice exposed to radiation without drug and with drug alone.

Results are shown in Figures 6 to 9. It can be seen that while the results are additive in the case of the KHT cells, providing destruction of tumour cells down to a surviving fraction of as low as less than 10 , the RIF-1 cells show a synergistic effect with surviving fraction being less than 10 .

Example 53: Effect of compound 54 (RB 94S77Ϊ with fractionated dose radiation ∑SgjmSΑ-

Four daily doses of 2.5 Gy of X-rays were administered to RIF-1 cells followed immediately by 20mg/Kg of compound 52 after each fraction. Results are given in the Table 4 below.

TABLE 4 RB94577 T54Ϊ + X-ravs in fractionated treatments

4 daily doses of 2.5 Gy X-rays followed immediately by 20 mg/kg RB94577 after each fraction. Tumours were excised 24 hours after last dose of drug to assay cell survival.

TABLE 5

Effect of fractionated treatments with RB94577 f54^ and radiation on RIF-1 tumours.

TABLE 6

Interaction between RB94577 (54 and cisPlatin in RTF tumour-no-radiation.

*The time between adminstration of the first mention agent and the second is indiated in minutes. Effect of the compound of the invention and cisPlatin is greater than additive in this assay.

IΔBLE Zi

Compound 53 as compared to EQ9.

DTD= DT diaphorase activity in nmol cytochrome c reduced per min per mg protein IC5 (nM) concentration required to kill 50% of cells following four days incubation of cells with drug in ajj.

* Compound 24 ** Compound 84

ΪΔBLE_2

Comparative tumour toxicities (relative surviving fraction of compounds 21 and 54.

Both drugs were given at 90 mg/kg ip and tumours were excised at 24 hours.

TABLE 10

Effects of Compound 54 as compared to EO9 of US 5097257.

Compound 54 was given at 30 mg/kg i-P- while EO9 was given at 5 mg/kg. The maximum tolerated dose of Compound 54 is about 100 mg.kg while the max tolerated dose of EO9 is 5 mg/kg. H249 is human small cell cancer type; HT29 is human colon cancer type and H647 is human non-small cell lung cancer type.

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