DITHIOLE COMPOUNDS AS COX INHIBITORS

FIELD

The present invention relates to compounds useful as pharmaceuticals, in particular as analgesic, anti-inflammatory and/or anti-neurodegenerative agents, processes for their preparation and related uses.

BACKGROUND

Paracetamol (also known as acetaminophen or 4-(acetylamino)phenol) is a widely-used analgesic and antipyretic drug used for the relief of mild pain and fever.

4-(Acetylamino)phenol

Irrespective of its widespread use and availability, paracetamol is subject to a number of disadvantages. One problem associated with paracetamol is that excessive doses cause liver damage. In the case of large overdoses, liver function deteriorates leading to jaundice, confusion, and loss of consciousness. If the dose is sufficiently excessive to cause death, the death is usually due to liver failure.

An associated disadvantage is that the difference between a typical dose effective for the relief of pain and/or inflammation is very close to the toxic dose - that is it has a relatively narrow therapeutic index. The toxic dose of paracetamol in adults varies due to several factors, however doses above 10 grams or regular doses over 5 grams per day in a healthy adult can cause significant damage to the liver,

As a consequence there is a need for alternatives to paracetamol that have similar therapeutic indications, but preferably without one of more of the disadvantages associated with paracetamol.

It would be advantageous for such alternatives to have wider therapeutic indexes than that of paracetamol, to minimise the risk of overdose, and to avoid liver

damage.

It is alternatively or additionally desirable to identify paracetamol alternatives that are more lipophilic than paracetamol, to improve potency.

Common analgesics other than paracetamol include the non-steroidal anti-inflammatory drugs (NSAIDs) such as ibuprofen and aspirin. These drugs act by blocking the cyclooxygenase (COX) enzymes COX-I and COX-2. Cyclooxygenase blocking inhibits the production of prostaglandin - the body's inflammatory response to injury - and thus reduces pain and inflammation. Whilst paracetamol has no significant action on COX-I and COX-2, recent research has indicated that paracetamol selectively inhibits COX-3, found in the brain and spinal cord.

It is of interest to explore new compounds and to study the activity of these compounds on the range of cyclooxygenases.

It would also be advantageous to obtain a compound that was useful to treat other diseases, disorders or conditions associated with cyclooxygenase, if any paracetamol alternatives are found having such activity.

SUMMARY

The present application provides a compound of formula (I):

wherein:

R] and R ₂ are the same or different and are independently selected from H and a shielding group;

X and Y are each independently selected from N and CH;

R ₃ is hydroxy, alkoxy, acyloxy or an ester group;

R ₄ is a direct bond to R ₅ , -CH ₂ - or -CH=; the broken line — represents an optional double bond between R ₄ and R ₅ ; and R ₅ is a 5- or 6-membered substituted or unsubstituted unsaturated heterocyclic or heteroaromatic ring, or R ₅ is a substituted heterocyclic ring containing the substituent = O or = S, wherein the heteroatom is selected from S, O or P, and wherein R ₅ is optionally attached to R ₄ through a substituent on the heterocyclic or heteroaromatic ring; or a pharmaceutically acceptable prodrug, metabolite, ester, salt, derivative, tautomer or isomer thereof.

The present invention also provides use of compounds of formula (I) as pharmaceuticals. Thus, the present invention provides a pharmaceutical agent comprising a compound of formula (I), in which the pharmaceutical agent is an analgesic, anti-inflammatory agent, anti-neurodegenerative agent and/or a COX inhibitor. A particular class of compounds of formula (I) in which R ₃ is alkoxy, acyloxy or ester, provides compounds which are particularly useful as selective COX-2 inhibitors. The compounds of formula (I) may also be used to treat a COX-mediated disease, disorder or condition.

The present invention further provides a method for the treatment of pain, inflammation or a neurodegenerative disease, disorder or condition, comprising administering a therapeutically effective amount of a compound according to formula (I) to a subject in need thereof.

The present invention further provides a method for the inhibition of COX, comprising administering a therapeutically effective amount of a compound according to formula (I) to a subject in need thereof.

The present invention further provides a method for the selective inhibition of COX-2, comprising administering a therapeutically effective amount of a compound according to formula (I) in which R ₃ is alkoxy, acyloxy or ester to a subject in need thereof.

The present invention further provides a method for the treatment and/or prophylaxis of a COX-mediated disease, disorder or condition, comprising

- A -

administering a therapeutically effective amount of a compound according to formula (I) to a subject in need thereof.

The present invention provides for the use of a compound according to formula (I) in the manufacture of a medicament for the treatment and/or prophylaxis of the above-mentioned diseases, disorders or conditions.

The present invention provides for the use of a compound according to formula (I) in the manufacture of a medicament for COX inhibition. According to one embodiment, the medicament is for selective COX-2 inhibition.

The compound of formula (I) is advantageously administered in the form of a pharmaceutical composition together with a pharmaceutically acceptable carrier, excipient or diluent. Thus, the present invention provides a pharmaceutical composition comprising the compound of formula (I) and a pharmaceutically acceptable carrier, excipient or diluent.

The present invention further provides a method of inhibiting COX in a cell comprising contacting the cell with a compound of formula (I).

The present invention also provides a process for preparing the compound of formula (I) comprising the step of: converting a compound of formula (II) below:

wherein R ₁ , R ₂ , R ₃ , X and Y are as defined above; into a compound of formula (I).

The present invention also provides a process for preparing the compound of formula (I) comprising the step of:

deprotecting a compound of formula (III) below:

wherein PG is a hydroxy protecting group and R ₁ , R ₂ , X, Y, R ₄ and R ₅ are as defined above; to produce a compound of formula (I).

DETAILED DESCRIPTION

As used herein, the singular forms "a," "an" and "the" include plural reference unless the context clearly dictates otherwise.

Compounds

The compounds of formula (I) are structurally related to paracetamol, which is known for its analgesic properties. Previously 2,6-di-t-butylphenols have been shown to possess anti-inflammatory properties (Lazer et al (1989) and Kramer et al (1995)), but no extensive research has been performed on paracetamol derivatives with the particular ring structures now claimed in place of the acetamide group of paracetamol, and the efficacy of these compounds in the treatment and/or prophylaxis of diseases and/or disorders.

For ease of explanation, the ring system containing atoms X and Y, and substituents R ¹ , R ² , and R ³ will be referred to as the "base" ring.

• Heterocyclic or Hetero aromatic Ring

The compounds of formula (I) possess a substituted or unsubstituted unsaturated heterocyclic or heteroaromatic ring represented by R ₅ .

The term "heterocyclic" used either alone or in compound words such as "substituted or unsubstituted unsaturated heterocyclic ring" denotes monocyclic heterocyclic rings containing at least one heteroatom selected from P, O or S. Suitable heterocyclic rings include unsaturated 5 to 6-membered heteromonocyclic rings containing 1 to 4 P, O or S atoms. One such example is l,2-dithiole-3-thione. This ring may be attached to the base ring via the 4- or 5- position. The other of the 4- and 5- position may be unsubstituted or may contain a substituent such as an alkyl or aryl group.

The term "heteroaromatic" used either alone or in compound words such as "substituted or unsubstituted heteroaromatic ring", denotes single aromatic rings containing at least one heteroatom selected from P, O or S. Suitable heteroaromatic rings include unsaturated 5 to 6-membered heteromonoaromatic rings containing 1 to 4 P, O or S atoms. Suitable heteroaromatic rings include furanyl and the like.

R ₅ preferably comprises a pair of conjugated double bonds. The pair of conjugated double bonds can be present in the following combinations:

(i) the first double bond is constituted by a double bond between R ₄ and R ₅ , the second is constituted by a double bonded substituent on R ₅ , or by a ring- forming double bond in R ₅ ; or

(ii) the first double bond is a ring-forming double bond in R ₅ and the second is a double bonded substituent on R ₅ ; or

(iii) the conjugated double bonds are both ring-forming double bonds in the ring of R ₅ .

The unsaturated heterocyclic or heteroaromatic ring represented by R ₅ can be optionally substituted by one or more of the following groups ("substituents"): straight or branched chained alkyl, alkenyl, hydroxy, alkoxy, alkenyloxy, alkoxyalkyl, alkoxyalkenyl, halo, haloalkyl, haloalkenyl, haloalkoxy, haloalkenyloxy, haloalkoxyalkyl, haloalkoxyalkenyl, mercapto, thio, alkylthio, alkylthioalkyl, alkylthioalkenyl, alkenylthio, alkenylthioalkyl, alkenylthioalkenyl, thioalkoxy, thioalkoxyalkyl, thioalkoxyalkenyl, thiocarbonyl, carbonyl, acyl, alkenylacyl, acyloxy, alkenylacyloxy, acylthio, alkenylacylthio, alkylsulfmyl, alkenylsulfinyl, alkylsulfonyl, alkenylsulfonyl, alkylsufenyl, alkenylsulfenyl, nitro, nitroalkyl, nitroalkenyl, amino, alkylamino, dialkylamino, alkenylamino, dialkenylamino, acylamino, diacylamino or the like.

In the case of 'carbonyP or 'thiocarbonyP, it is noted that the carbon of these substituents may be within the ring system of R ₅ , and therefore these groups may alternatively be considered to be double bonded oxygen or sulphur substituents, that is =0 or =S substituents on the ring.

It follows that R ₅ encompasses the following types of groups: 5-membered rings with 1 double bond and 1 heteroatom — for example:

5-membered rings with 1 double bond and 2 heteroatoms - for example:

5-membered rings with 2 double bonds and 1 heteroatom - for example:

5-membered rings with 2 double bonds and 2 heteroatoms — for example:

5 le bonds and 3 heteroatoms - for example:

6-membered rings with 1 double bond and 1 heteroatom - for example:

6-membered rings with 1 double bond and 2 heteroatoms - for example:

6-membered rings with 2 double bonds and 1 heteroatom - for example:

6-membered rings with 2 double bonds and 2 heteroatoms - for example:

In each of the above examples, the heteroatom-containing ring is attached to R ₄ (or to the base ring directly) via any available site on the ring or through a substituent on the ring. Preferably, attachment is from an atom in the ring.

R ₅ may be a saturated ring, containing the substituent = O or = S, for example:

According to one embodiment, R ₅ is selected from:

5-membered rings with 1 double bond and 3 heteroatoms; or 5-membered rings with 2 double bonds and 3 heteroatoms; or 5-membered rings with 2 double bonds and 2 heteroatoms; or

6-membered rings with 2 double bonds and 2 heteroatoms.

More preferably, R ₅ is selected from the following:

It is noted in the above that the point of attachment of the group R ₅ to R ₄ can be from any suitable atom in the ring or R ₅ , or through a substituent on the ring of R ₅ as the following examples demonstrate.

Some examples of compounds of formula (I) are as follows:

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• Shielding Group

One subclass of the compounds of formula (I) possess a shielding group at Ri and/or R ₂ - preferably shielding groups at both Rj and R ₂ . The function of the shielding group is to slow down the rate of conjugation of the group R ₃ with sulfate and glucuronide in the kidneys, which slows down the rate of excretion of the conjugated compound from the body, thereby lengthening the therapeutic effect that the compound of formula (I) has on a subject. This lengthened therapeutic effect can also result in smaller doses of the compound having to be taken, and less frequent dosages in order to be effective compared to a compound which does not bear one or more shielding groups. Examples of such shielding groups are straight or branched chained alkyl, alkoxy, alkoxyalkyl, haloalkyl, haloalkoxy, halo or haloalkoxyalkyl groups. Methyl and t-butyl groups are notable examples.

• R ₃

The group R ₃ of the compounds of the present invention forms a conjugate with the sulfhydryl group of glutathione in the kidneys when being metabolised. The group R ₃ is hydroxy, alkoxy (such as methoxy or ethoxy) acyloxy (such as benzyloxy) or an ester group. Compounds of formula (I) have shown cyclooxygenase (COX) activity, resulting in an anti-inflammatory effect and analgesia. Certain compounds of formula (I) have also been shown to have selective COX-2 activity, often (although not exclusively) when R ₃ is alkoxy or acyloxy.

In the case where R ₃ is an ester, the term "ester" is used in the broadest sense to encompass any organic esters, in which the ester may contain any other functional groups such as alkyl, aryl, amine, alkylamine. One notable example of a suitable ester is the carboxylate group. The esters may contain other functional groups such as amines, alkylamines and so on. One example is diethylglycine ester.

• Subclasses of compounds

Notable subclasses of compounds of the present invention are as follows:

- (Ia) - Ri and R ₂ are each shielding groups;

R ₃ is alkoxy, acyloxy or ester; and X, Y and R ₄ to R ₅ are as broadly defined for formula I.

(Ib) - any of compounds of formula (I), with the proviso that:

(i) when R ₄ is a direct bond to R ₅ , located para to R ₃ , and R ₅ is:

and is attached direct to the aromatic ring of formula (I) from the carbon atom at position 4 marked, then either

Z is a substituent other than hydrogen mercapto or thioester (preferably it is alkyl), or - R ₃ is alkoxy or ester, or

R ₁ and R ₂ are both H,

or

(ii) when R ₄ is a direct bond to R ₅ , located para to R ₃ , and R ₅ is:

and is attached direct to the aromatic ring of formula (I) from the carbon atom at position 5 marked, then

Ri and R ₂ are both shielding groups, or Z is H or a substituent other than alkyl, or Ri and R ₂ are H, and R ₃ is selected from OH, ester, ethoxy and benzyloxy.

(Ic) X = N and Y = CH, and Rj to R ₅ are as broadly defined for formula I

(Id) R ₅ is a 6-membered ring, and X, Y and Ri to R ₄ are as broadly defined for formula I

(Ie) R ₅ is a 5-membered ring containing =0, R ₁ and R ₂ are shielding groups, and X, Y, R ₃ and R ₄ are as broadly defined for formula I.

5 Uses

The compounds of formula (I) have been shown to be COX inhibitors. Certain subclasses of the compounds have been found to be selective COX-2 inhibitors.

The two principal forms of cyclooxygenase (COX) that have been 0 studied are the constitutive isoform (COX-I) and an inducible isoform (COX-2).

Expression of these enzymes is upregulated at sites of inflammation (Vane, J. R. et. al., (1994)). COX-I appears to play a physiological role and to be responsible for gastrointestinal and renal protection. COX-2 appears to play a pathological role and is believed to be the predominant isoform present in inflammation conditions. The use of 5 conventional COX-I inhibitors are limited due to side effects such as ulceration and liver and renal toxicity. Compounds that selectively inhibit COX-2 exert antiinflammatory effects without the adverse side effects associated with COX-I inhibition.

COX inhibition refers to inhibition of at least one form of O cyclooxygenase, and therefore encompasses inhibition of one or both of COX-I and

COX-2 to a degree considered to be statistically significant to a person skilled in the art.

The term "selective COX-2 inhibitor" refers to a compound able to inhibit COX-2 without significant inhibition of COX-I, e.g., the degree of inhibition of 5 COX-2 compared to COX-I inhibition that would be considered statistically significant by people of ordinary skill in this art. Preferably, this includes compounds which have a COX-2 IC ₅₀ of less than about 25 μM, and also have a selectivity ratio of COX-I inhibition over COX-2 inhibition of at least about 5, and more preferably of at least about 25. Preferably, the compounds have a COX-I IC ₅₀ of greater than about lOμM, O and more preferably of greater than about 1 OOμM.

The compounds of formula (I) are useful as pharmaceuticals, in particular as analgesics, as anti-inflammatory agents and as anti-neurodegenerative agents. 5

Analgesia refers to the relief of pain. The term "pain" encompasses both acute and chronic pain. The term "acute pain" means immediate, generally high

threshold, pain brought about by chemical stimulation. The term "chronic pain" means pain other than acute pain. It is understood that chronic pain often is of relatively long duration, for example, months or years and can be continuous or intermittent. Such pain includes inflammatory pain, neuropathic pain, acute pain, chronic pain, post- operative pain and pain associated with migraine, arthralgia, nerve injury, neurodegeneration, neuropathies, diabetic neuropathy, hyperactive urinary bladder, arthritis, hypersensitive urinary bladder, urinary incontinence, interstitial cystitis, bladder disorders, irritable bowel syndrome, inflammatory bowel disease, inflammatory disease, asthma, chronic obstructive pulmonary disease, digestive tract ulcer, skin irritation, eye irritation and mucous membrane irritation.

The term "inflammation" as used herein refers to short-term and chronic inflammation. Chronic inflammation is marked by inflammation lasting many days, months or even years, and may lead to the formation of a chronic wound. Short-term inflammation means inflammation other than chronic inflammation. Examples of inflammation include appendicitis, gastritis, laryngitis, and meningitis.

The term "neurodegenerative disease, disorder or condition" as used herein refers to a condition which affects brain function. They are divided into two groups; conditions causing problems with movements and conditions affecting memory and conditions related to dementia. Examples of neurodegenerative diseases and/or disorders include: Alexander disease, Alper's disease, Alzheimer disease, Amyotrophic lateral sclerosis, Ataxia telangiectasia, Canavan disease, Cockayne syndrome, Corticobasal degeneration, Creutzfeldt- Jakob disease, Huntington disease, Kennedy's disease, Krabbe disease, Lewy body dementia, Machado- Joseph disease

(Spinocerebellar ataxia type 3), Multiple sclerosis, Multiple System Atrophy, Parkinson disease, Pelizaeus-Merzbacher Disease, Pick's disease, Primary lateral sclerosis, Refsum's disease, Sandhoff disease, Schilder's disease, Spinocerebellar ataxia (multiple types with varying characteristics), Spinal muscular atrophy, Steele-Richardson- Olszewski disease, Tabes dorsalis, cerebral amyloid angiopathy, cognitive disorders, progeria, epileptic dementia, pre-senile dementia, post-traumatic dementia, senile dementia, vascular dementia, HIV-I -associated dementia, post-stroke dementia, Down's Syndrome and motor neuron disease.

Substituents

In order to provide a clear and consistent understanding of the terms used in this specification, the following definitions are provided.

• Compounds

The term "alkyl" used either alone or in a compound word such as "optionally substituted alkyl" denotes straight chain, branched or mono- or poly- cyclic alkyl, preferably Ci _-30 alkyl or cycloalkyl. Examples of straight chain and branched alkyl include methyl, ethyl, propyl, isopropyl, the different butyl isomers and the like.

The term "alkenyl" used either alone or in compound words such as "alkenyloxy" denotes groups formed from straight chain, branched or cyclic alkenes including ethylenically mono-, di- or poly-unsaturated alkyl or cycloalkyl groups as defined above, preferably C ₂ - ₂₀ alkenyl. Examples of alkenyl include vinyl, allyl, 1- methylvinyl, butenyl, and the like.

The term "acyl" used either alone or in compound words such as "optionally substituted acyl" denotes groups formed from an acyl group, preferably C ₁ - C ₃ o acyl. Examples of acyl groups include straight chain or branched alkanoyl such as formyl, acetyl, propanoyl and butanoyl.

The term "alkoxy" used either alone or in compound words such as "optionally substituted alkoxy" denotes straight chain or branched alkoxy, preferably Ci-C ₃₀ alkoxy. Examples of alkoxy include methoxy, ethoxy, n-propyloxy, isopropyloxy and the different butoxy isomers. The term "alkoxy" is used broadly to encompass substituted and unsubstituted alkoxy. However, unsubstituted alkoxy are one subclass of particular interest.

The term "acyloxy" denotes aromatic ring-containing groups, such as phenyloxy, benzyl oxy, napthyloxy, and derivatives thereof in which one or more of the aromatic ring atoms contain a substituent. Preferably the acyloxy is unsubstituted acyloxy.

The term "halo" either used alone or in combination with words such as "haloalkyl" refers to fluorine, chlorine, bromine or iodine.

The present invention includes within its scope "prodrugs" of the compounds of formula (I). In general, such prodrugs will be functional derivatives of the compound of formula (I) which are readily convertible in vivo into the required compound of formula (I).

Preferably the "derivative" is a "pharmaceutically acceptable derivative". By "pharmaceutically acceptable derivative" is meant any pharmaceutically acceptable salt, solvate, ester, ether, amide, active metabolite, analogue, residue or any other compound which is not biologically or otherwise undesirable and induces the desired pharmacological and/or physiological effect. Conventional procedures for the selection and preparation of suitable prodrug derivatives are described, for example, in Design of Prodrugs ed. H. Bundgaard.

The salts of the compound of formula (I) are preferably pharmaceutically acceptable, but it will be appreciated that non-pharmaceutically acceptable salts also fall within the scope of the present invention, since these are useful as intermediates in the preparation of pharmaceutically acceptable salts. Examples of pharmaceutically acceptable salts include sodium, potassium, lithium, calcium, and the like. In addition, some of the compounds of the present invention may form solvates with water (e.g. hydrates) or common organic solvents. Such solvates are encompassed within the scope of the invention.

The term "tautomer" is used herein in its broadest sense to include compounds of formula (I) which are capable of existing in a state of equilibrium between two isomeric forms. Such compounds may differ in the bond connecting two atoms or groups and the position of these atoms or groups in the compound.

The term "isomer" is used herein in its broadest sense and includes structural, geometric and stereo isomers. As the compound of formula (I) may have one or more chiral centres, it is capable of existing in enantiomeric forms.

• Uses

The term "subject" as used herein refers to any animal having a disease or condition which requires treatment with a pharmaceutically-active active agent. The subject may be a mammal, preferably a human.

The term "therapeutically effective amount" refers to an amount of a compound of the present invention effective to yield a desired therapeutic response, for example, to treat, ameliorate or prevent a disease and/or condition. The term

"therapeutically effective amount" will, obviously, vary with such factors as the particular condition being treated, the physical condition of the subject, the type of

subject being treated, the duration of the treatment, the nature of concurrent therapy (if any), and the specific formulations employed and the structure of the compound or its derivatives.

Generally, the term "treatment" means affecting a subject, tissue or cell to obtain a desired pharmacological and/or physiological effect and include: (a) preventing the disease from occurring in a subject that may be predisposed to the disease, but has not yet been diagnosed as having it; (b) inhibiting the disease, i.e., arresting its development; or (c) relieving or ameliorating the effects of the disease, i.e., cause regression of the effects of the disease.

• Pharmaceutical Compositions

The compositions of the present invention comprise at least one compound of formula (I) together with one or more pharmaceutical acceptable carriers, diluents and/or excipients. Such carriers, diluents and/or excipients can include solubilising agents, such as cyclodextrins. The pharmaceutical acceptable carriers, diluents and/or excipients must be pharmaceutically "acceptable" in the sense of being compatible with the other ingredients of the composition and not injurious to the subject. Compositions include those suitable for oral, rectal, nasal, topical, vaginal or parenteral (including subcutaneous, intramuscular, intravenous, intrathecal, intracranial, and intradermal) administration. Methods and carriers for preparation of pharmaceutical compositions are well known in the art, as set out in textbooks such as Remington's Pharmaceutical Sciences.

The pharmaceutical compositions are preferably prepared and administered in dose units. Solid dose units may be tablets, capsules and suppositories. Doses may be administered as a single dose unit or as several smaller dose units to a subject.

Dosage levels of the compound of formula (I) of the present invention are of the order of about 0.5 mg to about 100 mg per kilogram body weight, with a preferred dosage range between about 0.5 mg to about 50 mg per kilogram body weight per day (from about 0.5 grams to about 5 grams per patient per day). The amount of active ingredient that may be combined with the carrier materials to produce a single dosage will vary depending upon the host treated and the particular mode of administration. For example, a formulation intended for oral administration to humans may contain about 5 mg to 5 g of an active compound with an appropriate and

convenient amount of carrier material which may vary from about 5 to 95 percent of the total composition. Dosage unit forms will generally contain between from about 5 mg to 5000 mg of active ingredient.

Optionally the compounds of the invention are administered in a divided dose schedule, such that there are at least two administrations in total in the schedule. Administrations are given preferably at least every two hours for up to four hours or longer; for example the compound may be administered every hour or every half hour. In one preferred embodiment, the divided-dose regimen comprises a second administration of the compound of the invention after an interval from the first administration sufficiently long that the level of active compound in the blood has decreased to approximately from 5-30% of the maximum plasma level reached after the first administration, so as to maintain an effective content of active agent in the blood. Optionally one or more subsequent administrations may be given at a corresponding interval from each preceding administration, preferably when the plasma level has decreased to approximately from 10-50% of the immediately-preceding maximum.

EXAMPLES

The invention will now be described in detail by way of reference only to the following non-limiting examples.

General methods

Thin layer chromatography (TLC) was performed with Merck Silica Gel 60 F ₂₅₄ , using mixtures of petroleum spirits-ethyl acetate. Detection was effected by visualization in UV light. NMR spectra were obtained on a Unity 400, Inova 400 or a Inova 500 machine (Melbourne, Australia) operating at 400 MHz for and 500 MHz for ¹ H and at 100 MHz or 125 MHz for ¹³ C. Infra-red spectra were obtained as thin films using a Perkin-Elmer Spectrum One FTIR spectrometer with a zinc selenide/diamond Universal

ATR sampling accessory. Flash chromatography was performed according to the method of Still et al. with Scharlau Silica Gel 60, using adjusted mixtures of ethyl acetate-petroleum spirits. Solvents were evaporated under reduced pressure using a rotary evaporator. Melting points were obtained using a Reichert-Jung hot stage and are corrected. Elemental analyses were performed by Chemical and Micro Analysis Services Pty. Ltd. (Belmont, Victoria). High resolution mass spectra were performed by Mr Chris Barlow at the School of Chemistry, University of Melbourne.

Glossary of Abbreviations

AlCl ₃ = Aluminium chloride Ac ₂ O = Acetic anhydride

CuI = Copper (I) iodide

DMF = Dimethyl formamide

DMPU = Tetrahydro-l,3-dimethyl-2(lH)-pyrimidinone

EtI = Ethyl Iodide EtOAc = Ethyl acetate

Hg(OAc) ₂ = Mercuric acetate

HMDO = Hexamethyldisiloxane

HMDT =1,1,1,3,3 ,3 -Hexamethyldisilathiane

HOAc = Acetic acid KH = Potassium Hydride

K ₂ CO ₃ = Potassium carbonate

Magnesium sulfate = MgSO ₄

Me ₂ CO ₃ = Dimethyl carbonate

MeOH = Methanol NaHCO ₃ = Sodium bicarbonate

NaH = Sodium Hydride

Na ₂ SO ₄ = Sodium sulfate

P ₄ Si ₀ ⁼ Phosphorous pentasulfide dimmer

Petrol = Petroleum Spirit 40-60 ⁰ C S = SuIfUr

ZnCl ₂ = Zinc chloride

l-(3,5-Di-fertf-butyl-4-methoxymethoxyphenyl)ethanone

Acetyl chloride (1.6 niL, 23 mniol) was added dropwise to a solution of dimethoxymethane (2.0 niL, 23 mmol), and ZnCl ₂ (0.6 mg, 4.4 μmol) in toluene (6.0 niL) and the reaction stirred at rt for 4 h. l-(3,5-Di-tert-butylhydroxyphenyl)ethanone (1.00 g, 4.03 mmol) was added followed by iV-ethyl-N-(l-methylethyl)-2-propanamine (3.5 mL, 20 mmol) and the reaction was stirred at rt overnight. Water was added and the mixture stirred for 15 min. The aqueous layer was extracted with EtOAc (x 2) and the combined organic extracts washed with sat. NaHCO ₃ (x 1), water (x 2), brine (x 1), dried (Na ₂ SO ₄ ) and concentrated. The residue was purified by flash chromatography (10% EtO Ac/petrol) and the residue recrystallised from EtOH/water to afford l-(3,5-di- fert-butyl-4-methoxymethoxyphenyl) ethanone as a light yellow solid (1.67 g, 71%); mp 60-61 ⁰ C; ¹ H NMR (500 MHz, CDCl ₃ ) δ 0.88 (s, 18H, t-Bu x 2), 1.98 (s, 3H, COCH ₂ ), 3.06 (s, 3H, OMe) ₅ 4.33 (s, 2H, CH ₂ ), 7.32 (s, 2H, Ar); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 26.4 (CH ₃ ), 31.8 (C(CH ₃ ) ₃ ), 35.8 (CH ₃ ), 57.4 (CH ₃ ), 100.7 (CH ₂ O), 127.0, 132.1, 144.7, 159.0 (4Ar), 197.8 (CO); IR v 2956, 2873, 1762, 1676, 1589, 1227, 881 cm-'HRMS ESI ⁺ [M+Na] ⁺ = 315.1931, requires 315.1931 for C ₁₈ H ₂₈ O ₃ Na;

Microanalysis: Found C, 73.92; H, 9.70; Cj ₈ H ₂₈ O ₃ requires C, 73.93; H, 9.65%.

3-(3,5-Di-tert-butyI-4-methoxymethoxyphenyl)-3-oxopropion ic acid methyl ester

l-(3,5-Di-tert-butylmethoxymethylphenyl)ethanone (1.00 g, 3.42 mmol) in THF (4.0 mL) was added dropwise over 2 h to a refluxing solution of washed (hexane) sodium hydride (680 mg, 17.0 mmol, 60% dispersion in mineral oil) and dimethyl carbonate (1.5 mL, 18 mmol) in THF (8.0 mL). The reaction was heated for a further 30 min then cooled to rt. Water (50 mL) was then added followed by ether. The organic extract was washed with water (x 3), sat. NaHCO ₃ (x 1), brine (x 1), dried (Na ₂ SO ₄ ) and concentrated. The residue was purified by flash chromatography (10% EtO Ac/petrol) and the residue recrystallised from EtOH/water to afford 3-(3,5-di-fert-butyl-4- methoxymethoxyphenyl)-3-oxopropionic acid methyl ester as a colourless solid, (1.15 g, 97%); mp 79-80 °C; ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.43 (s, 18H, t-Bu x 2), 3.69 (s, 3H, OMe) ₃ 3.71 (s, 3H, OMe), 3.74 (s, 3H, OMe), 3.79 (s, 3H, OMe), 4.00 (s, 2H,

CH ₂ ), 5.59 (s, IH, C=CH), 7.66 (s, 2H, Ar), 7.87 (s, 2H, Ar), 12.53 (CR=COH); ¹³ C NMR (100 MHz ₃ CDCl ₃ ) δ 32.0 (C(CH ₃ ) ₃ ), 36.2 (CH ₃ ), 46.1 (CH ₂ ), 52.7 OCH ₃ ), 57.8 (CH ₃ ), 101.1 (CH ₂ O), 127.6, 131.1, 145.4, 160.0 (Ar-4), 168.4 (CO), 192.1 (CO); IR v 2956, 2876, 1744, 1680, 1619, 1203, 883 cm- ¹ , HRMS ESI ⁺ [M+Naf = 373.1984, requires 373.1985 C ₂₀ H ₃₀ O ₅ Na; Microanalysis: Found C, 68.60; H, 8.61; C ₂₀ H ₃₀ O ₅ requires C, 68.54; H, 8.63%.

5-(3,5-Di-ter^-butyl-4-hydroxyphenyl)-3JS-l,2-dithiole-3- thione

P ₄ S ₁₀ (1.15 g, 2.59 mmol), sulfur (130 mg, 4.05 mmol) and HMDO (2.4 mL, 11 mmol) were added to a solution of 3-(3,5-di-ter^butyl-4-methoxymethoxyphenyl)-3- oxopropionic acid methyl ester (1.3O g, 3.71 mmol) in xylene (15 mL). The reaction was heated under reflux for 4 h, cooled to rt and concentrated. The residue was purified by flash chromatography (5% EtO Ac/petrol) and the residue recrystallised from EtO Ac/petrol to afford 5-(3,5-di-før^-butyl-4-hydroxyphenyl)-3/f-l,2-dithiole-3-th ione 1 as a yellow-brown solid, (540 mg, 43%); mp 180-183 ⁰ C; ¹ H NMR (500 MHz, CDCl ₃ ) δ 1.47 (s, 18H, r-Bu x 2), 5.72 (s, IH, OH), 7.41 (s, IH, CH), 7.48 (s, 2H, Ar); ¹³ C NMR (125 MHz, CDCl ₃ ) δ 30.3 (C(CH ₃ ) ₃ ), 34.8 (CH ₃ ), 123.5 (C4), 124.5 (C5), 134.7, 137.5, 158.1, 175.1 (Ar-4), 215.2 (CS); IR v 3429, 2960, 1593, 1514, 1419, 889, 715 cm- ¹ ; HRMS ESI ⁺ [M+H] ⁺ = 339.0908, requires 339.0911 for C _n H ₂₄ OS ₃ .

3-(3,5-DWert-butyl-4-methoxyphenyl)-3-oxopropionic acid methyl ester

l-(3,5-Di-tert-butylmethoxyphenyl)ethanone (420 mg, 1.60 mmol) in THF (4.0 mL) was added dropwise over 2 h to a refluxing solution of washed (hexane) sodium hydride

(192 mg, 8.00 mmol, 60% dispersion in mineral oil) and dimethyl carbonate (0.80 mL, 9.5 mmol) in THF (8.0 mL). The reaction was heated for a further 45 min then cooled to rt. Acetic acid (5 mL in 20 mL of water) was added followed by ether. The organic extract was washed with water (x 3), sat. NaHCO ₃ (x 1), brine (x 1), dried (Na ₂ SO ₄ ) and concentrated. The residue was purified by flash chromatography (5%

EtOAc/petrol) to afford 3-(3,5-di-fert-butyl-4-methoxyphenyl)-3-oxopropionic acid methyl ester as a brown oil, (490 mg, 96%); ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.43 (s, 18H, t-Bu x 2), 3.69 (s, 3H, OMe), 3.71 (s, 3H, OMe), 3.74 (s, 3H, OMe), 3.79 (s, 3H ₃ OMe), 4.00 (s, 2H, CH ₂ ), 5.59 (s, IH, C=CH), 7.66 (s, 2H, Ar), 7.87 (s, 2H, Ar), 12.53 (CH=COH); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 31.7 (C(CH ₃ ) ₃ ), 35.8 (CH ₃ ), 35.9, 45.8 (CH ₂ ), 51.2 (CH ₃ ), 52.3 (CH ₃ ), 64.3, 64.4, 85.9, 124.6, 127.4, 130.5, 144.0, 144.4, 164.6 (Ar-4), 168.1 (CO), 191.8 (CO); IR v 2956, 2876, 1744, 1680, 1619, 1203, 883 cm ^"1 ; HRMS ESI ⁺ [M+Na] ⁺ = 343.1877, requires 343.1885 for Ci ₉ H ₂₈ O ₄ Na.

5-(3,5-Di-ter/-butyl-4-methoxyphenyl)-3H-l,2-dithioIe-3-t hione

P ₄ Si ₀ (356 mg, 0.801 mmol), sulfur (400 mg, 12.5 mmol) and ηMDO (1.4 mL, 6.47 mmol) were added to a solution of 3-(3,5-di-ført-butyl-4-methoxyphenyl)-3- oxopropionic acid methyl ester (363 mg, 1.13 mmol) in xylene (2.0 mL). The reaction was heated under reflux for 3 h and then cooled to rt. The reaction mixture was applied to silica gel and purified by flash chromatography (5% EtO Ac/petrol). The residue recrystallised from EtOAc/petrol to afford 5-(3,5-di-ferr-butyl-4-methoxyphenyl)-3//- l,2-dithiole-3-thione 2 as an orange solid, (117 mg, 28%); mp 106-107 °C; (lit. ² mp

100-101 ⁰ C) ₅ ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.45 (s, 18H, /-Bu x 2), 3.74 (s, 3H, OCH ₃ ), 7.41 (s, IH, C=CH), 7.53 (s, 2H, Ar); ¹³ C NMR (100 MHz ₅ CDCl ₃ ) δ 31.8 (C(CH ₃ ) ₃ ), 36.2 (CH ₃ ), 64.6 (OCH ₃ ), 125.5 (C4), 126.2 (C5), 135.2, 145.6, 163.5, 174.2 (Ar-4), 215.2 (CS); IR v 2948, 2865, 1744, 1587, 1498, 1304, 1 110, 782 cm ^"1 ; HRMS ESI ⁺ [MH-H] ⁺ = 353.1063, requires 353.1062 for Ci ₈ H ₂₅ OS ₃ ; Microanalysis: Found C, 61.33; H, 6.85; S, 27.34. C ₁₈ H ₂₃ OS ₃ , requires C, 61.32; H, 6.86; S, 27.28%.

3-(3,5-Di-fø/*t-butyl-4-ethoxyphenyl)-2-methyI-3-oxoprop ionic acid methy! ester

l-(3,5-Di-tert-butylethoxyphenyl)ethanone (780 mg, 2.82 mmol) in THF (5.0 niL) was added dropwise over 2 h to a refluxing solution of washed (hexane) sodium hydride (564 mg, 14.1 mmol, 60% dispersion in mineral oil) and dimethyl carbonate (1,4 mL, 17 mmol) in THF (7.0 mL). The reaction was then heated for a further 30 min then cooled to rt. Water (20 mL) was then added followed by ether. The organic extract was washed with water (x 3), sat. NaHCO ₃ (x 1), brine (x 1), dried (Na ₂ SO ₄ ) and concentrated. The residue was purified by flash chromatography (10% EtO Ac/petrol) to afford 3-(3,5-di-fert-butyl-4-ethoxyphenyl)-2-methyl-3-oxopropionic acid methyl ester as a brown oil, (511 mg, 55%); ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.01 (t, 3H, J= 7.2 Hz, CH ₂ CH ₃ ), 1.02 (s, 18H, t-Bu x 2), 3.32 (s, 3H, CH ₃ ), 3.37 (q, 2H, J= 7.2 Hz ₅ CH ₂ CH ₃ ), 3.56 (s, 3H, CH ₃ ), 7.47 (s, 2H ₅ Ar); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 14.6 (CH ₃ ), 31.6 (C(CHs) ₃ ), 35.7 (CH ₃ ), 45.6 (CH ₂ ), 52.1 (OCH ₃ ), 71.9 (CH ₂ ), 124.5, 127.4, 130.3, 144.2, 163.0 (Ar-5), 168.0 (CO), 191.6 (CO); IR v 2956, 1742, 1680, 1435, 1383, 1197, 733 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 335.2217, requires 335.2217 for C ₂₀ H ₃₁ O ₄ .

5-(3,5-Di-før^butyl-4-ethoxyphenyI)-3#-l,2-dithiole-3-th ione

P ₄ Si ₀ (400 mg, 0.900 mmol), sulfur (55.0 mg, 1.71 mmol) and HMDO (1.9 mL, 8.9 mmol) were added to 3-(3,5-di-te?t-butyl-4-ethoxyphenyl)-2-methyl-3-oxopropionic acid methyl ester (500 mg, 1.49 mmol) in xylene (3.0 mL). The reaction mixture was heated under reflux for 1 h. The reaction mixture was applied to silica gel and was purified by flash chromatography (5% EtO Ac/petrol). The residue was recrystallised from EtO Ac/petrol to afford 5-(3,5-di-terf-butyl-4-ethoxyphenyl)-3H-l,2-dithiole-3-

thione 3 as an orange solid, (238 mg, 43%); mp 87-89°C; ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.44 (s, 18H, t-Bu x 2), 1.44 (t, J= 7.2 Hz, 3H, OCH ₂ CH ₃ ), 3.80 (q, J= 7.2 Hz, 2H, OCH ₂ CH ₃ ), 7.41 (s, IH, CH), 7.52 (s, 2H, Ar); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 14.9 (CH ₃ ), 31.8 (C(CHs) ₃ ), 36.1 (CH ₃ ), 72.3 (CH ₂ ), 125.6 (C4), 126.0 (C5), 135.1, 145.6, 162.1, 174.3 (Ar-4), 215.2 (CS); IR v 2963, 1510, 1426, 1384, 1217, 1056, 888 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 367.1219, requires 367.1219 for Ci ₉ H ₂₇ OS ₃ .

5-(3,5-Di-tert-butyl-4-hydroxyphenyl)-3JEr-l,2-dithiol-3- one

A hot solution OfHg(OAc) ₂ (2.18 g, 6.84 mmol) in acetic acid (85 rnL) was added to a hot solution of 5-(3,5-di-tert-butyl-4-hydroxyphenyl)-3H-l,2-dithiole-3-thio ne 1 (1.16 g, 2.99 mmol) in acetic acid (85 mL) and the reaction heated under reflux for 2.5 h. The reaction was cooled to rt and concentrated. The residue was dissolved in hot ethyl acetate, filtered and allowed to crystallise to afford 5-(3,5-di-fert-butyl-4- hydroxyphenyl)-3H-l,2-dithiol-3-one 4 as a light brown solid, (450 mg, 47%); mp 223- 226 °C; ¹ H NMR (500 MHz, CDCl ₃ ) δ 1.47 (s, 18H, t-Bu x 2), 5.68 (s, 1H.0H), 6.75 (s, IH, CH), 7.44 (s, 2H, Ar); ¹³ C NMR (125 MHz ₅ CDCl ₃ ) δ 30.0 (C(CH ₃ ) ₃ ), 34.5(CH ₃ ), 116.0 (C4), 123.7 (C5), 124.2, 136.9, 157.2, 171.7 (Ar-4), 194.3 (CO); IR v 3514, 2954, 1632, 1549, 1116, 888, 734 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 323.1134, requires 323.1134 for C _n H ₂₃ O ₂ S ₂ .

5-(3,5-Di-te^-butyI-4-methoxyphenyl)-3iϊ-l,2-dithiol-3-o ne

A hot solution Of Hg(OAc) ₂ (2.18 g, 6.58 mmol) in acetic acid (85 niL) was added to a hot solution of the 5-(3,5-dDi-fert-butyl-4-methoxyphenyl)-3H-l,2-dithiol-3-thio ne 2 (1.16 g, 3.29 mmol) in acetic acid (85 mL) and the reaction was heated under reflux for 2.5 h. The reaction was cooled to rt and concentrated. The residue was dissolved in hot ethyl acetate, filtered and allowed to crystallise to afford 5-(3,5-di-ferf-butyl-4- methoxyphenyl)-3H-l,2-dithiol-3-one 5 as a light brown solid, (650 mg, 59%); mp 109- 112 ⁰ C; ¹ H NMR (500 MHz, CDCl ₃ ) δ 1.46 (s, 18H, t-Bu x 2), 3.74 (s, 3H, OMe), 6.77 (s, IH, CH), 7.50 (s, 2H, Ar); ¹³ C NMR (125 MHz, CDCl ₃ ) δ 31.8 (C(CH ₃ ) ₃ ), 36.0 (CH ₃ ), 64.1 (CH ₃ ), 116.9 (C4), 124.9 (C5), 127.2, 145.3, 162.9, 171.2 (Ar-4), 194.1 (CO); IR v 2959, 1658, 1545, 1407, 1215, 1006, 887 Cm- ¹ J HRMS ESI ⁺ [M+H] ⁺ = 337.1288, requires 337.1290 for Ci ₈ H ₂₅ O ₂ S ₂ .

3-(3,5-Di-tert-butyl-4-methoxymethoxyphenyl)-2-methyl-3-o xopropionic acid methyl ester

K ₂ CO ₃ (318 mg, 2.30 mmol) and methyl iodide (142 μL, 2.29 mmol) were added to a solution of 3-(3,5-di-ført-butyl-4-methoxymethoxyphenyl)-3-oxopropionic acid methyl ester (820 mg, 2.34 mmol) in methanol (5.0 mL) and the mixture heated under reflux overnight. The reaction mixture was then cooled to rt and concentrated. Ether was added and the organic layer washed with water (x 3), brine (x 1), dried (MgSO ₄ ) and concentrated. The residue was purified by flash chromatography (5% EtO Ac/petrol) to afford 3 -(3 ,5-di-tert-butyl-4-methoxymethoxyphenyl)-2-methyl-3 -oxopropionic acid methyl ester as a colourless oil (604 mg, 72%); ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.45 (s,

18H, t-Bu x 2), 1.48 (d, J= 6.8 Hz, 3H, CH(CH ₃ )), 3.64 (s, 3H, OMe), 3.69 (s, 3H, OMe), 4.36 (q, J= 6.8 Hz, IH, CH(CH ₃ )), 4.91 (s, 2H, CH ₂ ), 7.93 (s, 2H, Ar); ¹³ C NMR (IOO MHz, CDCl ₃ ) δ 13.8 (CH ₃ ), 31.6 (C(CH ₃ ) ₃ ), 35.8 (CH ₃ ), 48.0 (CH), 52.3 (OCH ₃ ), 57.3 (OCH ₃ ), 100.7 (CH ₂ ), 127.4, 130.3, 144.9, 159.4 (Ar-4), 171.4 (CO), 194.9 (CO); IR v 2952, 1742, 1680, 1591, 1159, 1076, 856 Cm ^-1 ; HRMS ESI ⁺ [M+Na] ⁺

= 387.2140, requires 387.2142 for C ₂₀ H ₃₃ ONa.

5-(3,5-Di-tor-butyl-4-hydroxyphenyl)-4-methyl-3H-l,2-dith iole-3-thione

P ₄ Si ₀ (950 mg, 2.14 mmol), sulfur (100 mg, 3.12 mmol) and HMDO (3.5 niL, 16 mmol) were added to 3-(3,5-di-tert-butyl-4-methoxymethoxyphenyl)-2-methyl-3- oxopropionic acid methyl ester (1.07 g, 2.94 mmol) in xylene (7.0 mL). The reaction mixture was heated under reflux for 1 h cooled to rt and the reaction mixture applied to silica gel and purified by flash chromatography (5% EtO Ac/petrol). The residue was recrystallised from EtOAc/petrol to afford 5-(3,5-di-ført-butyl-4-hydroxyphenyl)-4-

(methyl)-3H-l,2-dithiole-3-thione 6 as an orange solid, (600 mg, 81%); mp 174-175°C; ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.47 (s, 18H, t-Bu x 2), 2.24 (s, 3H, CH ₃ ), 5.61 (s, IH, OH), 7.30 (s, 2H, Ar); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 16.9 (CH ₃ ), 30.1 (C(CH ₃ ) ₃ ), 34.5 (CH ₃ ), 124.7 (C4), 125.7 (C5), 136.8, 140.8, 156.2, 170.2 (Ar-4), 215.4 (CS); IR v 3624, 2959, 2911, 1594, 1430, 1120, 885 cm ^'1 ; Microanalysis: Found C, 61.37; H, 6.91 ; S, 27.15. Ci ₈ H ₂₄ OS ₃ , requires C, 61.32; H, 6.86; S, 27.28%.

3-(3,5-Di-fer^butyI-4-methoxyphenyI)-2-methyl-3-oxopropio nic acid methyl ester

K ₂ CO ₃ (1.15 g, 8.32 mmol) and methyl iodide (515 μL, 8.31 mmol) were added to a solution of 3-(3,5-di-ter^butyl-4-methoxyphenyl)-3-oxopropionic acid methyl ester (2.66 g, 8.30 mmol) in methanol (20 mL) and the mixture was heated under reflux overnight. The reaction mixture was then cooled to rt and concentrated. Ether and water were added and the aqueous phase extracted with ether (x 2). The combined extracts were washed with water (x 2), brine (x 1), dried (MgSO ₄ ) and concentrated. The residue was purified by flash chromatography (5% EtOAc/petrol) to afford 3-(3,5-di-

tert-butyl-4-methoxyphenyl)-2-rnethyl-3-oxopropionic acid methyl ester as a colourless oil, (1.92 g, 69%); ¹ H NMR (500 MHz, CDCl ₃ ) δ 1.45 (s, 18H, f-Bu x 2), 1.50 (d, J= 7.0 Hz, 3H, CH ₃ ), 3.71 (s, 3H, OCH ₃ ), 3.73 (s, 3H, OCH ₃ ), 4.38 (q, J= 7.0 Hz, IH, CH), 7.94 (s, 2H, Ar); ¹³ C NMR (100 MHz ₅ CDCl ₃ ) δ 14.2 (CH ₃ ), 32.1 (C(CH ₃ ) ₃ ), 36.2 (CH ₃ ), 48.4 (CH), 52.6 (OCH ₃ ), 64.6 (OCH ₃ ), 127.8, 130.4, 144.6, 164.7(Ar-4), 171.8 (CO), 195.3 (CO); IR v 2956, 2873, 1740, 1681, 1373, 1114, 873 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 335.2217, requires 335.2217 for C ₂₀ H ₃₁ O ₄ .

5-(3,5-Di-førr-butyl-4-methoxyphenyl)-4-(methyl)-3Jϊ-l, 2-dithiole-3-thione

P ₄ Si ₀ (1.51 mg, 3.37 mmol), sulfur (202 mg, 6.30 mmol) and HMDO (7.3 mL, 34 mmol) were added to 3-(3,5-di-tert-butyl-4-methoxyphenyl)-2-methyl-3-oxopropioni c acid methyl ester (1.92 g, 5.74 mmol) in xylene (5.0 mL). The reaction was heated under reflux for 1.5 h. The reaction was then cooled to rt and concentrated. The reaction mixture was applied to silica gel and purified by flash chromatography (5% EtO Ac/petrol) and the residue recrystallised from petrol to afford 5-(3,5-di-tert~butyl-4- methoxyphenyl)-4-(methyl)-3H-l,2-dithiole-3-thione 7 as an orange solid, (2.07 g, 98%); mp 83-84 °C; ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.46 (s, 18H, t-Bu x 2), 2.23 (s, 3H, C=C(Me)), 3.76 (s, 3H, OMe), 7.35 (s, Ar, 2H); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 16.9 (CH ₃ ), 31.9 (C(CH ₃ ) ₃ ), 36.0 (CH ₃ ), 64.3 (OCH ₃ ), 127.1 (C4), 127.9 (C5), 141.2, 145.0, 161.8, 169.4 (Ar-4), 215.5 (CS); IR v 2961, 2869, 1525, 1307, 1223, 1007, 732 cm ^"1 ; Microanalysis: Found C, 62.34; H, 7.25. Ci ₉ H ₂₃ OS ₃ requires C, 62.25; H, 7.15%.

3-(3,5-Di-tert-butyl-4-ethoxyphenyl)-2-methyl-3-oxopropio nic acid methyl ester

K ₂ CO ₃ (384 mg, 2.78 mmol) and methyl iodide (172 μL, 2.15 mmol) were added to a solution of 3-(3,5-di-tgrt-butyl-4-ethoxyphenyl)-3-oxopropionic acid methyl ester (929 mg, 2.78 mmol) in methanol (10 mL) and the mixture was heated under reflux overnight. The reaction mixture was then cooled to rt and concentrated. Ether and water were then added and the aqueous phase extracted with ether (x 2). The combined extracts were washed with water (x 2), brine (xl) and dried (MgSO4). The residue was purified by flash chromatography (5% EtOAc/petrol) to afford 3-(3,5-di-tert-butyl-4- ethoxyphenyl)-2-methyl-3-oxopropionic acid methyl ester as a colourless oil (632 mg, 65%); ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.41 (s, 18H, t-Bu x 2), 1.41 (t, J= 6.8 Hz, 3H, CH ₃ CH ₂ O), 1.47 (d, J= 7.2 Hz, 3H, CH(CH ₃ )), 3.68 (s, 3η, OMe), 3.76 (q, J= 6.8 Hz, 2H, CH ₂ O), 4.36 (q, J= 7.2 Hz, IH, CH(CH ₃ ), 7.91 (s, 2H, Ar); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 13.9 (CH ₃ ), 14.8 (CH ₃ ), 31.7 (C(CH ₃ ) ₃ ), 35.9 (CH ₃ ), 48.0 (CH), 52.3 (OCH ₃ ), 72.0 (CH ₂ ), 127.6, 129.9, 144.2, 162.9 (Ar-4), 171.5 (CO), 195.0 (CO); IR v 2960, 2904, 2873, 1742, 1678, 1209, 1032, 887 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 349.2373, requires 349.2373 for C ₂ iH ₃₃ O ₄ .

5-(3,5-Di-tert-butyI-4-ethoxyphenyl)-4-(methyl)-3H-l,2-di thiole-3-thione

P ₄ Si ₀ (484 mg, 1.09mmol), sulfur (64 mg, 2.00 mmol) and HMDO (2.3 mL, 11 mmol) were added to 3-(3,5-di-ter^butyl-4-ethoxyphenyl)-2-methyl-3-oxopropionic acid methyl ester (632 mg, 1.81 mmol) in xylene (3.0 mL). The reaction was heated under _^ reflux for 2 h then cooled to rt. The reaction mixture was applied to silica gel and purified by flash chromatography (5% EtO Ac/petrol) and the residue recrystallised from petrol to afford the 5-(3,5~di-fer^butyl-4-ethoxyphenyl)-4-(methyl)-3H-l,2- dithiole-3-thione 8 as an orange solid, (480 mg, 70%); mp 83-84 ⁰ C; ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.44 (bs, 2 IH, t-Bu x 2, CH ₃ CH ₂ O), 2.23 (s, 3H, CH ₃ ), 3.83 (q, 2H, CH ₃ CH ₂ O), 7.34 (s, 2η, Ar); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 14.9 (CH ₃ ), 16.8 (CH ₃ ),

31.9 (C(CHs) ₃ ), 36.1 (CH ₃ ), 72.1(CH ₂ ), 126.8 (C4), 127.6 (C5), 141.2, 145.0, 160.3, 169.5 (Ar-4), 215.6 (CS); IRv 2956, 1522, 1425, 1383, 1217, 1084, 886 cm ^"1 ; HRMS

ESI ⁺ [M+H] ⁺ = 381.1375, requires 381.1375 for C ₂₀ H ₂₉ OS ₃ ; Microanalysis: Found C, 63.15; H, 7.50; S, 25.11. C ₂₀ H ₂₈ OS ₃ requires C, 63.11; H, 7.41; S, 25.27%.

5-(3,5-Di-ter/-butyl-4-hydroxyphenyl)-4-(methyl)-3/-'-l,2 -dithiole-3-one

A hot solution of 5-(3,5-di-terr-butyl-4-hydroxyphenyl)-4-(methyl)-3H-l,2-dith iole-3- thione 6 (456 mg, 1.25 mmol) in acetic acid (1OmL) was added to a hot solution of Hg(OAc) ₂ (797 mg, 2.50 mmol) in acetic acid (10 mL) and the reaction mixture heated under reflux for 1.5 h. The reaction was cooled to rt and filtered. The filtrate was concentrated and the residue was purified by flash chromatography (10% EtO Ac/petrol) to afford 5-(3,5-di-^rt-butyl-4-hydroxyphenyl)-4-(methyl)-3H-l,2-dithi ole-3-one 9 as a colourless solid, (257 mg, 61%); mp 189-190 °C; ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.47 (s, 18H, t-Bu x 2), 2.06 (s, 3H, CH ₃ ), 5.56 (s, IH, OH), 7.26 (s, 2H, Ar); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 14.1 (CH ₃ ), 30.1 (C(CH ₃ ) ₃ ), 34.5 (CH ₃ ), 125.2 (C4), 125.7 (C5), 136.6, 155.9, 164.3 (Ar-4), 195.4 (CO); IR v 3532, 2951, 1623, 1556, 1432, 1113, 950, 658 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 337.1292, requires 337.1290 for C ₁₈ H ₂₅ O ₂ S ₂ ; Microanalysis: Found C, 64.31; H, 7.22, C ₁₈ H ₂₄ O ₂ S ₂ requires C, 64.25; H, 7.19%.

5-(3,5-Di-te^-butyl-4-methoxyphenyl)-4-(methyl)-3iϊ-l,2- dithioIe-3-one

A hot solution Of Hg(OAc) ₂ (829 mg, 2.60 mmol) in acetic acid (35 mL) was added to a hot solution of 5-(3,5-di-teτ-r-butyl-4-methoxyphenyl)-4-(methyl)-3H-l,2-di thiole-3- thione 7 (500 mg, 1.3 mmol) in acetic acid (35 mL) and the reaction mixture was heated

under reflux for 1 h. The reaction was cooled to rt and filtered. The filtrate was concentrated and the residue purified by flash chromatography (10% EtO Ac/petrol). The residue was recrystallised from EtOH to afford the 5-(3,5-di-tert-butyl-4- methoxyphenyl)-4-(methyl)-3/f-l,2-dithiole-3-one 10 as a colourless solid (370 mg, 79%); mp 112-114 °C; ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.45 (s, 18H, t-Bu x 2), 2.06 (s, 3H, CH ₃ ), 3.75 (s, 3H, OMe), 7.32 (s, 2H, Ar); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 14.1 (CH ₃ ), 31.9 (C(CH ₃ ) ₃ ), 36.0 (CH ₃ ), 64.4 (OCH ₃ ), 126.2 (C4), 126.6 (C5), 128.3, 144.8, 161.5, 163.8 (Ar-4), 195.3 (CO); IR v 2961, 2870, 1590, 1032 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 351.1447, requires 351.1447 for Ci ₉ H ₂₇ O ₂ S ₂ .

l-(3,5-Diisopropyl-4-methoxymethoxyphenyl)ethanone

Acetyl chloride (370 μL, 5.16 mmol) was added dropwise to a solution of dimethoxymethane (460 μL, 5.16 mmol), and ZnCl ₂ (3 mg, 0.037 mmol) in toluene (2 mL) and the reaction stirred at rt for 4 h. l-(4-Hydroxy-3,5-diisopropylphenyl)ethanone (380 mg, 1.72 mmol) was added, followed by N-ethyl-iV-(l-methylethyl)-2- propanamine (450 μL, 2.60 mmol) and the reaction was stirred at rt overnight. Water (15 mL) was added and the mixture stirred for 15 min. The aqueous layer was extracted with ether (x 2) and the combined organic extracts washed with water (x 2), brine (x 1), dried and concentrated. The residue was purified by flash chromatography (10% EtO Ac/petrol) to afford l-(3,5-diisopropyl-4-methoxymethoxyphenyl)ethanone as a yellow oil (206 mg, 37%); ¹ H NMR (500 MHz, CDCl ₃ ) δ 1.26 (d, J= 7 Hz, 12H, CH(CH ₃ ) ₂ x 2), 2.59 (s, 3H, COCH ₃ ), 3.36 (septet, J= 7.0 Hz, 2H, (CH ₃ ) ₂ C// x 2), 3.62 (s, 3H, OMe), 4.96 (s, 2H, OCH ₂ O), 7.73 (s, 2H, Ar); HRMS ESI ⁺ [M+H] ⁺ = 265.1798, requires 256.1798 for Ci ₆ H ₂₅ O ₃ .

5-(3,5-DiisopropyI-4-methoxymethoxyphenyl)-3H-l,2-dithiol e-3-thione

l-(3,5-Diisopropyl-4-methoxymethoxyphenyl)ethanone (1.39 g, 5.26 mmol) in THF (7.6 mL) was added dropwise to a suspension of KH (1.27 g, 11.0 mmol, 35% dispersion in mineral oil) in THF (12 mL) and DMPU (5.8 mL). The resulting suspension was stirred for 15 min. A solution of carbon disulfide (350 μL, 5.78 mmol) in THF (3.3 mL) and DMPU (1.6 mL) was then added and stirred for a further 10 min. HMDT (1.7 mL, 7.89 mmol) was then added and stirring was continued for 20 min. The reaction mixture was then cooled to O ⁰ C and a solution of hexachloroethane (1.24 g, 5.26 mmol) in THF (3.3mL) added and stirring continued for 30 min. Methanol was added and the reaction was stirred for 15 min. The solvent was removed and the residue dissolved in dichloromethane and purified by flash chromatography (10% EtO Ac/petrol). The residue was recrystallised from petrol to afford 5-(3,5-diisopropyl- 4-methoxymethoxyphenyl)-3H-l,2-dithiole-3-thione as an orange solid (1.30 g, 70%); mp 71-72 °C; ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.25 (d, J= 7 Hz, 12H, CH(C/f ₃ ) ₂ x 2),

3.37 (septet, J= 7.0 Hz, 2H, (CH ₃ ) ₂ CH x 2), 3.62 (s, 3H, OCH ₃ ), 4.97 (s, 2H, OCH ₂ ),

7.38 (s, 2H, Ar), 7.43 (s, IH, CH); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 23.7 (CH ₃ ), 27.0 (CH(CHs) ₂ ), 57.5 (OCH ₃ ), 100.5 (CH ₂ ), 123.1 (C4), 128.2 (C5), 135.5, 143.7, 156.0,

173.6 (Ar-4), 215.3 (CS); IR v 2962, 1512, 1456, 1262, 1159, 1097, 1034, 884, 666 cm- '; HRMS ESI ⁺ [M+H] ⁺ = 355.0853, requires 355.0855 for C ₁₇ H ₂₃ O ₂ S ₃ ; Microanalysis: Found C, 58.06; H, 5.79; S, 31.04% C _n H ₂₂ O ₂ S ₃ requires C, 58.02; H, 5.84; S, 30.98%

5-(3,5-DiisopropyI-4-hydroxyphenyl)-3H-l,2-dithiole-3-thi one

A mixture of 5-(3,5-diisopropyl-4-methoxymethoxyphenyl)-3/f-l,2-dithiole- 3-thione (903 mg, 2.55 mmol) and TFA (6.1 niL) in dichloromethane (30 niL) was stirred at rt for 2 h. The solvent was evaporated and the residue recrystallised from EtO Ac/petrol to afford 5-(3,5-diisoρropyl-4-hydroxyphenyl)-3/i-l,2-dithiole-3-thio ne 11 as a dark brown solid (606 mg, 77%); mp 154-155 °C; ¹ H NMR (400 MHz, DMSO) δ 1.29 (s, 12H, CH(CH ₃ )Z ^x 2), 3.10 (septet, J= 6.4 Hz, 2H, CH(CH ₃ ) ₂ ), 5.48 (bs, IH, OH), 7.35 (s, 2H, Ar), 7.43 (s, IH, CH); ¹³ C NMR (100 MHz, DMSO) δ 22.5 (CH ₃ ), 27.2 (CH(CH ₃ ) ₂ ), 122.8 (C4), 124.1 (C5), 134.4, 135.1, 154.2, 174.5 (Ar-4), 214.8 (CS); IR v 3197, 2958, 1594, 1507, 1286, 1030, 771 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 311.0592, requires 311.0593 Ci ₅ Hi ₉ OS ₃ ; Microanalysis: Found C, 57.59; H, 6.17; S, 27.21%. C ₅ H ₂₂ O ₂ S ₃ requires C, 57.49; H, 6.25; S, 27.13%.

3-(3,5-Diisopropyl-4-methoxyphenyl)-3-oxopropionic acid methyl ester

l-(3,5-Diisopropyl-4-methoxy-phenyl) ethanone (836 mg, 3.50 mmol) in THF (4.0 mL) was added dropwise over 2 h to a refluxing solution of washed (hexane) sodium hydride (700 mg, 17.5 mmol, 60% dispersion in mineral oil) and dimethyl carbonate (1.5 mL, 17.5 mmol) in THF (8.0 mL). The reaction was heated for a further 15 min then cooled to rt. Water (20 mL) was added followed by ether. The organic extract was washed with water (x 3), sat. NaHCO ₃ (x 1), brine (x 1), dried (MgSO ₄ ) and concentrated. The residue was purified by flash chromatography (10% EtO Ac/petrol) to afford 3 -(3, 5- diisopropyl-4-methoxyphenyl)-3-oxopropionic acid methyl ester as a yellow oil (595

mg, 58%); ( ¹ H NMR 400 MHz, CDCl ₃ ) δ 1.25 (d, J= 7.0 Hz, 12H, (CHs) ₂ CH x 2 (keto)), 3.36 (septet, J= 7.0 Hz, 2H, (CH ₃ ) ₂ CH x 2 (keto)), 3.75 (s, 3η, OMe (keto)), 3.75 (s, 3H, OMe (enol)), 3.77 (s, 3H, OMe (keto)), 3.79 (s, 3H, OMe (enol)), 3.99 (s, 2H, CH ₂ ), 5.63 (s, IH, CH (enol)), 7.52 (s, 2H, Ar (enol)), 7.65 (s, 2H, Ar (keto)), 12.53 (s, IH, OH); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 23.77 (CH ₃ ), 23.83 (CH ₃ ), 26.56

(CH(CH ₃ ) ₂ ), 26.59 (CH(CH ₃ ) ₂ ), 45.8 (CH ₂ ), 51.3 (OCH ₃ ), 52.3 (OCH ₃ ), 62.2, 86.1, 122.3, 125.1, 129.4, 132.3, 142.1, 142.5, 157.5, 159.6, 168.1, 172.0 (Ar-8), 173.5 (CO), 191.7 (CO); IR v 2963, 2872, 1743, 1682, 800 cm ^'1 ; HRMS ESI ⁺ [M+Na] ⁺ = 293.1745 requires 293.1747 for C ₁₇ H ₂₅ O ₄ .

5-(3,5-Diisopropyl-4-methoxyphenyl)-3H-l,2-dithiole-3-thi <me

P ₄ S ₁₀ (542 mg, 1.22 mmol), sulfur (72 mg, 2.424 mmol) and HMDO (2.6 rnL, 12 mmol) were added to a solution of 3-(3,5-diisopropyl-4-methoxyphenyl)-3~ oxopropionic acid methyl ester (595 mg, 2.04 mmol) in xylene (3 mL). The reaction was heated under reflux for 1 h then cooled to rt. The reaction mixture was applied to silica gel and purified by flash chromatography (10% EtO Ac/petrol) and the residue recrystallised from EtO Ac/petrol to afford 5-(3,5-diisopropyl-4-methoxyphenyl)-3H- l,2-dithiole-3-thione 12 as an orange solid mp 150-151 ⁰ C; (407 mg, 61%); ¹ H NMR (500 MHz, CDCl ₃ ) δ 1.26 (d, J= 7 Hz, 12H, CH(CH ₃ ) ₂ x 2), 3.35 (septet, J= 7.0 Hz, 2H, (CH ₃ ) ₂ CH ^χ 2), 3.78 (s, 3η, OMe), 7.37 (s, 2H, Ar), 7.43 (s, IH, CH); ¹³ C NMR (125 MHz, CDCl ₃ ) δ 23.8 (CH ₃ ), 26.7 (CH(CH ₃ ) ₂ ), 62.3 (CH ₃ ), 123.2 (C4), 127.9 (C5), 135.4, 143.6, 158.4, 173.8 (Ar-4), 215.3(CS); IR v 3045, 2958, 1595, 1500, 1383, 1163, 1001, 885, 670 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 325.0749, requires 325.0750 for C ₁₆ H ₂₁ OS ₃ .

3-(4-Ethoxy-3,5-diisopropylphenyl)-3-oxopropionic acid methyl ester

l-(4-Ethoxy-3,5-diisopropyl-phenyl) ethanone (772 mg, 3.11 mmol) in THF (3.0 mL) was added dropwise over 2 h. to a refluxing solution of washed (hexane) sodium hydride (622 mg, 17.5 mmol, 60% dispersion in mineral oil) and dimethyl carbonate (1.3 mL, 16 mmol) in THF (7.0 mL). The reaction was heated for a further 15 min then cooled to rt. Water was then added followed by ether. The organic extract was washed with water (x 3) sat. NaHCO ₃ (x 1), brine (x 1), dried (MgSO ₄ ) and concentrated. The residue was purified by flash chromatography (10% EtO Ac/petrol) to afford 3-(4- ethoxy-3,5-diisopropylphenyl)-3-oxopropionic acid methyl ester as a yellow oil (600 mg, 64%); (400 MHz, CDCl ₃ ) δ 1.20 (d, J= 6.8 Hz, 12H, (CH ₃ ) ₂ CH ), 1.42 (t, J= 7.0 Hz, OCH ₂ CH ₃ ), 3.28 (septet, J= 7.0 Hz, 2H, (CH ₃ ) ₂ CH x 2), 3.71 (s, 3η, OMe), 3.80 (q, J= 7.0 Hz ₅ OCH ₂ CH ₃ ) 3.95 (s, 2H, CH ₂ ), 7.68 (s, 2H, Ar); IR v 2963, 1743, 1682, 1622, 1461, 1290, 1163, 800 cm ^"1 ; HRMS ESI ⁺ [M+Na] ⁺ = 307.1904, requires C ₁₈ H ₂₇ O ₄ .

5-(3,5-Diisopropyl-4-ethoxyphenyl)-3 _J fiT-l,2-dithiole-3-thione

P ₄ S ₁₀ (533 mg, 1.20 mmol), sulfur (71 mg, 2.21 mmol) and HMDO (2.6 mL, 12 mmol) were added to a solution 3-(4-ethoxy-3,5-diisopropylphenyl)-3-oxopropionic acid methyl ester (600 mg, 2.00 mmol) in xylene (3.0 mL). The reaction was heated under reflux for 1 h then cooled to rt. The reaction mixture was applied to silica gel and purified by flash chromatography (10% EtO Ac/petrol). The residue was recrystallised from EtO Ac/petrol to afford 5-(3,5-Diisopropyl-4-ethoxyphenyl)-3H-l,2-dithiole-3-

thione 13 as an orange solid, (250 mg, 37%); mp 96-97 ⁰ C; ¹ H NMR (500 MHz, CDCl ₃ ) δ 1.25 (d, J= 7 Hz, 12H, CH(CH ₃ ) ₂ x 2), 1.47 (t, J= 7.0 Hz ₅ OCH ₂ CH ₃ ), 3.33 (septet, J= 7.0 Hz, 2H ₅ (CH ₃ ) ₂ CH x 2), 3.84 (q, J= 7.0 Hz, OCH ₂ CH ₃ ), 7.37 (s, 2H, Ar), 7.43 (s, IH ₅ CH); ¹³ C NMR (125 MHz, CDCl ₃ ) δ 15.7, 26.7, 70.7, 123.1, 127.8, 135.3, 143.7, 157.3, 173.9, 215.2; IR v 304O ₅ 2960, 2924, 2865, 1508, 1331, 1028, 1105, 777 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 339.0904, requires 339.0906 for Ci ₇ H ₂₃ OS ₃ .

l-(4-Methoxymethoxy-3,5-dimethylphenyl) ethanone

Acetyl chloride (640 μL, 707 mg, 9.00 mmol) was added dropwise to a solution of dimethoxymethane (800 μL, 685 mg, 9.00 mmol) and ZnCl ₂ (3 mg, 0.037 mmol) in toluene (3 mL) and the reaction was stirred at rt for 4 h. l-(4-Hydroxy-3,5- dimethylphenyl) ethanone (500 mg, 3.00 mmol) was then added followed by iV-ethyl-N- (l-methylethyl)-2-propanamine (783 μL, 582 mg, 4.50 mmol) and the reaction was stirred at rt overnight. Water (10 mL) was added and stirred for 15 min. The aqueous layer was extracted with ether (x 2) and the combined organic extracts washed with water (x 3), brine (x 1), dried and concentrated. The residue was purified by flash chromatography to afford l-(4-methoxymethoxy-3,5-dimethylphenyl) ethanone as a coulorless oil (427 mg, 68%); ¹ H νMR (500 MHz, CDCl ₃ ) δ 2.25 (s, 6H, Me x 2), 2.46 (s, 3H, COCH ₃ ), 3.52 (s, 3H, OMe), 4.92 (s, 2H ₅ OCH ₂ O), 7.56 (s, 2H ₅ Ar); ¹³ C νMR (100 MHz, CDCl ₃ ) δ 16.7, 26.1, 57.1, 98.8, 129.1, 131.1, 133.0, 158.9, 197.0; IR v 2925, 1677, 1596, 1155, 959, 769 Cm ^-1 ; HRMS ESI ⁺ [M+νa] ⁺ = 209.1170, requires 209.1172 for Ci ₂ H] ₇ O ₃ Na

5-(3,5-Dimethyl-4-methoxymethoxyphenyI)-3H-l,2-dithioIe-3 -thioαe

l-(4-Methoxymethoxy-3,5-dimethylphenyl) ethanone (1.06 g, 5.23 mmol) in THF (7.6 mL) was added dropwise to a suspension of KH (1.20 g, 10.5 mmol, 35% dispersion in mineral oil) in THF (11 mL) and DMPU (5.5 mL). The resulting suspension was then stirred for 15 min. A solution of carbon disulfide (440 μL, 5.76 mmol) in THF (3.3 mL) and DMPU (1.6 mL) was then added and the solution stirred for a further 10 min. HMDT (1.7 mL, 7.85 mmol) was then added and stirring was continued for 20 min. The reaction mixture was then cooled to O ⁰ C and a solution of hexachloroethane (1.24 g, 5.23 mmol) in THF (3.3 mL) was added and stirring was continued for 30 min. The solvent was removed and the residue purified by flash chromatography (20%

EtOAc/petrol) The residue was recrystallised from EtOAc/petrol to afford 5-(3,5- dimethyl-4-methoxymethoxyphenyl)-3/f-l,2-dithiole-3-thione as an orange solid (777 mg, 50%); mp 91-96 ⁰ C; ¹ H NMR (500 MHz, CDCl ₃ ) δ 2.34 (s, 6H, Me x 2), 3.62 (s, 3H, OMe) ₃ 5.01 (s, IH, OCH ₂ O), 7.32 (s, 2H, Ar), 7.37 (s, IH, CH); ¹³ C NMR (125 MHz, CDCl ₃ ) δ 17.0, 57.5, 99.1, 127.4, 127.5, 132.8, 135.3, 158.6, 172.9, 215.3; IR v 2951, 2923, 1509, 1473, 1272, 1061, 957, 665 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 299.0227, requires 299.0229 for C ₁₃ Hj ₅ O ₂ S ₃ .

5-(3,5-Dimethyl-4-hydroxyphenyl)-3iϊ-l,2-dithiole-3-thio ne

A mixture of 5-(3,5-dimethyl-4-methoxymethoxyphenyl)-3/f-l,2-dithiole-3-t hione (500 mg, 1.68 mmol) and TFA (4.1 mL) in dichloromethane (20 mL) was stirred at rt for 1 h. The solvent was evaporated and the residue recrystallised from acetone to afford 5-(3,5- dimethyl-4-hydroxyphenyl)-3/iT-l,2-dithiole-3-thione 14 as a dark orange solid (250 mg, 58%); mp 222-223 °C; ¹ H NMR (500 MHz, d ₆ -DMSO) δ 2.20 (s, 6H, Me x 2), 7.53 (s, 2H, Ar) 7.66 (s, IH, CH), 9.33 (s, IH, OH); ¹³ C NMR (100 MHz, d ₆ -DMSO) δ 16.4 (CH ₃ ), 122.1 (C4), 125.4 (C5), 127.5, 133.3, 157.9, 174.7 (Ar-4), 214.1 (CS); IR v 3438, 1569, 1500, 1295, 1165, 1077, 940, 696 cm ^"1 ; HRMS ESI ^" [M-H] ^" = 252.9818,

requires 252.9821 C ₁₁ H ₉ OS _3, ; Microanalysis: Found C, 57.59; H, 6.17; S, 27.21%. C ₁₅ H ₂₂ O ₂ S ₃ requires C, 57.49; H, 6.25; S, 27.13%.

l-(4-Methoxy-3,5-dimethylphenyl) ethanone

Dimethyl sulfate (2.8 mL, 29.1 mmol) and K ₂ CO ₃ (2.3 g, 16.7 mmol) were added to a solution of l-(4-hydroxy-3,5-dimethylphenyl) ethanone (1.60 g, 9.7 mmol) in acetone (10 mL) and the mixture was heated at reflux for 3 h. The solution was cooled and ether was added. The organic phase was washed with water (x 2), brine (x 1), dried (MgSO ₄ ) and concentrated. The residue was purified by flash chromatography (5% EtOAc/pertol) to afford l-(4-methoxy-3,5-dimethylphenyl)ethanone as a colourless oil (1.19 g, 70%); ¹ H NMR (400 MHz, CDCl ₃ ) δ 2.21 (s, 6H, Me x 2), 2.44 (s, 3H, CH ₃ ), 3.65 (s, 3H, OMe), 7.53 (s, 2H, Ar); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 15.9 (CH ₃ ), 26.1 (CH ₃ ), 59.2 (OCH ₃ ), 129.0, 130.7, 132.5, 160.9 (Ar-4), 197.1 (C=O); IR v 2940, 1674, 1591, 1482, 1092, 874 cm ^"1; HRMS ESI ⁺ [M+H] ⁺ = 179.1066, requires 179.1067 for CπHi ₅ θ ₂ .

3-(4-Methoxy-3,5-dimethylphenyl)-3-oxopropionic acid methyl ester

l-(4-Methoxy-3,5-dimethylphenyl) ethanone (1.19 g, 5.04 mmol) in THF (5 mL) was added dropwise over 2 h to a refluxing solution of washed (hexane) sodium hydride (1.33 g, 33.3 mmol, 60% dispersion in mineral oil) and dimethyl carbonate (2.8 mL, 33.3 mmol) in THF (8 mL). The reaction was heated for a further 30 min then cooled to it. Water (50 mL) was then added followed by ether. The organic extract was washed with water (x 3), sat. NaHCO ₃ , brine (x 1), dried (MgSO ₄ ) and concentrated. The

residue was purified by flash chromatography (10% EtO Ac/petrol) to afford 3-(4- methoxy-3,5-dimethylphenyl)-3-oxopropionic acid methyl ester as a yellow oil (935 mg, 60%); ¹ H NMR (500 MHz, CDCl ₃ ) δ 2.24 (s, 6H, Me (enol) x 2), 2.25 (s, 6H, Me (keto) x 2), 3.68 (s, 3H, CO ₂ Me (enol)), 3.69 (s, 3H, CO ₂ Me (keto)), 3.70 (s, 3H, OMe (keto)), 3.72 (s, 3H, OMe (enol)), 3.90 (s, 2H, CH ₂ ), 5.54 (s, IH, CH), 7.38 (s, 2H, Ar (enol)), 7.56 (s, 2H, Ar (keto)), 12.46 (s, IH, OH), ¹³ C NMR (100 MHz, CDCl ₃ ) δ 16.0 (CH ₃ ), 45.2 (CH ₂ ), 52.1 (OCH ₃ ), 59.4 (OCH ₃ ), 85.9 (CH), 126.6, 129.4, 130.9, 131.2, 131.4, 161.6, 167.9 (Ar-8), 171.4 (CO), 191.4 (CO); IR v 2952, 1744, 1680, 1597, 1137, 1004, 889 cm ^'1 ; HRMS ESI ⁺ [M+Na] ⁺ = 259.0938, requires 259.0941 Cj ₃ H ₁₆ NaO ₄

5-(3,5-Dimethyl-4-methoxyphenyl)-3H ^r -l,2-dithiole-3-thione

P ₄ Si ₀ (1.07 g, 2.41 mmol), sulfur (141 mg, 4.40 mmol) and HMDO (5.2 mL, 3.9 g, 24 mmol) were added to a solution of 3-(4-methoxy-3,5-dimethylphenyl)-3-oxopropionic acid methyl ester (935 mg, 4 mmol) in xylene (3 mL). The reaction was heated under reflux for 1.5 h. The reaction was then cooled to rt and reaction mixture applied to silica gel and purified by flash chromatography (5 to 15% EtO Ac/petrol) and the residue recrystallised from EtOAc/ petrol to afford 5-(3,5-dimethyl-4-methoxyphenyl)-3/f-l,2- dithiole-3-thione 15 as an orange solid mp 96-99 °C; (576 mg, 53%); ¹ H NMR (500 MHz, CDCl ₃ ) δ 2.33 (s, 6H, Me x 2), 3.77 (s, 3H, OMe), 7.32 (s, 2H, Ar), 7.37 (s, IH, CH); ¹³ C NMR (125 MHz, CDCl ₃ ) δ 16.2 (CH ₃ ), 59.8 (OCH ₃ ), 127.1 (C4), 127.5 (C5), 132.5, 135.3, 160.6, 173.0 (Ar-4), 215.3 (CS); IR v 2937, 1599, 1470, 1181, 1064, 832 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 269.0125, requires 269.1023 for C ₁₂ H ₁₃ OS ₃ .

l-(4-Ethoxy-3,5-dimethylphenyI) ethanone

Ethyl iodide (1.5 niL, 18 mmol) and K ₂ CO ₃ (2.53 g, 10.4 mmol) were added to a solution of l-(4-hydroxy-3,5-dimethylphenyl) ethanone (Ig, 6.10 mmol) in acetone (10 mL) and the mixture was heated at reflux for 3 h. The solution was cooled and ether was added. The organic phase was washed with water (x 2), brine (x 1), dried (MgSO ₄ ) and concentrated. The residue was purified by flash chromatography (5% EtOAc/pertol) to afford l-(4-ethoxy-3,5-dimethylphenyl) ethanone as a colourless oil, (94% 1.08 g); (400 MHz, CDCl ₃ ) δ 1.30 (t, J= 6.4 Hz, 3H, CH ₂ CH ₃ ), 2.27 (s, 6H, Me x 2), 2.50 (s, 3H, COCH ₃ ), 3.84 (q, J= 6.4 Hz, 2H, CH ₂ CH ₃ ), 7.59 (s, 2H, Ar); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 15.6 (CH ₃ ), 16.3 (CH ₃ ), 26.3 (CH ₃ ), 67.8 (CH ₂ ), 129.1, 131.0, 132.5, 160.3 (Ar-4), 197.4 (CO); IR v 2980, 2929, 1677, 1306, 1030, 900, 777 cm ^'1 ; HRMS ESI ⁺ [M+H] ⁺ = 193.1223, requires 193.1223 for Ci ₂ H ₁₇ O ₂ .

3-(4-Ethoxy-3,5-dimethyl-phenyl)-3-oxopropionic acid methyl ester

l-(4-Ethoxy-3,5-dimethylphenyl) ethanone (1.09 g, 5.67 mmol) in THF (5 mL) was added dropwise over 2 h to a refluxing solution of washed (hexane) sodium hydride (1.13 g, 28.3 mmol, 60% dispersion in mineral oil) and dimethyl carbonate (2.4 mL, 28 mmol) in THF (8 mL). The reaction was heated for a further 15 min then cooled to rt. Water (50 mL) was added followed by ether. The organic extract was washed with water (x 3) sat. NaHCO ₃ (x 1), brine (x 1), dried (MgSO ₄ ) and concentrated. The residue was purified by flash chromatography (10% EtO Ac/petrol) to afford 3-(4- ethoxy-3,5-dimethyl-phenyl)-3-oxopropionic acid methyl ester as a brown oil (971 mg, 68%); (400 MHz, CDCl ₃ ) δ 1.40 (t, J= 6.8 Hz, 3H, CH ₂ CH ₃ (keto)), 2.27 (s, 6η, Me x

2 (end)), 2.29 (s, 6H, Me x 2 (keto)), 3.72 (s, 3H, OMe (keto)), 3.76 (s, 3H, OMe (enol)), 3.88 (q, J= 6.8 Hz, 2H, CH ₂ CH ₃ ), 3.93 (s, 3H, OMe (keto)), 5.57 (s, IH, CH (enol)), 7.42 (s, 2H, Ar (enol)), 7.59 (s, 2H, Ar (keto)); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 15.6, 16.3 (enol), 16.4, 45.4, 51.2 (enol), 52.3, 67.9 (enol), 68.0, 86.0, 126.6, 128.3 (enol), 129.4, 131.2 (enol), 131.3, 131.5, 158.9 (enol), 161.0, 168.1, 171.6 (enol), 173.5 (enol), 191.6; IR v 2980, 2956, 1746, 1680, 1483, 1140, 1108, 987, 662 cm ^'1 ; HRMS ESI ⁺ [MHhNa] ⁺ = 273.1089, requires 273.1097 for C ₁₄ H ₁₈ NaO ₄ .

5-(3,5-Dimethyl-4-ethoxyphenyl)-3Jϊ-l,2-dithiole-3-thion e

P ₄ Si ₀ (1-03 g, 2.33 mmol), sulfur (137 mg, 4.27 mmol) and HMDO (5 niL, 23 mmol) were added to a solution 3-(4-ethoxy-3,5-dimethyl-phenyl)-3-oxopropionic acid methyl ester (971 mg, 3.88 mmol) in xylene (3 mL). The reaction was heated under reflux for 1 h then cooled to rt. The reaction mixture was applied to silica gel and purified by flash chromatography (10% EtO Ac/petrol). The residue was recrystallised from EtO Ac/petrol to afford 5-(3,5-dimethyl-4-ethoxyphenyl)-3H-l,2-dithiole-3-thione 16 as an orange solid, (709 mg, 65%); mp 110-112 °C; ¹ H NMR (500 MHz, CDCl ₃ ) δ 1.41 (t, 3H, OCH ₂ CH ₃ ), 2.29 (s, 6η, Me x 2), 3.86 (q, 2H, OCH ₂ CH ₃ ), 7.28 (s, 2H, Ar), 7.34 (s, IH, CH); ¹³ C NMR (125 MHz, CDCl ₃ ) δ 15.7 (CH ₃ ), 16.4 (CH ₃ ), 68.2 (CH ₂ ), 126.8 (C4), 127.5 (C5), 132.7, 135.1, 159.8, 173.1 (Ar-4), 215.2 (CS); IR v 2972, 2920, 1596, 1471, 1165, 1062, 900 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 283.0280, requires 283.0280 for C ₁₃ H ₁₅ OS ₃ .

5-(3,5-Dimethyl-4-methoxyphenyl)-3Jy-l,2-dithiole-3-one

A hot solution of Hg(OAc) ₂ (570 mg, 1.79 mmol) in acetic acid (15 niL) was added to a hot solution of 5-(3,5-dimethyl-4-methoxyphenyl)-3H-l,2-dithiole-3-thione (240 mg, 0.89 mmol) in acetic acid (15 mL) and the reaction was heated under reflux for 1.5 h. The reaction was cooled to rt and filtered. The filtrate was concentrated and the residue purified by flash chromatography (10% EtO Ac/petrol). The residue was recrystallised from EtO Ac/petrol to afford 5-(3,5-dimethyl-4-methoxyphenyl)-3H-l ,2-dithiole-3-one 17 as a colourless solid (108 mg, 48%); mp 95-96 °C; ¹ H NMR (500 MHz, CDCl ₃ ) δ 2.34 (s, 6H, Me x 2), 3.77 (s, 3H, OMe), 6.76 (s, IH, CH), 7.29 (s, 2H, Ar); ¹³ C NMR (125 MHz, CDCl ₃ ) δ 16.2 (CH ₃ ), 59.8 (OCH ₃ ), 117.0 (C4), 127.1 (C5), 128.1, 132.3, 160.2, 170.2 (Ar-4), 194.1 (CO); IR v 2924, 1659, 1546, 1237, 1001, 859 cm ^"1 _; HRMS ESI ⁺ [M+H] ⁺ = 253.0351, requires 253.0351 for C ₁₂ H ₁₃ O ₂ S ₂ .

3-(4~Benzyloxy-3,5-dimethyIphenyl)-3-oxopropionic acid methyl ester

l-(4-Benzyloxy-3,5-dimethyl-ρhenyl)-ethanone (670 mg, 2.64 mmol) in THF (3 mL) was added dropwise over 2 h to a refluxing solution of washed (hexane) sodium hydride (530 mg, 13 mmol, 60% dispersion in mineral oil) and dimethyl carbonate (1.3 mL, 13 mmol) in THF (4 mL). The reaction was heated for a further 15 min then cooled to rt. Water (20 mL) was then added followed by ether. The organic extract was washed with water (x3), sat. NaHCO ₃ (xl), brine (x 1) dried (MgSO ₄ ) and concentrated. The residue was purified by flash chromatography (10% EtO Ac/petrol) to afford 3~(4-benzyloxy- 3,5-dimethylphenyl)-3-oxopropionic acid methyl ester as a yellow oil (704 mg, 85%);

(400 MHz, CDCl ₃ ) δ 2.32 (s, 6H, Me x 2 (enol)), 2.34 (s, 6H, Me x 2, (keto)) 3.76 (s ₅ 3H, OMe (keto)), 3.80 (s, 3H, OMe (enol)), 3.97 (s, 2H, CH ₂ ), 4.84 (s, 2H, OCH ₂ Ph (enol)), 4.86 (s, 2H, OCH ₂ Ph (keto)), 5.62 (s, IH, CH (enol)), 7.35-7.48 (m, 5H, Ar), 7.65 (s, 2H, Ar); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 16.5 (CH ₃ ), 16.6 (CH ₃ ), 45.5, 51.3, 52.4, 74.0, 86.2, 126.8, 127.8, 128.1, 128.2, 128.5, 128.53, 128.7, 129.6, 131.4, 131.7, 131.8, 136.8, 137.1, 158.5, 159.7, 160.6, 168.1, 171.5 (CO), 173.5 (CO), 191.6 (CO); IR v 2956, 1744, 1681, 1621, 1330, 1146, 896 cm ^"1 ; HRMS ESI ⁺ [M+H] ⁺ = 313.1440, requires 313.1440 for Ci ₉ H ₂ ]O ₄ .

5-(3,5-DimethyI-4-benzyIoxyphenyl)-3Jϊ-l,2-dithiole-3-th ione

P ₄ S ₁₀ (600 mg, 1.35 mmol), sulfur (80 mg, 2.48 mmol) and HMDO (2.9 niL, 14 mmol) was added to a solution 3-(4-benzyloxy-3,5-dimethylphenyl)-3-oxopropionic acid methyl ester (704 mg, 2.25 mmol) in xylene (3 mL). The reaction was heated under reflux for 1 h then cooled to rt. The reaction mixture was applied to silica gel and purified by flash chromatography (15-20% EtO Ac/petrol). The residue was recrystallised from dichloromethane/petrol to afford 5-(3,5-dimethyl-4- benzyloxyphenyl)-3H-l,2-dithiole-3-thione 18 as an orange solid, (388 mg, 50%); mp 138-140 °C; ¹ H NMR (500 MHz, CDCl ₃ ) δ, 2.34 (s, 6H, Me x 2), 4.87 (s, 2H, OCH ₂ Ph) , 7.35-7.47 (m, 8η, Ar, CH); ¹³ C NMR (125 MHz, CDCl ₃ ) δ 16.6 (CH ₃ ), 74.3 (CH ₂ ), 127.2 (C4), 127.6 (C5), 127.8, 128.3, 128.6, 132.9, 135.2, 136.7, 159.3, 173.0 (Ar-9), 215.2 (CS); HRMS ESI ⁺ [M+H] ⁺ = 345.0436, requires 345.0436 for C ₁₈ H ₇ OS ₃ ; Microanalysis: Found C, 62.80; H, 4.75; S, 27.80. Ci ₈ Hi ₆ OS ₃ requires C, 62.75; H, 4.68; S, 27.92%.

Reference

Still, C. W; Khan, M.; Mitra, A. Rapid chromatographic technique for preparative separations with moderate resolution. J. Org. Chem. 1978, 43, 2923-2925.

W

- 44 -

Assays For Testing COX-I and COX-2 Activity of Compounds 1 to 18 • Cyclooxygenase-1 (COX-I) Assay

Blood was collected from healthy humans and mixed with one-tenth volume of anticoagulant citrate solution (65 mM citric acid, 85 mM sodium citrate, 2 % glucose) and centrifuged at 200 x g for 10 minutes in a refrigerated centrifuge. The supernatant was mixed with 50% volume of Hanks' balanced solution buffered with 15 mM Herpes buffer (pH 7.4) and 30 % volume of the above citrate solution. This solution was then centrifuged at 750 x g for 10 minutes in a refrigerated centrifuge and the supernatant was discarded. The pellet was resuspended in Hanks' balanced buffer (as above) to give approximately 40 million platelets per millilitre. Platelets (25 millilitres) was added to an equal volume of 0.1 M Tris-HCl, pH 7.4 containing 10 mM EDTA, leupeptin (2 mg/mL), 2 mg/mL soybean trypsin inhibitor, 2 mg/mL aprontinin and 1 mM phenylmethylsulfonyl fluoride. The cell suspension was sonicated four times for 10 seconds and then centrifuged for 10,000 x g for 10 minutes at 40 C. The supernatant was then centrifuged at 100,000 x g for 60 minutes at 40 C and the resulting pellet was resuspended in 0.1 M Tris HCl, 10 mM EDTA (pH 7.4) and aliquots stored frozen at -80°C as a source of platelet microsomes for COX-I assays.

Microsomes were thawed and briefly sonicated and diluted to a protein concentration of 125 μg/mL in 0.1 M Tris-HCl, 10 mM EDTA buffer (pH 7.4) containing 0.5 mM phenol, 1 mM glutathione and 1 μM haematin. Drugs were dissolved in dimethyl sulfoxide and diluted in 3 -fold serial dilutions and then tested at 3 final concentrations of 0.3, 3 and 30 μM. Each tube contained 200 μL of microsomal suspension and 25 μL of drug dilution. After pre-incubation at room temperature for 15 minutes, 25 μL of arachidonic acid (peroxide free) (1 mM in Tris-HCl EDTA buffer) was added, mixed and incubated at room temperature for 40 minutes. Incubation was stopped by the addition of 1 M hydrochloric acid. Samples were then neutralized by the addition of 1 M sodium hydroxide (25 μL). The formation of prostaglandin E2 was then quantitated by specific radioimmunoassay. COX-I activity was calculated in the absence of drug as the difference between prostaglandin E2 levels in samples incubated in the presence of arachidonic acid versus the DMSO vehicle. The percentage of inhibition of prostaglandin E2 synthesis is calculated from the difference between prostaglandin E2 levels in samples incubated in the absence or presence of drug concentrations. The concentration of drug causing 50 % inhibition of enzyme activity (IC50) was calculated graphically from the three concentrations of added drug.

• Cyclooxygenase-2 (COX-2) Assay

Instead of human platelets as a source of COX-I enzyme, the gene for human COX-2 was cloned and expressed in Sf21 insect cells using a baculovirus expression system as described by Gierse, J.K. et al (1995) Biochemical Journal, Vol. 305, page 479-484. Cells expressing human COX-2 enzyme were homogenized in the same buffer as used for the COX-I assay and incubated with arachidonic acid (0.3 μM). COX-2 enzyme activity was determined by monitoring prostaglandin E2 production by specific radioimmunoassay. The percentage of inhibition of prostaglandin E2 synthesis is calculated from the difference between prostaglandin E2 levels in samples incubated in the absence or presence of drug concentrations. The concentration of drug causing 50 % inhibition of enzyme activity (IC50) was calculated graphically from the three concentrations of added drug.

Test Results

Compounds 1 and 2 were tested in the above assays and the results were as follows:

The test results show that compound 2 has selective COX-2 activity of at least 100, and compound 1 has significant COX-I activity and also COX-2 selectivity of approximately 10.

Compounds 1, 2, 4, 5, 6, 7, 10, 11, 12, 17 and 18 were further subjected to testing in the above in vitro assays at 1 μm concentrations and the results are as follows:

Comp ID # Chemical Structure MW % COX-I % COX-2 1 uM I uM

Compound 1 338.55 44 98

Compound 2 352,58 69

Compound 3 366.6

Compound 4 323 12 53

Compound 5 337 22 51

Compound 6 352.58 34 73

Compound 7 366.6 31 29

Compound 8 380.63

Compound 9 336.51

Compound 10 350.54 1 1 43

Compound 11 310.5 10 65

Compound 12 324.52 35

Compound 13 338.55

Compound 14 254.39

Compound 15 268.42

Compound 16 282.44

Compound 17 252.35 41

Compound 18 344.51 30

EXAMPLE 2 - Additional detail on synthesis

Routes for the synthesis of further exemplary compounds were devised as detailed in the following schematic flowcharts.

Compounds 1 and 2 from Example 1 were prepared via alternative routes, as detailed below. Details for the repeated synthesis of other compounds of the invention are also outlined below.

Compound 1 (5-(3,5-di-tert-butyl-4-hvdroxyphenyl)-3H-l ,2-dithiole-3-thione)

l-(3,5-Di-tert-butyl-4-hydroxyphenyl)ethanone (0.61 g, 2.45 mmol) in dry acetone (20 mL) was added potassium carbonate ( 0.5 g, 3.61 mmol) followed by chloromethyl methyl ether(0.30 mL, 3.94 mmol). The reaction mixture was stirred overnight with a calcium chloride drying tube. The solid was filtered and the filtrate was evaporated in vacuo. The residue was dissolved in dichloromethane and washed with IN sodium hydroxide followed by distilled water. The organic layer was dried over sodium sulphate and the solvents were removed in vacuo. The residue was chromatographed on silica gel (1 :6 ethyl acetate /hexane) to give l-(3,5-di-tert-butyl-4- (methoxymethoxy)phenyl)ethanone as a brown oil (0.64 g, 89% yield). This compound

was reacted with KH, CS ₂ , and then with HMDT, hexachloroethane following the same procedure for compound (1) to give 5-(3,5-di-tert-butyl-4-methoxymethylphenyl)-3H- l,2-dithiole-3-thione as an orange crystalline solid after column chromatography (0.30 g, 35 % yield).

5-(3,5-di-tert-butyl-4-methoxymethylphenyl)-3H-l,2-dithiole- 3-thione (0.30 g, 0.77 mmol) in dichloromethane (10 mL) was cooled in an ice bath and trifiuoroacetic acid (2 mL, 25.8 mmol) was added and stirred under nitrogen atmosphere for 30 min. The reaction mixture was evaporated in vacuo and the residue was chromatographed on silica gel (1:6 ethylacetate/hexane) to give compound 2 (5- (3,5-di-tert-butyl-4-hydroxyphenyl)-3H-l,2-dithiole-3-thione ) as a bronze colour solid (0.16 g, 61% yield). Mass Spec: m/z 339.0905. [M+H] ⁺ requires 339.0911.

¹ H NMR (300 MHz, CDCl ₃ ) 7.46 (2H, s), 7.38 (IH, s), 5.69 (IH, s), 1.45 (18H, s)

13 ^J ,C NMR(75 MHz, CDCl ₃ ) 214.93, 174.79, 157.80, 137.27, 134.40, 124.26, 123.25, 34.48, 30.03

Compound 2 (5-(3,5-di-tert-butyl-4-methoxyphenyl)-3H-L2-dithiole-3-thio ne)

Anhydrous aluminum chloride (1.29 g, 9.69 mmol) in dry dichloromethane (20 mL) under N ₂ atmosphere was cooled in an ice bath. This was added 2,6-di-tert-butylphenol (2.0 g, 9.69 mmol) followed by acetyl chloride (2.01 mL, 29.1 mmol) in dichloromethane (10 mL) using a dropping funnel. The reaction mixture was stirred for 1 h in ice and poured into ice (100 mL). The white emulsion was extracted with dichloromethane. The combined dichloromethane extractions were dried over sodium sulphate and concentrated. The brown residue was chromatographed on silica gel (4:1 hexane/ ethyl acetate) to give l-(3,5-di-tert-butyl-4-

hydroxyphenyl)ethanone as a yellow solid (2.06 g, 86% yield).

l-(3,5-Di-tert-butyl-4-hydroxyphenyl)ethanone (0.52 g, 2.09 mmol) in dry acetone (10 mL) was added anhydrous potassium carbonate (0.50 g, 3.61 mmol) followed by methyl iodide (0.40 mL, 6.05 mmol) and the reaction mixture was refluxed overnight with a calcium chloride drying tube. After cooling to room temperature the white precipitate was filtered off and the filtrate was concentrated. The residue was chromatographed on silica gel (1:10 ethyl acetate/hexane) to give l-(3,5-di-tert-butyl-4- methoxyphenyl)ethanone as a light brown liquid (0.4 g, 73% yield).

Ref: Thomas J. Curphey and Adam H. Libby, Tetrahedron Letters 41, 2000, 6977-6980.

To a well stirred suspension of potassium hydride (0.12 g, 3.07 mmol) in dry tetradydrofuran (5 mL) and dry DMPU (N,N'-dimethylpropyleneurea) (2.5 mL) under N ₂ atmosphere was added l-(3,5-di-tert-butyl-4-methoxyphenyl)ethanone (0.40 g, 1.5 mmol) in dry THF (1 mL). The enolate suspension was stirred for an additional 15 min after the gas evolution had ceased, then a solution of carbon disulfide (0.1 mL, 1.65 mmol) in dry THF (1.5 ml) and dry DMPU (0.75 mL) was added. The resulting red solution was stirred for 10 min and HMDT (hexamethyldisilathiane) (0.5 mL, 2.25 mmol) was added. After stirring for an additional 20 min, the reaction mixture was cooled to O ⁰ C, treated with a solution of hexachloroethane (0.36 g, 1.5 mmol) in THF (2 ml), and stirred for 30 min. Methanol (2 mL) was added cautiously to destroy any unreacted hydride and the reaction mixture was allowed to stand for 15 min. THF was removed in vacuo and excess distilled water was added. The bright orange emulsion was extracted with dichloromethane and dried over sodium sulphate. The dichloromethane was removed in rotary evaporator and DMPU was removed at 85 ⁰ C at 5 mm Hg to give a reddish black thick liquid which was chromatographed on silica gel (1 :10 ethyl acetate/hexane). 5-(3,5-di-tert-butyl-4-methoxyphenyl)-3H-l,2-dithiole-3- thione (0.17 g, 33 % yield) was obtained as an orange crystalline solid. MP: 100-101 ⁰ C

Microanalysis: C: 60.43, H: 6.96, S: 27.12, Ci ₈ H ₂₄ OS ₃ requires C: 61.32, H: 6.86, S: 27.28

¹ H NMR (300 MHz, CDCl ₃ ) 7.51 (2H, s), 7.38 (IH, s), 3.71 (3H, s), 1.43

(18H, s)

¹³ C NMR (75 MHz, CDCl ₃ ) 215.27, 174.12, 163.49, 145.63, 135.18, 126.25, 125.47, 64.55, 36.02, 31.76

5-(3,5-di-tert-butyl-4-methoxyphenyl)-3H-l,2-dithiole-3-t hione

P ₄ S ₁₀ (356 mg, 0.801 mmol), sulfur (400 mg, 12.5 mmol) and HMDO (1.4 mL, 6.47 mmol) was added to a solution of the methyl ester Ia (363 mg, 1.13 mmol) in xylene (2 mL). The reaction was heated under reflux for 3 h and then cooled to rt. Acetone was then added (3 mL) followed by 5.3 M K ₂ CO ₃ (1.3 mL) and the mixture stirred for 1 h. Toluene was then added and the organic layer was washed with 2.15 M K ₂ CO ₃ , water, brine, dried (Na ₂ SO ₄ ) and concentrated. The residue was passed through a plug of silica eluting with 10 % EtO Ac/petrol and the product was recrystallised from EtO Ac/petrol to afford the dithiole-3-thione 2a as an orange solid (117 mg, 28 %); mp 106-107 ⁰ C (lit. ¹ mp 100-101 ⁰ C) ₃ ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.45 (s, 18H, f-Bu x 2), 3.74 (s, 3H, OCH ₃ ),7.41 (s, IH, C=CH) 7.53 (s, 2H, Ph); ¹³ C NMR (100 MHz, CDCl ₃ ) 31.8, 36.2, 64.6, 125.5, 126.2, 135.2, 145.6, 163.5, 174.2, 215.2 (C=S); IR 2948, 2865, 1744, 1587, 1498, 1304, 1110, 782 cm ^"1 ; HRMS [M+H] ⁺ = found 353.1063, requires 353.1062 Ci ₈ H ₂₆ OS ₃ , Microanalysis Found C, 61.33; H, 6.85, S, 27.34 Ci ₈ H ₂₄ OS ₃ requires C, 61.32; H, 6.86, S, 27.28 %.

5-(3,5-Di-tert-butyl-4-hydroxyphenyl)-3H-l,2-dithiole-3-t hione

P ₄ Si ₀ (1.15 g, 2.59 mmol), sulfur (130 mg, 4.05 mmol) and HMDO (2.4 mL, 11 mmol) was added to the methyl ester 3a (1.3Og, 3.71 mmol) in xylene (15 mL). The reaction was heated under reflux for 4 h. The reaction was then cooled to rt and concentrated. The reside was chromatographed on silica gel eluting with 5 % EtO Ac/petrol and the product was recrystallised from EtO Ac/petrol to afford the dithiole-3-thione 4a as an yellow/brown solid (540 mg, 43 %); mp 180-183 ⁰ C; ¹ H NMR (500 MHz, CDCl ₃ ) δl.47 (s, 18H, t-Bu x 2), 5.72 (s, IH, OH), 7.41 (s, IH, C=CH), 7.48 (s, 2H, Ph); ¹³ C NMR (125 MHz, CDCl ₃ ) δ 30.3, 34.8, 123.5, 124.5, 134.7, 137.5, 158.1, 175.1, 215.2 (C=S); IR 3429, 2960, 1593, 1514, 1419, 889, 715 cm- ¹ ; HRMS [M+H] ⁺ = found 339.0908, requires 339.0911 C ₁₇ H ₂₄ OS ₃ .

5-(3,5-Di-tert-butyl-4-hydroxyphenyl)-4-(methyl)-3H-l,2-d ithiole-3-thione

P ₄ S] ₀ (950 mg, 2.14 mmol), sulfur (100 mg, 3.12 mmol) and HMDO (3.5 mL, 16.5 mmol) was added to a solution of the methyl ester 5a (1.07 g, 2.93 mmol) in xylene (7 mL). The reaction was heated under reflux for 1 h. then cooled to rt and concentrated. The reside was chromatographed on silica gel eluting with 5 % EtO Ac/petrol and the solid recrystallised from EtO Ac/petrol to afford the dithiole-3- thione 6a as an orange solid (600 mg, 81 %); mp 174-175 ⁰ Cj ¹ H NMR (400 MHz,

CDCl ₃ ) δl.47 (s, 18H, t-Bu x 2), 2.24 (s, 3H, CH ₃ ), 5.61 (s, IH, OH), 7.30 (s, 2H, Ph); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 16.9, 30.1, 34.5, 104.7, 124.7, 125.7, 136.8, 140.8, 156.2, 170.2, 215.4 (C=S); IR 3624, 2959, 2911, 1594, 1430, 1120, 885 cm ^"1 ; Microanalysis Found C, 61.37, H, 6.91, S, 27.15 C ₁₈ H ₂₄ OS ₃ requires C, 61.32; H, 6.86, S, 27.28 %.

5-(3,5-Di-tert-butyl-4-hydroxyphenyl)-4-(methyl)-3H-l _! 2-dithiole-3-one

A solution of the dithiole-3-thione 6a (441 mg, 1.25 mmol) in hot acetic acid (5 mL) was added to a solution OfHg(OAc) ₂ (797 mg, 2.50 mmol) in hot acetic acid (5 mL) and the reaction heated under reflux for 1 h. The reaction was cooled to rt and filtered. The filtrate was concentrated and the residue chromatographed on silica gel then recrystallised from EtO Ac/petrol to afford the dithiole-3-one 7a as a colourless soild (257 mg, 61%); mp 189-190 °C; ¹ H NMR (400 MHz, CDCl ₃ ) δl.47 (s, 18H, t-Bu x 2), 2.06 (s, 3H, CH ₃ ), 5.56 (s, IH, OH), 7.26 (s, 2H, Ph); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 14.1, 30.1, 31.8, 34.5, 125.2, 125.7, 136.6, 155.9, 164.3, 195.4 (C=O); IR 3532, 2951, 1623, 1556, 1432, 1113, 950, 658 cm ^"1 ; HRMS [M+H] ⁺ = found 337.1292, requires 337.1290 C ₁₈ H ₂₅ O ₂ S ₂ ; Microanalysis Found C, 64.31; H, 7.22, C] ₈ H ₂₄ O ₂ S ₂ requires C, 64.25; H, 7.19%.

5-(3,5-Di-tert-butyl-4-methoxyphenyl)-4-(methyl)-3H-l,2-d ithiole-3-thione

P ₄ S ₁₀ (1.51 mg, 3.40 mmol), sulfur (202 mg, 6.30 mmol) and HMDO (7.3 rriL, 34.3 mmol) was added to a solution of the methyl ester 8a (1.92 g, 5.74 mmol) in xylene (5 mL). The reaction was heated under reflux for 1.5 h. then cooled to rt. The reaction mixture was loaded on a silica column and chromatographed eluting with 5 % EtO Ac/petrol and the resulting solid recrystallised from petrol to afford the dithiole-3-thione 9a as an orange solid (2.07 g, 98 %); mp 83-84 ⁰ C; ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.46 (s, 18H, t-Bu x 2), 2.23 (s, 3H ₅ C(Me)), 3.76 (s, 3H, OMe), 7.35 (s, Ph, 2H); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 16.9, 31.9, 36.0, 64.3, 127.1, 127.9, 141.2, 145.0, 161.8, 169.4, 215.5 (C=S); IR 2961, 2869, 1525, 1307, 1223, 1007, 732 cm ^"1 ; Microanalysis Found C, 62.34; H, 7.25, Ci ₉ H ₂₃ OS ₃ requires C, 62.25; H, 7.15%.

5-(3,5-Di-tert-butyl-4-ethoxyphenyl)-4-(methyl)-3H-l,2-di thiole-3-thione

P ₄ Si ₀ (484 mg, 1.09mmol), sulfur (64 mg, 2.00 mmol) and HMDO (2.3 mL, 10.8 mmol) was added to a solution of the methyl ester 10a (632 mg, 1.81 mmol) in xylene (3 mL). The reaction was heated under reflux for 2 h. then cooled to rt. The reaction mixture was then loaded on a silica column and chromatographed eluting with 5 % EtO Ac/petrol and the resulting solid recrystallised from petrol to afford the dithiole-3-thione as an orange solid (480 mg, 70 %); mp 83-84 °C; ¹ H NMR (400 MHz, CDCl ₃ ) 1.44 (bs, 21H, r-Bu x 2, CH ₃ CH ₂ O), 2.23 (s, 3H, CH ₃ ), 3.83 (q, 2H, CH ₃ CH ₂ O), 7.34 (s, 2η, Ph); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 14.9, 16.8, 31.9, 36.1, 72.1, 126.8, 127.6, 127.7, 141.2, 145.0, 160.3, 169.5, 215.6 (C-S); IR 2956, 1522, 1425, 1383, 1217, 1084, 886 cm ^"1 ; HRMS [M+H] ⁺ = found 381.1375, requires 381.1375 C ₂₀ H ₂₉ OS ₃ ; Microanalysis Found C, 63.15; H, 7.50, S, 25.11 C ₂₀ H ₂₈ OS ₃ requires C, 63.11; H, 7.41, S, 25.27%.

5-(3,5-Di-tert-butyl-4-methoxyphenyl)-4-(methyl)-3H-l,2-d ithiole-3-one

A hot solution Of Hg(OAc) ₂ (829 mg, 2.60 mmol) in hot acetic acid (35 niL) was added to a hot solution of the dithiole-3-thione 9a (500 mg, 1.36 mmol) in hot acetic acid (35 mL) and the reaction was heated under reflux for 1 h. The reaction was cooled to rt and filtered. The filtrate was concentrated and the residue chromato graphed on silica gel (10 % EtO Ac/petrol) to afford a brown oil which was crystallised and was then recrystallised from ethanol/water to afford the dithiole-3-one 12a as a colourless soild (370 mg, 79 %); mp 112-114 ⁰ C; ¹ H NMR (400 MHz, CDCl ₃ ) δ 1.45 (s, 18H, t-Bu x 2), 2.06 (s, 3H ₅ CH ₃ ), 3.75 (s, 3H, OMe), 7.32 (s, 2H, Ph); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 14.1, 31.9, 36.0, 64.4, 126.2, 126.6, 128.3, 144.8, 161.5, 163.8, 195.3 (C=O); IR 2961, 2870, 1590, 1032 cm ^"1 ; HRMS [M+H] ⁺ = found 351.1447, requires 351.1447 Ci ₉ H ₂₇ O ₂ S ₂ .

5-(4-Methoxy-3, 5-dimethylphenyl)-[l, 2] -3H-dithiole-3-thione

P ₄ Si ₀ (1.07 g, 2.41 mmol), sulfur (141 mg, 4.40 mmol) and HMDO (5.2 mL, 25 mmol) was added to the methyl ester (935 mg, 4.40 mmol) in xylene (3 mL). The reaction was heated under reflux for 1.5 h. then cooled to rt. The reaction mixture was then chromatographed on silica gel eluting with 5 to 15 % EtO Ac/petrol. The residue was recrystallised from EtOAc/ petrol to afford the dithiole-3-thione as an orange solid mp 96-99 °C; (576 mg, 53 %); ¹ H NMR (500 MHz, CDCl ₃ ) δ 2.33 (s, 6H,

Me x 2), 3.77 (s, 3H, OMe), 7.32 (s, 2H, Ph), 7.37 (s, IH, CH); ¹³ C NMR (1250 MHz, CDCl ₃ ) δ 16.2, 59.8, 127.1, 127.5, 132.5, 135.3, 160.6, 173.0, 215.3 (C=S); IR 2937, 1599, 1470, 1181, 1064, 832 cm ^"1 ; HRMS [M+H] ⁺ = 269.0125 found, requires 269.1023 C] ₂ H ₁₃ OS ₃ .

5-(4-Methoxy-3, 5-dimethylphenyl)-[l , 2] -3H-dithiole-3-one

A hot solution OfHg(OAc) ₂ (570 mg, 1.79 mmol) in hot acetic acid (15 mL) was added to a hot solution of the dithiole-3-thione 14a (240 mg, 0.894 mmol) in hot acetic acid (15 mL) and the reaction was heated under reflux for 1.5 h. The reaction was cooled to rt and filtered. The filtrate was concentrated and the residue chromato graphed on silica gel (10 % EtO Ac/petrol) to afford a pale yellow solid which was recrystallised from EtO Ac/Petrol to afford the dithiole-3-one as a colourless soild (108 mg, 48 %); mp 95-96 ⁰ C; ¹ H NMR (500 MHz, CDCl ₃ ) δ 2.34 (s, 6H, Me x 2), 3.77 (s, 3H, OMe), 6.76 (s, IH ₃ CH), 7.29 (s, 2H, Ph); ¹³ C NMR (125 MHz, CDCl ₃ ) δ 16.2, 59.8, 117.0, 127.1, 128.1, 132.3, 160.2, 170.2, 194.1 (C=O); IR 2924, 1659, 1546, 1237, 1001, 859 cm ^"1 ; HRMS [M+H] ⁺ = found 253.0351, requires 253.0351 Ci ₂ Hi ₃ O ₂ S ₂ .

EXAMPLE 3 - FURTHER TEST RESULTS

Comparative Testing l,2-dithiole-3-thione, the R ⁵ ring system in compounds 1 and 2 above was also tested for COX-I and COX-2 activity using the assays outlined above. It was found that l,2-dithiole-3-thione did not significantly inhibit either COX-I or COX-2 at lOμm concentrations - the percentage inhibition was 9% and 35%, respectively. It is however reported in the literature to have neuroprotective properties.

In Vivo Assays

1. Anti-inflammatory activity

The method used was the carrageenan-induced rat paw oedema assay which is based on the method described in CA. Winter et al (1962) Proceedings of the Society for Experimental Biology & Medicine, Vol. Ill: page 544-547.

Groups of six male mice weighing approximately 25 gm were acclimatised in the laboratory for seven days prior to use. Three groups were treated daily with either compound (1) or compound (2) or compound (11) at a dose of 50 mg/kg by gastric lavage; one group was treated with the vehicle (2% tween 80) and another group was treated with a maxium dose of aspirin (150 mg/kg). On the third day of treatment, an injection of carrageenan (50 μl of a 1% suspension) was given into the right hind paw by intra-plantar injection one hour after the third dose of drugs. Four hours later, the volume of the right hind paw (inflamed) and the volume of the left hind paw (control) was measured using a plethysmometer (Ugo Basile, Italy).

The results are ; set out in the following table:

Group Dosage % Inhibition - Right Paw

Compound (1) 50 mg/kg x 3 35%

Compound (2) 50 mg/kg x 3 25%

Compound (11) 50 mg/kg x 3 25%

Aspirin 150 mg/kg 100%

Vehicle 0.25 ml x 3 0%

2. Peripheral Analgesia Analgesia will be evaluated in mice using the abdominal constriction test which is based on the method described in a paper by H.O.J. Collier et al (British Journal of Pharmacology, Vol. 32: page 295-310, 1968). Male adult mice will be fasted overnight and then dosed with a suspension of the test drug in 0.5 mL by gastric lavage or with the vehicle for a control group. Thirty minutes later, mice will be injected with 0.1 mL of acetic acid solution (0.7%) and placed in a clear plastic box for observation. The number of full abdominal constrictions (comprising a stretching of the hind limbs to full extent allowing the abdomen to touch the floor) will be counted over the next 15 minutes. Five mice per group will be tested at four dose levels together with a control group. The total number of constrictions will be summed for the five mice in each group. Analgesic activity will be recorded as the percentage inhibition of abdominal constrictions when the drug is present compared to the control group. The effective

dose to inhibit the constrictions by 50% (ED50) will be calculated from the dose- response plot.

3. Central analgesia The injection of formalin into the hindpaw of mice causes two periods of intense licking with the first period due to a direct peripheral action and the second period due to a central action. Analgesic drugs are capable of blocking one or both periods depending on their mechanism of action. Thus analgesia will be evaluated in mice based on the method described in a paper by S. Hunskaar and K. Hole (Pain, Vol. 30: page 103-114, 1987). Male adult mice will be placed individually in a transparent plastic chamber and injected intraperitoneally with the test drug (0.3 mL) or vehicle and left for 30 minutes. Formalin (1 %, 20 uL) will then be injected into the dorsal hind paw using a microsyringe and a 26-gauge needle. The mouse is then placed back in the chamber and the video recorded over the next 30 minutes so that the number and amount of time spent licking the injected paw could be determined accurately. The effect of the test drug on licking parameters can then be expressed as the percentage inhibition.

4. Neuroprotection in a mouse model of Parkinson 's disease Inflammation in the brain is now believed to play a major role in the loss of neurons that characterize Parkinson's and Alzheimer's diseases. Injection of rodents with 1 -methyl -4-phenyl-l, 2,3, 6-tetrahydropyridine (MPTP) causes central inflammation and leads to biochemical and behavioural symptoms characteristic of Parkinson's disease while a known cyclooxygenase-2 inhibitor, meloxicam blocks this action of MPTP. This is the basis of a method described in a paper by P. Teismann and B. Ferger (Synapse, Vol. 39: page 167-174, 2001) for assessing neuroprotective drugs in Parkinson's disease. Adult male rats (C57BL/6 strain) in groups of five will be injected intraperitoneally with either test drug (10 or 30 mg/kg) or meloxicam (10 or 30 mg/kg) immediately before a single injection of MPTP (30 mg/kg, subcutaneously) or saline (controls). Seven days after MPTP or saline, with or without drug coadministration, mice will be killed and the midbrain taken for histological examination by staining the dopamine-containing neurons with tyrosine hydroxylase antiserum as described by Teismann and Ferger (2001). The number of tyrosine hydroxylase staining neurons will be counted by quantitative microscopy and the ability of the test drugs to block loss of tyrosine hydroxylase staining cell bodies after MPTP will be calculated.

It will be apparent to the person skilled in the art that while the invention has been described in some detail for the purposes of clarity and understanding, various modifications and alterations to the embodiments and methods described herein may be made without departing from the scope of the inventive concept disclosed in this specification.