WO2008046155A1 - Methods for regulating glucose homeostasis and agents therefor - Google Patents

Abstract

The invention particularly relates to methods of regulating glucose homeostasis and to the therapeutic or prophylactic treatment of diseases and conditions, such as diabetes, syndrome X, obesity, hyperglycaemia, cardiovascular disease, vascular disease and kidney disease, in which impaired glucose uptake due to insulin resistance is involved or implicated. The present invention further relates to compounds and agents and compositions thereof for use in the treatment methods.

Description

METHODS FOR REGULATING GLUCOSE HOMEOSTASIS AND AGENTS THEREFOR

FIELD OF THE INVENTION

The present invention relates generally to the field of therapy. The invention particularly relates to methods of regulating glucose homeostasis and to the therapeutic or prophylactic treatment of diseases and conditions, such as diabetes, syndrome X, obesity, hyperglycaemia, cardiovascular disease, vascular disease and kidney disease, in which impaired glucose uptake due to insulin resistance is involved or implicated. The present invention further relates to compounds and agents and compositions thereof for use in the treatment methods.

DESCRIPTION OF THE PRIOR ART •

The reference in this specification to any prior publication (or information derived from it), or to any matter which is known, is not, and should not be taken as an acknowledgment or admission or any form of suggestion that that prior publication (or information derived from it) or known matter forms part of the common general knowledge in the field of endeavour to which this specification relates.

Bibliographic details of references provided in the subject specification are listed at the end of the specification.

Glucose is the body's preferred energy source. Once entered into the blood stream, it requires the assistance of the insulin to enter hepatic, muscle and adipose cells in order to be stored or utilised. In a healthy individual, glucose homeostasis is controlled primarily by insulin. As blood glucose levels rise, such as after eating, specialised β-cells within the pancreas release insulin which promotes glucose uptake, intracellular metabolism and glycogen synthesis by the body's target tissues. Thus, in healthy individuals, blood glucose concentrations are strictly controlled, typically in the range of 80-110 mg/dl. However, where the pancreas produces an inadequate insulin response, or the target cells do not respond appropriately to the insulin produced, the glucose cannot enter the body's cells. This results in a rapid accumulation of glucose in the blood stream (hyperglycemia).

High blood glucose levels over time may cause cardiovascular disease, retinal damage, renal failure, nerve damage, erectile disfunction and gangrene (with the risk of amputation). Furthermore, in the absence of available glucose, cells turn to fats as an alternative energy source. Resulting ketones, a product of fat hydrolysis, can accumulate in the blood stream instigating hypotension and shock, coma and even death.

Chronically elevated blood glucose levels (greater than about 126 mg/dl) from either inadequate insulin secretion and/or an inadequate response or sensitivity to insulin is referred to as diabetes, a disease which is now suffered by more than 10 percent of adults in the USA. The disease may be characterised by persistent hyperglycemia, polyuria, polydipsia and/or hyperphagia, chronic microvascular complications such as retinopathy, nephropathy and neuropathy, and macrovascular complications, such as hyperlipidemia and hypertension which can lead to blindness, end-stage renal disease, limb amputation and myocardial infarction. The three most common types of diabetes are type 1, type 2 and gestational.

Type 1, known as insulin dependent diabetes mellitus (IDDM), or juvenile-onset diabetes, occurs in 10-15% of all cases. It is most commonly diagnosed in children and adolescents but can occur in young adults as well. It is characterised by β-cell destruction resulting in a loss of insulin secretory function. Most cases relate to autoimmune destruction of the β- cells. Treatment is via insulin injection and must be continued indefinitely.

In contrast, in type 2 diabetes, known as non-insulin dependent diabetes mellitus (NIDDM) or late-onset diabetes, insulin levels are initially normal but the body's target cells lose their responsiveness to insulin. This is known as insulin resistance or insensitivity. By way of compensation, the pancreas begins to secrete excess insulin, however, in time the pancreas becomes less able to produce enough insulin resulting in chronic hyperglycaemia. Initial symptoms are typically milder than for type 1 and the condition may go undiagnosed for years before more severe symptoms present. Lifestyle (poor diet and inactivity) is considered to be a major mediating factor of the condition, although a genetic predisposition increases the risk. Treatment focuses on lifestyle modification, insulin therapy and/or anti-diabetic agents such as insulin sensitisers and insulin releasers.

Gestational diabetes occurs in about 2-5% of all pregnancies. It is temporary, but if untreated may cause foetal complications. Most sufferers make a complete recovery after the birth. However, a proportion of women who develop gestational diabetes go on to develop type 2 diabetes.

Other, less common, causes of diabetes include genetic defects in β-cells, genetically related insulin resistance, diseases of the pancreas, hormonal defects, malnutrition and chemical or drug influences.

Impaired glucose tolerance and impaired fasting glucose, are pre-type 2 diabetic states, closely related to type 2 and occur when the blood glucose level is higher than normal but not high enough to be classified as diabetes (about 110-126 mg/dl). As with type 2, the body produces insulin but in an insufficient amount or the target tissues are unresponsive to the insulin produced.

Syndrome X, also known as Insulin Resistance Syndrome (IRS) is a cluster of risk factors for heart disease and is associated with insulin resistance. It presents symptoms or risk factors for the development of type 2 diabetes and heart disease including obesity, atheriosclerosis, hypertriglyceridemia, low HDL cholesterol, hyperinsulinemia, hyperglycaemia, hypertension, impaired glucose tolerance and impaired fasting glucose.

High blood glucose levels and insulin resistance are also associated with fatty liver disease, which can progress to chronic inflammation, fibrosis and cirrhosis.

It is therefore apparent that insulin resistance, or insensitivity, can play a significant role in diabetic and other hyperglycemic-related conditions.

The latest WHO estimate (for the number of people with diabetes, worldwide, in 2003) is 194 million. This is expected to increase to at least 330 million by 2025 and it is estimated that there are around 4 million deaths per year are related to the disorder.

Given the prevalence of diabetes and other diseases and conditions with which insulin resistance is associated, and their potential consequences, there remains a continuing need for new therapeutic or preventative treatments for insulin resistance.

SUMMARY OF THE INVENTION

Insulin sensitisers resensitise cells to the action of insulin thereby reducing blood glucose levels and triglyceride levels. The present invention is predicated on compounds related to strictosidine, a key intermediate in the biosynthesis of terpenoid indole alkaloids, as a new class of insulin sensitizers and provides new methods for modulating glucose uptake by a cell and treating insulin resistance and conditions associated with insulin resistance.

The compounds contemplated by the invention include those of Formula (I):

wherein,

is a single or a double bond;

R₁-R₄ are independently selected from hydrogen, hydroxy, thiol, halo, CO₂H, carboxy ester, amino, nitro, cyano, amido, sulfoxide, sulfonamide, sulfonyl, sulfate, sulfonate, phosphate, phosphonate, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted acyl, optionally substituted alkoxy, optionally substituted alkenyloxy, optionally substituted alkynyloxy, optionally substituted aryloxy, optionally substituted heteroaryloxy, optionally substituted carbocyclyloxy, optionally substituted heterocyclyloxy, optionally substituted aralkyloxy, optionally substituted acyloxy, optionally substituted alkylthio, optionally substituted alkenylthio, optionally substituted alkynylthio, optionally substituted arylthio, optionally substituted carbocyclylthio, optionally substituted aralkylthio, optionally substituted heteroarylthio, optionally substituted heterocyclylthio, optionally substituted acylthio or any 2 adjacent Ri-R₄ together form a -0-(CHi)_n-O- or -NR-(CH₂)_n-NR-group wherein n is 1 or 2 and each R is independently H or C_1-6alkyl;

R₆ is absent when "^"- is a double bond;

R₅ and R₆ (when present) independently are selected from hydrogen, optionally substituted alkyl, optionally substituted acyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aralkyl, optionally substituted heterocyclyl, optionally substituted carbocyclyl, and amido;

R₇ is selected from hydrogen, optionally substituted alkyl, CO₂H, carboxy ester, amido, optionally substituted heteroaryl and optionally substituted aryl;

R₈ is selected from hydrogen, optionally substituted alkyl, optionally substituted aryl, optionally substituted heteroaryl, CO₂H, carboxy ester and SO₃H;

R_7a and R_8a are independently selected from hydrogen, optionally substituted alkyl and optionally substituted aryl.

COX forms a group selected from optionally substituted acyl, CO₂H, carboxy ester and amido, Y is selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclyl, optionally substituted carbocyclyl, optionally substituted acyl and a saccharide group,

Z is selected from optionally substituted at least C₂alkyl, optionally substituted alkenyl and optionally substituted alkynyl,

and pharmaceutically acceptable salts and prodrugs thereof.

In a first aspect, the present invention thus provides a method of increasing glucose uptake by a cell comprising contacting said cell with a compound of Formula (I) or a pharmaceutically acceptable salt or prodrug thereof.

Methods for increasing glucose uptake by a cell, for example adipocytes and muscle and hepatic cells, can be performed in vivo or in vitro.

A second aspect of the invention relates to a method for regulating glucose homeostasis in a subject in need thereof comprising the administration of a compound of Formula (I) or a pharmaceutically acceptable salt or prodrug thereof.

In another aspect, the present invention provides a method of treating insulin resistance in a subject in need thereof comprising the administration of a compound of Formula (I) or a pharmaceutically acceptable salt or prodrug thereof.

A further aspect of the invention relates to a method of lowering blood glucose levels in a subject in need thereof comprising the administration of a compound of Formula (I) or a pharmaceutically acceptable salt or prodrug thereof.

Yet a further aspect of the invention provides a method for treating a disease or condition, or symptom thereof, in which insulin resistance is involved comprising the administration of a compound of Formula (I) or a pharmaceutically acceptable salt or prodrug thereof to subject in need thereof.

In yet another aspect, the invention provides a compound of Formula (I) or a pharmaceutically acceptable salt or prodrug thereof provided that the compound is not (3R) or (3S) strictosidine, strictosidine ethyl ester and allyl ester, strictosidinic acid, 5- carboxystrictosidine, 3,4-dehydro-5-carboxystrictosidine, 3,4-dehydro5- carbomethoxystrictosidine or gluco-acetylated forms thereof.

Further aspects of the invention provide a compound of Formula (I), or pharmaceutically acceptable salt or prodrug thereof for use in therapy, particularly for regulating glucose homeostasis, treating insulin resistance, lowering blood levels and/or treating a disease or condition, or one or more symptoms thereof, in which insulin resistance is involved.

The present invention also provides for agents and compositions comprising compounds of Formula (I), or pharmaceutically acceptable salt or prodrug thereof and the use of said compounds in treatment in the manufacture of medicaments for regulating glucose homeostasis, treating insulin resistance, lowering blood levels and/or treating a disease or condition, or one or more symptoms thereof, in which insulin resistance is involved. Advantageously, in certain embodiments, said compositions and medicaments comprise the compound of Formula (I), or pharmaceutically acceptable salt or prodrug thereof, as at least 50%, such as at least 90 or 95% of the active component.

DETAILED DESCRIPTION OF THE INVENTION

Throughout this specification and the claims which follow, unless the context requires otherwise, the word "comprise" and variations such as "comprises" and "comprising" will be understood to imply the inclusion of a stated integer or step or group of integers but not the exclusion of any other integer or step or group of integers.

The singular forms "a", "an" and "the" include plural aspects unless the context clearly dictates otherwise.

As used herein, the term "alkyl" or "alk", used either alone or in compound words denotes straight chain, branched or cyclic alkyl, preferably C_1-20 alkyl, eg C_1-10 or C_1-6. Examples of straight chain and branched alkyl include methyl, ethyl, «-propyl, isopropyl, «-butyl, sec- butyl, t-butyl, n-pentyl, 1,2-dimethylpropyl, 1,1-dimethyl-propyl, hexyl, 4-methylpentyl, 1- methylpentyl, 2-methylpentyl, 3-methylpentyl, 1,1-dimethylbutyl, 2,2-dimethylbutyl, 3,3- dimethylbutyl, 1,2-dimethylbutyl, 1,3-dimethylbutyl, 1,2,2,-trimethylρropyl, 1,1,2- trimethylpropyl, heptyl, 5-methylhexyl, 1-methylhexyl, 2,2-dimethylpentyl, 3,3- dimethylpentyl, 4,4-dimethylpentyl, 1,2-dimethylpentyl, 1,3-dimethylpentyl, 1,4- dimethyl-pentyl, 1,2,3-trimethylbutyl, 1,1,2-trimethylbutyl, 1,1,3-trimethylbutyl, octyl, 6- methylheptyl, 1-methylheptyl, 1,1,3,3-tetramethylbutyl, nonyl, 1-, 2-, 3-, 4-, 5-, 6- or 7- methyl-octyl, 1-, 2-, 3-, 4- or 5-ethylheptyl, 1-, 2- or 3-propylhexyl, decyl, 1-, 2-, 3-, 4-, 5-, 6-, 7- and 8-methylnonyl, 1-, 2-, 3-, 4-, 5- or 6-ethyloctyl, 1-, 2-, 3- or 4-propylheptyl, undecyl, 1-, 2-, 3-, A-, 5-, 6-, 7-, 8- or 9-methyldecyl, 1-, 2-, 3-, A-, 5-, 6- or 7-ethylnonyl, 1-, 2-, 3-, 4- or 5-propylocytl, 1-, 2- or 3-butylheptyl, 1-pentylhexyl, dodecyl, 1-, 2-, 3-, A-, 5-, 6-, 7-, 8-, 9- or 10-methylundecyl, 1-, 2-, 3-, A-, 5-, 6-, 7- or 8-ethyldecyl, 1-, 2-, 3-, A-, 5- or 6-propylnonyl, 1-, 2-, 3- or 4-butyloctyl, 1-2-pentylheptyl and the like. Examples of cyclic alkyl include mono- or polycyclic alkyl groups such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, cyclodecyl and the like. Where an alkyl group is referred to generally as "propyl", butyl" etc, it will be understood that this can refer to any of straight, branched and cyclic isomers where appropriate. An alkyl group may be optionally substituted by one or more optional substitutents as herein defined.

The term "alkenyl" as used herein denotes groups formed from straight chain, branched or cyclic hydrocarbon residues containing at least one carbon to carbon double bond including ethylenically mono-, di- or poly-unsaturated alkyl or cycloalkyl groups as previously defined, preferably C_2-20 alkenyl (eg C_2-1O or C_2-6)- Examples of alkenyl include vinyl, allyl, 1-methylvinyl, butenyl, iso-butenyl, 3-methyl~2-butenyl, 1-pentenyl, cyclopentenyl, 1-methyl-cyclopentenyl, 1-hexenyl, 3-hexenyl, cyclohexenyl, 1-heptenyl, 3-heρtenyl, 1-octenyl, cyclooctenyl, 1-nonenyl, 2-nonenyl, 3-nonenyl, 1-decenyl, 3- decenyl, 1,3-butadienyl, l-4,pentadienyl, 1,3-cyclopentadienyl, 1,3-hexadienyl, 1,4- hexadienyl, 1,3-cyclohexadienyl, 1,4-cyclohexadienyl, 1,3-cycloheρtadienyl, 1,3,5- cycloheptatrienyl and 1,3,5,7-cyclooctatetraenyl. An alkenyl group may be optionally substituted by one or more optional substitutents as herein defined.

As used herein the term "alkynyl" denotes groups formed from straight chain, branched or cyclic hydrocarbon residues containing at least one carbon-carbon triple bond including ethynically mono-, di- or poly- unsaturated alkyl or cycloalkyl groups as previously defined. Unless the number of carbon atoms is specified the term preferably refers to C_2-2O alkynyl (eg C_2-10 or C₂.₆). Examples include ethynyl, 1-propynyl, 2-propynyl, and butynyl isomers, and pentynyl isomers. An alkynyl group may be optionally substituted by one or more optional substitutents as herein defined.

Terms written as " [group] oxy" refer to a particular group when linked by oxygen, for example, the terms "alkoxy", "alkenoxy", . "alkynoxy" and "aryloxy" and "acyloxy" respectively denote alkyl, alkenyl, alkynyl, aryl and acyl groups as hereinbefore defined when linked by an oxygen atom. Terms written as "[group]thio" refer to a particular group when linked by sulfur, for example, the terms "alkylthio", "alkenylthio", alkynylthio" and

"arylthio" respectively denote alkyl, alkenyl, alkynyl, aryl groups as hereinbefore defined when linked by a sulfur atom. Similarly, a term written as ^M[groupA]groupB" is intended to refer to a groupA when linked by a divalent form of groupB, for example, "hydroxyalkyl" is a hydroxy group when linked by an alkylene group.

The term "halogen" ("halo") denotes fluorine, chlorine, bromine or iodine (fluoro, chloro, bromo or iodo).

The term "aryl" (or "carboaryl)" or the abbreviated form "ar" used in compound words such as "aralkyl" denotes any of single, polynuclear, conjugated and fused residues of aromatic hydrocarbon ring systems. Examples of aryl include phenyl, biphenyl, terphenyl, quaterphenyl, naphthyl, tetrahydronaphthyl, anthracenyl, dihydroanthracenyl, benzanthracenyl, dibenzanthracenyl, phenanthrenyl, fluorenyl, pyrenyl, idenyl, azulenyl, chrysenyl. Preferred aryl include phenyl and naphthyl. An aryl group may be optionally substituted by one or more optional substituents as herein defined.

The term "carbocyclyl" includes any of non-aromatic monocyclic, polycyclic, fused or conjugated cyclic hydrocarbon residues, preferably C_3-20 (e.g. C₃-_J0 or C_3-8). The rings may be saturated, for example cycloalkyl, or may possess one or more double bonds (cycloalkenyl) and/or one or more triple bonds (cycloalkynyl). Particularly preferred carbocyclyl are 5-6-membered or bicyclic 9-10 membered ring systems. Suitable examples include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, cyclodecyl, cyclopentenyl, cyclohexenyl, cyclooctenyl, cyclopentadienyl, cyclohexadienyl, cyclooctatetraenyl, indanyl, decalinyl and indenyl. A carbocyclyl group may be optionally substituted by one or more optional substituents as herein defined.

The term "heterocyclyl" when used alone or in compound words includes any of monocyclic, polycyclic, fused or conjugated hydrocarbon residues, preferably C_3-20 (e.g. C_3-10 or C_3-8) wherein one or more carbon atoms are replaced by a heteroatom so as to provide a non-aromatic residue. Suitable heteroatoms include, O, N, S, P and Se₅ particularly O, N and S. Where two or more carbon atoms are replaced, this may be by two or more of the same heteroatom or by different heteroatoms. The heterocyclyl group may be saturated or partially unsaturated, i.e. possess one or more double bonds. Particularly preferred heterocyclyl are 5-6 and bicyclic 9-10 membered heterocyclyl. Suitable examples of heterocyclyl groups may include azridinyl, oxiranyl, thiiranyl, azetidinyl, oxetanyl, thietanyl, 2H-pyrrolyl, pyrrolidinyl, pyrrolinyl, piperidyl, piperazinyl, morpholinyl, indolinyl, imidazolidinyl, imidazolinyl, pyrazolidinyl, thiomorpholinyl, dioxanyl, tetrahydrofuranyl, tetrahydropyranyl, tetrahydropyrrolyl, tetrahydrothiophenyl, pyrazolinyl, dioxalanyl, thiazolidinyl, isoxazolidinyl, dihydropyranyl, oxazinyl, thiazinyl, thiomorpholinyl, oxathianyl, dithianyl, trioxanyl, thiadiazinyl, dithiazinyl, trithianyl, azepinyl, oxepinyl, thiepinyl, indenyl, indanyl, 3H-indolyl, isoindolinyl, 4H-quinolazinyl, chromenyl, chromanyl, isochromanyl, pyranyl and dihydropyranyl. A heterocyclyl group may be optionally substituted by one or more optional substituents as herein defined.

The term "heteroaryl" includes any of monocyclic, polycyclic, fused or conjugated hydrocarbon residues, wherein one or more carbon atoms are replaced by a heteroatom so as to provide an aromatic residue. Preferred heteroaryl have 3-20 ring atoms, e.g. 3-10. Particularly preferred heteroaryl are 5-6 and 9-10 membered bicyclic ring systems. Suitable heteroatoms include, O, N, S, P and Se, particularly O, N and S. Where two or more carbon atoms are replaced, this may be by two or more of the same heteroatom or by different heteroatoms. Suitable examples of heteroaryl groups may include pyridyl, pyrrolyl, thienyl, imidazolyl, furanyl, benzothienyl, isobenzothienyl, benzofuranyl, isobenzofuranyl, indolyl, isoindolyl, pyrazolyl, pyrazinyl, pyrimidinyl, pyridazinyl, indolizinyl, quinolyl, isoquinolyl, phthalazinyl, 1,5-naphthyridinyl, quinozalinyl, quinazolinyl, quinolinyl, oxazolyl, thiazolyl, isothiazolyl, isoxazolyl, triazolyl, oxadialzolyl, oxatriazolyl, triazinyl, tetrazolyl and furazanyl. A heteroaryl group may be optionally substituted by one or more optional substituents as herein defined.

The term "acyl" either alone or in compound words denotes a group containing the moiety C=O (and not being a carboxylic acid, ester or amide) Preferred acyl includes C(O)-R, wherein R is hydrogen or an alkyl, alkenyl, alkynyl, aryl, heteroaryl, carbocyclyl, or heterocyclyl residue. Examples of acyl include formyl, straight chain or branched alkanoyl (eg. C₁^o) such as, acetyl, propanoyl, butanoyl, 2-methylpropanoyl, pentanoyl, 2,2-dimethylpropanoyl, hexanoyl, heptanoyl, octanoyl, nonanoyl, decanoyl, undecanoyl, dodecanoyl, tridecanoyl, tetradecanoyl, pentadecanoyl, hexadecanoyl, heptadecanoyl, octadecanoyl, nonadecanoyl and icosanoyl; cycloalkylcarbonyl such as

• cyclopropylcarbonyl cyclobutylcarbonyl, cyclopentylcarbonyl and cyclohexylcarbonyl; aroyl such as benzoyl, toluoyl and naphthoyl; aralkanoyl such as phenylalkanoyl (e.g. phenylacetyl, phenylpropanoyl, phenylbutanoyl, phenylisobutylyl, phenylpentanoyl and phenylhexanoyl) and naphthylalkanoyl (e.g. naphthylacetyl, naphthylpropanoyl and naphthylbutanoyl]; aralkenoyl such as phenylalkenoyl (e.g. phenylpropenoyl, phenylbutenoyl, phenylmethacryloyl, phenylpentenoyl and phenylhexenoyl and naphthylalkenoyl (e.g. naphthylpropenoyl, naphthylbutenoyl and naphthylpentenoyl); aryloxyalkanoyl such as phenoxyacetyl and phenoxypropionyl; arylthiocarbanioyl such as phenylthiocarbamoyl; arylglyoxyloyl such as phenylglyoxyloyl and naphthylglyoxyloyl; arylsulfonyl such as phenylsulfonyl and napthylsulfonyl; heterocycliccarbonyl; heterocyclicalkanoyl such as thienylacetyl, thienylpropanoyl, thienylbutanoyl, thienylpentanoyl, thienylhexanoyl, thiazolylacetyl, thiadiazolylacetyl and tetrazolylacetyl; heterocyclicalkenoyl such as heterocyclicpropenoyl, heterocyclicbutenoyl, heterocyclicpentenoyl and heterocyclichexenoyl; and heterocyclicglyoxyloyl such as thiazolyglyoxyloyl and thienylglyoxyloyl. The R residue may be optionally substituted as described herein.

In this specification "optionally substituted" is taken to mean that a group may or may not be further substituted or fused (so as to form a condensed polycyclic group) with one, two, three or more of organic and inorganic groups, including those selected from: alkyl, alkenyl, alkynyl, carbocyclyl, aryl, heterocyclyl, heteroaryl, acyl, aralkyl, alkylaryl, alkylheterocyclyl, alkylheteroaryl, alkylcarbocyclyl, halo, haloalkyl, haloalkenyl, haloalkynyl, haloaryl, halocarbocyclyl, haloheterocyclyl, haloheteroaryl, haloacyl, haloaryalkyl, hydroxy, hydroxyalkyl, hydroxyalkenyl, hydroxyalkynyl, hydroxycarbocyclyl, hydroxyaryl, hydroxyheterocyclyL hydroxyheteroaryl, hydroxyacyl, hydroxyaralkyl, alkoxyalkyl, alkoxyalkenyl, alkoxyalkynyl, alkoxycarbocyclyl, alkoxyaryl, alkoxyheterocyclyl, alkoxyheteroaryl, alkoxyacyl, alkoxy aralkyl, alkoxy, alkenyloxy, alkynyloxy, aryloxy, carbocyclyloxy, aralkyloxy, heteroaryloxy, heterocyclyloxy, acyloxy, haloalkoxy, haloalkenyloxy, haloalkynyloxy, haloaryloxy, halocarbocyclyloxy, haloaralkyloxy, haloheteroaryloxy, haloheterocyclyloxy, haloacyloxy, nitro, nitroalkyl, nitroalkenyl, nitroalkynyl, nitroaryl, nitroheterocyclyl, nitroheteroaryl, nitrocarbocyclyl, nitroacyl, nitroaralkyl, amino (NH₂), alkylamino, dialkylamino, alkenylamino, alkynylamino, arylamino, diarylamino, aralkylamino, diaralkylamino, acylamino, diacylamino, heterocyclamino, heteroarylamino, carboxy, carboxyester, amido, alkylsulphonyloxy, arylsulphenyloxy, alkylsulphenyl, arylsulphenyl, thio, alkylthio, alkenylthio, alkynylthio, arylthio, aralkylthio, carbocyclylthio, heterocyclylthio, heteroarylthio, acylthio, sulfoxide, sulfonyl, sulfonamide, aminoalkyl, aminoalkenyl, aminoalkynyl, aminocarbocyclyl, aminoaryl, aminoheterocyclyl, aminoheteroaryl, aminoacyl, aminoaralkyl, thioalkyl, thioalkenyl, thioalkynyl, thiocarbocyclyl, thioaryl, thioheterocyclyl, thioheteroaryl, thioacyl, thioaralkyl, carboxyalkyl, carboxyalkenyl, carboxyalkynyl, carboxycarbocyclyl, carboxyaryl, carboxyheterocyclyl, carboxyheteroaryl, carboxyacyl, carboxyaralkyl, carboxyesteralkyl, carboxyesteralkenyl, carboxyesteralkynyl, carboxyestercarbocyclyl, carboxyesteraryl, carboxyesterheterocyclyl, carboxyesterheteroaryl, carboxyesteracyl, carboxyesteraralkyl, amidoalkyl, amidoalkenyl, amidoalkynyl, amidocarbocyclyl, amidoaryl, amidoheterocyclyl, amidoheteroaryl, amidoacyl, amidoaralkyl, formylalkyl, formylalkenyl, formylalkynyl, formylcarbocyclyl, formylaryl, formylheterocyclyl, formylheteroaryl, formylacyl, formylaralkyl, acylalkyl, acylalkenyl, acylalkynyl, acylcarbocyclyl, acylaryl, acylheterocyclyl, acylheteroaryl, acylacyl, acylaralkyl, sulfoxidealkyl, sulfoxidealkenyl, sulfoxidealkynyl, sulfoxidecarbocyclyl, sulfoxidearyl, sulfoxideheterocyclyl, sulfoxideheteroaryl, sulfoxideacyl, sulfoxidearalkyl, sulfonylalkyl, sulfonylalkenyl, sulfonylalkynyl, sulfonylcarbocyclyl, sulfonylaryl, sulfonylheterocyclyl, sulfonylheteroaryl, sulfonylacyl, sulfonylaralkyl, sulfonamidoalkyl, sulfonamidoalkenyl, sulfonamidoalkynyl, sulfonamidocarbocyclyl, sulfonamidoaryl, sulfonamidoheterocyclyl, sulfonamidoheteroaryl, sulfonamidoacyl, sulfonamidoaralkyl, nitroalkyl, nitroalkenyl, nitroalkynyl, nitrocarbocyclyl, nitroaryl, nitroheterocyclyl, nitroheteroaryl, nitroacyl, nitroaralkyl, cyano, sulfate, sulfonate, phosphonate and phosphate groups. Optional substitution may also be taken to refer to where a CH₂ group in a chain or ring is replaced by a carbonyl group (C=O) or where 2 adjacent carbon atoms are substituted by one end each of a -O-(CH₂)_n-O- or -NH-(CH2)_n-NH- group, wherein n is 1 or 2.

Examples of optional substitutents include alkyl, (e.g. C_1-6alkyl such as methyl, ethyl, propyl, butyl, cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl), hydroxyalkyl (e.g. hydroxymethyl, hydroxyethyl, hydroxypropyl), alkoxyalkyl (e.g. methoxymethyl, methoxyethyl, methoxypropyl, ethoxymethyl, ethoxy ethyl, ethoxypropyl etc) alkoxy (e.g. Ci_-6 alkoxy such as methoxy, ethoxy, propoxy, butoxy, cyclopropoxy, cyclobutoxy), halo, trifluoromethyl, trichloromethyl, tribromomethyl, hydroxy, phenyl (which itself may be further substituted e.g., by one or more C_1-6alkyl, halo, hydroxy, hydroxyCi_-6alkyl, C_{1- 6}alkoxy, haloC_1-6alkyl, cyano, nitro, OC(O)C μβalkyl, and amino), benzyl (wherein benzyl itself may be further substituted e.g., by one or more of C_1-6alkyl, halo, hydroxy, hydroxyd-βalkyl, C_1-6alkoxy, haloCi_-6alkyl, cyano, nitro, OC(O)Ci_-6alkyl, and amino), phenoxy (wherein phenyl itself may be further substituted e.g., by one or more of C_1- βalkyl, halo, hydroxy, hydroxyC_1-6alkyl, C_1-OaIkOXy, haloC^alkyl, cyano, nitro, OC(O)Ci- ₆alkyl, and amino), benzyloxy (wherein benzyl itself may be further substituted e.g., by one or more of C_1-6alkyl, halo, hydroxy, hydroxyC_1-6alkyl, C_1-6alkoxy, haloCi-βalkyl, cyano, nitro, OC(O)C _1-6alkyl, and amino), amino, alkylamino (e.g. C_1-6alkyl, such as methylamino, ethylamino, propylamino etc), dialkylamino (e.g. C_1-6alkyl, such as dimethylamino, diethylamino, dipropylamino), acylamino (e.g. NHC(O)CH₃), phenylamino (wherein phenyl itself may be further substituted e.g., by one or more of C_{1- 6}alkyl, halo, hydroxy hydroxyCi^alkyl, Ci_-6alkoxy, haloC_1-6alkyl, cyano, nitro, OC(O)Ci- ₆alkyl, and amino), nitro, formyl, -C(O)-alkyl (e.g. C_1-6 alkyl, such as acetyl), O-C(O)- alkyl (e.g. C^alkyl, such as acetyloxy), benzoyl (wherein benzyl itself may be further substituted e.g., by one or more of C_1-6alkyl, halo, hydroxy hydroxyC_1-6alkyl, C^alkoxy, haloC_1-6alkyl, cyano, nitro, 0C(0)Ci-₆alkyl, and amino), benzoyloxy (wherein benzyl itself may be further substituted e.g., by one or more of C_halky!, halo, hydroxy hydroxyCμ ₆alkyl, C₁^aIkOXy, haloC_t-galkyl, cyano, nitro, 0C(0)Ci-₆alkyl, and amino), CO₂H, C0₂alkyl (e.g. C_1-6 alkyl such as methyl ester, ethyl ester, propyl ester, butyl ester), COaphenyl (wherein phenyl itself may be further substituted e.g., by one or more of C_{1- 6}alkyl, halo, hydroxy, hydroxyCi.₆alkyl, C_1-6alkoxy, haloCi_-6alkyl, cyano, nitro, OC(O)C_{1- 6}alkyL and amino), CC^benzyl (wherein benzyl itself may be further substituted e.g., by one or more of C_1-6alkyl, halo, hydroxy, hydroxyC^ealkyl, Ci_-6alkoxy, haloC_1-6alkyl, cyano, nitro, 0C(O)C_1-6alkyl, and amino), CONH₂, CONHphenyl (wherein phenyl itself may be further substituted e.g., by one or more of C_1-6alkyl, halo, hydroxy, hydroxyd, ₆alkyl, C_1-6alkoxy, haloC^alkyl, cyano, nitro, OC(O)Ci_₆alkyl₅ and amino), CONHbenzyl (wherein benzyl itself may be further substituted e.g., by one or more of C_1-6alkyl, halo, hydroxy hydroxyC_1-6alkyl, C_1-6alkoxy, haloC_1-6alkyl, cyano, nitro, OC(O)C _1-6alkyl, and amino), CONHalkyl (e.g. C_1-6 alkyl such as methyl ester, ethyl ester, propyl ester, butyl amide) CONHdialkyl (e.g.

6alkyl- and

thioalkyl (e.g., HSd.₆alkyl-), carboxyalkyl (e.g., Hθ₂CC_1-6alkyl-), carboxyesteralkyl (e.g., C_1-6alkylO₂CC_1-6alkyl-), amidoalkyl (e.g., H₂N(O)CC i-βalkyl-, H(C₁.₆alkyl)N(O)CC₁.₆alkyl-), formylalkyl (e.g., OHCCi_-6alkyl-), acylalkyl (e.g., C_1-6alkyl(O)CC_1-6alkyl-), nitroalkyl (e.g., O₂NCι.₆a\kyl-), sulfoxidealkyl (e.g., R(O)Sd.₆alkyl, such as C_1-6alkyl(O)SC_1-6alkyl-), sulfonylalkyl (e.g., R(O)₂SC_{1- 6}alkyl- such as C_1-6alkyl(O)₂SC_1-6alkyl-), sulfonamidoalkyl (e.g., H₂N(O)SC _1-6alkyl, H(Ci- ₆alkyl)NS(O)C_1-6alkyl-), replacement of CH₂ with C=O, and where 2 adjacent carbon atoms are substituted by one end each of a -0-(CHa)_n-O- or -NH-(CH₂)_n-NH- group, wherein n is 1 or 2.

The term "sulfoxide", either alone or in a compound word, refers to a group -S(O)R wherein R is selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, heterocyclyl, carbocyclyl, and aralkyl, each of which may be optionally substituted. Examples of R include hydrogen, C_1-20alkyl, phenyl and benzyl.

The term "sulfonyl", either alone or in a compound word, refers to a group S(O)₂-R, wherein R is selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, heterocyclyl, carbocyclyl, acyl, and aralkyl, each of which may be optionally substituted. Examples of R include hydrogen, C^oalkyl, phenyl and benzyl.

The term "sulfonamide", either alone or in a compound word, refers to a group S(O)₂NRR wherein each R is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, heterocyclyl, carbocyclyl, acyl, and aralkyl, each of which may be optionally substituted. Examples of R include C_1-20alkyl, phenyl and benzyl. In one embodiment at least one R is hydrogen. In another embodiment, both R are hydrogen.

A "sulfate" group refers to a group OS(O)₂OR wherein each R is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, heterocyclyl, carbocyclyl, acyl, and aralkyl, each of which may be optionally substituted. Examples of R include hydrogen, C_1-2oalkyl, phenyl and benzyl.

The term "sulfonate" refers to a group OS(O)₂R wherein each R is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, heterocyclyl, carbocyclyl, acyl, and aralkyl, each of which may be optionally substituted. Examples of R include hydrogen, C_1-20alkyl, phenyl and benzyl.

The term "phosphonate" refers to a group P(O)(OR₂) wherein R is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, heterocyclyl, carbocyclyl, acyl, and aralkyl, each of which may be optionally substituted. Examples of R include hydrogen, C^oalkyl, phenyl and benzyl.

The term "phosphate" refers to a group OP(O)(OR)₂ wherein R is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, heterocyclyl, carbocyclyl, acyl, and aralkyl, each of which may be optionally substituted. Examples of R include hydrogen, C_1-2oalkyl, phenyl and benzyl.

The term "thio" is intended to include groups of the formula "SR" wherein R can be hydrogen (thiol), alkyl, alkenyl, alkynyl, aryl, carbocyclyl, heteroaryl, heterocyclyl, aralkyl, and acyl, each of which may be optionally substituted. Examples of R include hydrogen, C_1-2oalkyl, phenyl and benzyl.

The term, "amino" is used here in its broadest sense as understood in the art and includes groups of the formula NR^AR^B wherein R^A and R^B may be any independently selected from hydrogen, hydroxy alkyl, alkoxyalkyl, alkenyl, alkynyl, aryl, carbocyclyl, heteroaryl, heterocyclyl, aralkyl, and acyl, each of which may be optionally substituted. R^A and R^B, together with the nitrogen to which they are attached, may also form a monocyclic, or polycyclic ring system e.g. a 3-10 membered ring, particularly, 5-6 and 9-10 membered systems. Examples of "amino" include NH₂, NHalkyl (e.g. Ci-₂₀alkyl), NHalkoxyalkyl, NHaryl (e.g. NHphenyl), NHaralkyl (e.g. NHbenzyl), NHacyl (e.g. NHC(O)C _1-20alkyl, NHC(O)phenyl), Ndialkyl (wherein each alkyl, for example C_1-2O, may be the same or different) and 5 or 6 membered rings, optionally containing one or more same or different heteroatoms (e.g. O, N and S). Reference to groups written as "[group] amino" is intended to reflect the nature of the R^A and R^B groups. For example, "alkylamino" refers to NR^AR^B where one of R^A or R^B is alkyl. "Dialkylamino" refers to NR^AR^B where R^A and R^B are each (independently) an alkyl group.

The term "amido" is used here in its broadest sense as understood in the art and includes groups having the formula C(O)NR^AR^B, wherein R^A and R^B are as defined as above.

Examples of amido include C(O)NH₂, C(O)NHalkyl (eg C_1-2Oalkyl), C(O)NHaryl (eg

C(O)NHphenyl), C(O)NHaralkyl (eg C(O)NHbenzyl), C(O)NHacyl (eg C(O)NHC(O)C₁.

_2Oalkyl, C(O)NHC(O)phenyl), C(O)Nalkylalkyl (wherein each alkyl, for example C_1-20, may be the same or different) and 5 or 6 membered rings, optionally containing one or more same or different heteroatoms (eg O, N and S).

In a further form of "amido", particularly when R⁷ is such, -NR^AR^B may be the amino terminus of an amino acid or peptide molecule. Such "amido" groups can be formed by coupling the amino terminus of an amino acid or peptide molecule with a precursor carboxylic acid group in the usual manner. As referred to herein a peptide molecule comprises 2, 3, 4, 5 or more same or different amino acid molecules covalently linked by peptide or amide bonds formed between the amino terminus of one amino acid and the carboxylic acid terminus of another. The amino acids contemplated include natural amino acids and non-natural amino acids, and modified and protected forms thereof. The term "carboxy ester" is used here in its broadest sense as understood in the art and includes groups having the formula CO₂R, wherein R may be selected from groups including alkyl, alkenyl, alkynyl, aryl, carbocyclyl, heteroaryl, heterocyclyl, aralkyl, and acyl, each of which may be optionally substituted. Examples of carboxy ester include CQA-aoalkyl, CO₂aryl (eg. CO₂ρhenyl), CO₂aralkyl (eg CO₂ benzyl).

"Saccharide" groups contemplated herein include mono-, di- and tri-saccharides of pyranose and furanose groups. In preferred embodiments there are linked through Cl. Examples include, glucose, idose, allose, tallose, mannose, galactose, maltose, lactose, arabinose, glucosamine, ribose, xylose, cellobiose, maltotriose, cellotriose, isomaltotriose. In one embodiment, the saccharides are monosaccharides. In a preferred embodiment, the saccharides are the pyranoses. In a particularly preferred embodiment, the saccharide group is glucose. It will be understood that the saccharide groups can exist in their "free hydroxyl" or "free amino" form or one or more, or all, of the hydroxyl and/or amino groups can be protected by a suitable protecting group (eg acetate groups). A saccharide group may be removed and/or replaced by known methods in the art.

Reference to the following numbering system will be used throughout:

Compounds contemplated by the invention may be obtained by a variety of means including isolation from natural sources, synthetic methods for the preparation of isoquinolines/tetrahydro-β-carbolines and dehydro forms thereof, or by enzyme catalysed coupling. Thus, naturally occurring compounds such as strictosidine (i), strictosidinic acid (ii), 5(S)- 5-carboxystrictosidine (iii) and 3,4-dehydro-5(S)-5-carboxystrictosidine (iv) (see below) can be isolated from their natural sources in accordance with described procedures (see for example, Kitajima, M. et al., 2002; Arbain, D. et al, 1993 and Stδckigt, J. et al, 1977). Once isolated, further chemical manipulations may be performed, e.g. acylation, alkylation, arylation or benzylation of some or all of the free hydroxy or amino groups, amidation or esterification of carboxylic acid groups, de-esterification of ester groups, reduction of carboxylic ester or acid groups to form aldehydes or removal of the glucosyl group and optionally replacing it with another saccharide moiety or other non-H Y group as defined herein. Methods for performing such chemical manipulations or transformations are known in the art and are extensively described, for example, Larock, R.E., 1989.

(i) R = H₅ R' = OMe (iv) (ii) R = H, R¹ = OH (iii) R = CO₂H₅ R' = OMe

Alternatively, compounds where - represents a single bond can be prepared by acid- mediated or strictosidine synthase-catalysed coupling of an appropriately substituted/protected tryptamine compound and secologanin, or derivative thereof, as depicted, for example in general Scheme I below.

Scheme I

Compounds of Formula (I), wherein "" is a single bond can be prepared in a stereoselective manner (at C3 of the resulting carbinol) by strictosidane synthase coupling of (1) and (2) in an analogous manner to that described by Patthy-Lukats, A. et ah, 1999 and Treimer, J.F. et ah, 1979, where, in the presence of strictosidine synthase, the coupling of tryptamine and secologanin afforded strictosidine with complete stereoselectivity (Scheme II) :

Scheme II

Alternatively, appropriate tryptamine and secologanin compounds may be coupled under acidic conditions (Pictet Spengler reaction) such as described in US 6,720,331, Patthy- Lukats, A. et ah, 1999 and The Merck Index, Thirteenth Edition. Compounds where ^'• ^'-"-" is a double bond, referred to herein as 3,4-dehydro compounds, may be prepared by coupling an appropriate tryptamine compound and a modified secologanin compound, wherein the aldehyde has been oxidized to a carboxylic acid group, followed by dehydrating ring closure in accordance with the methods described by US 6,350,757 and Kitajima, M. et al, 2002, and as described below in general Scheme III. Thus, coupling of (1) and (2) followed by ring closure of (3) with a suitable dehydrating agent such as DCC or POCl₃, yields the 3,4-dehydro compounds (4).

Alternatively, 3,4-dehydro compounds may be prepared from the 3,4-dihydro compounds by appropriate oxidative treatment, such as treatment with DDQ (see for example US 6,350,757 or US 6,720,331).

Starting tryptamine compounds may be prepared by any one of the many methods known in the art for indole synthesis, or alternatively, may be obtained from commercial sources. Some examples of starting tryptamines contemplated by the present invention include:

Secologanin may be obtained commercially from Sigma (Fluka), USA, or isolated from natural sources (Galan et al, 2006). It may be modified prior to coupling, e.g. removal or replacement of the glucose moiety, protection of free OH groups, oxidation to the carboxylic acid (for synthesis of 3,4-dehydro compounds) or modification of the ester moiety. Alternatively, the vinyl group may be modified in accordance with the olefin cross metathesis methods described by Galan et al., 2006, to afford further secologanin analogues. In some particularly preferred embodiments obtained thereby, Z is selected from (trans)CH=CH-R or (CHa)₂R wherein R is Cnoalkyl, for example t-butyl, n-butyl, n- pentyl, and n-heptyl.

In one embodiment of the invention, a preferred compound for coupling to tryptamine compounds is secologanin itself (X = OMe, Y = GIc, Z = CH=CH₂). (optionally protected) or the oxidised carboxylic acid form, e.g.

R = H or OH

R¹ = H or protecting group

Other preferred secologanin-type compounds for coupling include the aglycone form (or protected form thereof) of those described above.

It will be recognised that in some circumstances it may be appropriate to perform any chemical manipulations on the tryptamine and/or secologanin compounds prior to coupling. In others, for example, where the modification could hinder or block the coupling, the appropriate modification can be performed after coupling.

It will be recognised that during the processes for the preparation of compounds contemplated by the present invention, it may be necessary or desirable to protect certain functional groups which may be reactive or sensitive to the reaction or transformation conditions undertaken (eg OH (including diols), NH₂, CO₂H, SH, C=O). Suitable protecting groups for such functional groups are known in the art and may be used in accordance with standard practice. As used herein, the term "protecting group", refers to an introduced functionality which temporarily renders a particular functional group inactive under certain conditions. Such protecting groups and methods for their installation and subsequent removal at an appropriate stage are described in Protective Groups in Organic Chemistry, 3^rd Edition, T.W.Greene and P. G. Wutz, John Wiley and Sons, 1999, the entire contents of which are incorporated herein by reference. Exemplary forms of protected groups include: for amino (NH₂) - carbamates (such as Cbz, Boc, Fmoc), benzylamines, acetamides (e.g. acetamide, trifluoroacetamide); for carbonyl - acetals, ketals, dioxanes, dithianes, and hydrazones; for hydroxy - ethers (e.g. alkyl ethers, alkoxylalkyl ethers, allyl ethers, silyl ethers, benzyl ethers, tetrahydropyranyl ethers), carboxylic acid esters, acetals (e.g. acetonide and benzylidene acetal); for thio (SH) -ethers (e.g. alkyl ethers, benzyl ethers), esters for CO₂H - esters (e.g. alkyl esters, benzyl esters).

It will also be recognised that certain compounds of formula (I) may possess asymmetric centres and are therefore capable of existing in more than one stereoisomeric form, such as enantiomers and diastereomers. The invention thus also relates to compounds in substantially pure (e.g. about 95%, preferably at least 97%, or more preferably at least

99%) isomeric form at one or more asymmetric centres, as well as stereoisomeric mixtures.

Such isomers may be prepared by asymmetric synthesis, for example using chiral intermediates or reagents, enzymes, or mixtures may be resolved by conventional methods, eg., chromatography, recrystallisation or use of a resolving agent. In some embodiments of the invention, the compounds of Formula (I) have the same stereochemistry at 1, 2, 3 or all 4 of the chiral centres depicted for strictosidinic acid (i), or I₅ 2, 3 or all 4 chiral centres as depicted for 3,4-dehydro 5(S)-5-carboxystrictosidinic acid (iv).

In some embodiments of the invention, R_1-4 are independently selected from hydrogen, alkyl, (e.g. C^alkyl such as methyl, ethyl, propyl, butyl, cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl), hydroxyalkyl (e.g. hydroxymethyl, hydroxyethyl, hydroxypropyl), alkoxyalkyl (e.g. methoxymethyl, methoxyethyl, methoxypropyl, ethoxymethyl, ethoxyethyl, ethoxypropyl etc) alkoxy (e.g. C_1-6 alkoxy such as methoxy, ethoxy, propoxy, butoxy, cyclopropoxy, cyclobutoxy), alkylthio (e.g. MeS-, EtS-, PrS-, BuS-), halo, trifluoromethyl, trichloromethyl, tribromomethyl, hydroxy, phenyl (which itself may be further substituted e.g., by Ci_-6alkyl, halo, hydroxy, hydroxyC_1-6alkyl, C₁. ₆alkoxy, haloC_1-6alkyl, cyano, nitro OC(O)C _1-6alkyl, and amino), benzyl (wherein benzyl itself may be further substituted e.g., by one or more of C_halky!, halo, hydroxy, hydroxyCi._δalkyl, Cμβalkoxy, haloCi.₆alkyl, cyano, nitro, OC(O)C_1-6alkyl, and amino), phenoxy (wherein phenyl itself may be further substituted e.g., by one or more of C₁. ₆alkyl, halo, hydroxy,

cyano, nitro, OC(O)C₁. ₆alkyl, and amino), benzyloxy (wherein benzyl itself may be further substituted e.g., by one or more of Ci_-6alkyl, halo, hydroxy,

C_1-6alkoxy, haloC_1-6alkyl, cyano, nitro, OC(O)C₁.₆alkyl, and amino), amino, alkylamino (e.g. C_1-6alkyl, such as methylamino, ethylamino, propylamino etc), dialkylamino (e.g. C_1-6alkyl, such as dimethylamino, diethylamino, dipropylamino), acylamino (e.g. NHC(O)CHs), phenylamino (wherein phenyl itself may be further substituted e.g., by one or more Of C₁. ₆alkyl, halo, hydroxy hydroxyC^ealkyl, Ci_-6alkoxy, haloCi-βalkyl, cyano, nitro, OC(O)C₁. ₆alkyl, and amino), nitro, formyl, -C(O)-alkyl (e.g. C_1-6 alkyl, such as acetyl), 0-C(O)- alkyl (e.g. C^alkyl, such as acetyloxy), benzoyl (wherein benzyl itself may be further substituted e.g., by one or more of C^ancyl, halo, hydroxy hydroxyC_1-6alkyl, C_1-6alkoxy, halod.₆alkyl, cyano, nitro, OC(O)C μβalkyl, and amino), benzoyloxy (wherein benzyl itself may be further substituted e.g., by one or more of C_1-6alkyl, halo, hydroxy hydroxyCi- ₆alkyl, C_1-6alkoxy, haloC_1-6alkyl, cyano, nitro, 0C(0)d-₆alkyl, and amino), CO₂H, Cθ2alkyl (e.g. C_1-6alkyl such as methyl ester, ethyl ester, propyl ester, butyl ester), Cθ2phenyl (wherein phenyl itself may be further substituted e.g., by one or more of Ci- ₆alkyl, halo, hydroxy, hydroxyC_t-δalkyl, C_1-6alkoxy, haloCi_-6alkyl, cyano, nitro, OC(O)C₁. ₆alkyl, and amino), COibenzyl (wherein benzyl itself may be further substituted e.g., by one or more of C^alkyl, halo, hydroxy, hydroxyC_1-6alkyl, C_1-6alkoxy, haloC_1-6alkyl, cyano, nitro, OC(O)C _1-6alkyl, and amino), CONH₂, CONHphenyl (wherein phenyl itself may be further substituted e.g., by one or more of C_1-6alkyl, halo, hydroxy, hydroxyCϊ. βalkyl, C_1-6alkoxy, haloC_1-6alkyl, cyano, nitro, OC(O)C_1-6alkyl, and amino), CONHbenzyl (wherein benzyl itself may be further substituted e.g., by one or more of Ci-βalkyl, halo, hydroxy hydroxyC_1-6alkyl,

haloCi_-6alkyl, cyano, nitro, OC(O)C _1-6alkyl, and amino), CONHalkyl (e.g. C_1-6 alkyl such as methyl ester, ethyl ester, propyl ester, butyl amide) CONdialkyl (e.g. C_1-6alkyl) aminoalkyl (e.g., HNC_1-6alkyl-, C_1-6alkylHN-C_1-6alkyl- and (C_1-6alkyl)₂N-C_1-6alkyl-), thioalkyl (e.g., HSC_1-6alkyl-), carboxyalkyl (e.g., HO₂CCi- βalkyl-), carboxyesteralkyl (e.g., C₁.₆alkyl0₂CC_1-6alkyl-), amidoalkyl (e.g., H₂N(O)CC_1- ealkyl-, H(C_1-6alkyl)N(O)CC₁._όalkyl-), formylalkyl (e.g., OHCC^alkyl-), acylalkyl (e.g.,

nitroalkyl (e.g., O₂NC_1-6alkyl-), sulfoxidealkyl (Cg₁₅ R(O)SC_1- ealkyl, such as C₁.₆alkyl(O)SC₁.₆alkyl-), sulfonylalkyl (e.g., R(O)₂SC i_-6alkyl- such as C_{1- 6}alkyl(O)₂SC_1-6alkyl-), sulfonamidoalkyl (e.g., ₂HN(O)₂SC_1-6alkyl, H(C₁.₆alkyl)N(O)₂SCi- ₆alkyl-), and where 2 adjacent carbon atoms are substituted by one end each of a -O- (CH₂)_n-0- or -NH-(CH₂)_n-NH- group, wherein n is 1 or 2.

In some embodiments of the invention, at least one of R₁-R₄ is hydrogen, for example, at least 2 are hydrogen. In other embodiments, 3 or all 4 OfR₁-R₄ are hydrogen.

In some embodiments of the invention, R₅ and R₆ are selected from hydrogen, optionally substituted C_1-6alkyl, optionally substituted C(O)C ^alkyl, optionally substituted C(O)phenyl, optionally substituted C(0)beri2yl.

In some embodiments of the invention, R₇ is selected from hydrogen, CO₂H, CO₂C_1-6alkyl₅ e.g. methyl, ethyl, propyl or butyl), CONH₂, CONHC_1-6alkyl, CONHaryl. In particularly preferred embodiments, R₇ is selected from H and CO₂H.

In some embodiments of the invention, Rg is hydrogen.

In some embodiments of the invention, at least one of R_7a and Rg_a is hydrogen, for example, both are hydrogen. In some embodiments of the invention, X include OH, OCi.ioalkyl, NHC^oalkyl, NHphenyl and NHbenzyl.

In some embodiments of the invention, Y is H or a saccharide, such as glucose.

In some embodiments of the invention, Z is CH₂=CH₂ or C^oalkyl.

Some non-limiting examples of compounds contemplated by the invention include combinations of any two, three, four, five, six, seven or eight of the embodiments for R_1-4, R₅ and R₆, R₇, R₈, R_7a and Rg₈, X, Y and Z described above.

In a particular embodiment, the compound for use in the invention is strictosidinic acid.

Strictosidinic acid can be isolated from natural sources, in accordance with the procedures described in the literature, for example, described by Arbain et ah, 1993; Yamakazi et ah, 2003 and Geerlings et al. , 2000, and in accordance with the Examples described herein. Thus, in certain embodiments, strictosidinic acid, or other compounds contemplated herein are used in extracted, isolated or substantially purified form (for example, at least 50, 75, 90, 95 or 99% pure).

As used herein, "regulating glucose homeostasis" refers to the regulation or control of blood glucose levels to lower hyperglycaemic, or preferably achieve or maintain normal fasting state, blood glucose levels. Normal fasting state blood glucose levels are typically less than 6.1 mmolL^" (110 mgdL^"1). Hyperglycaemic levels refer to blood glucose levels greater than or equal to 6.1 mmolL^'1 (110 mgdL^"1).

Impaired fasting glycemia (IFG) is characterised by a fasting plasma glucose concentration greater than or equal to 6.1 mmol (110 mgdL^"1) but less than 7.0 mmolL^"1 (126 mgdL^"1) and a 2-h plasma glucose concentration during the oral glucose tolerance test (OGTT) (if measured) less than 7.8 mmolL^"1 (140 mgdL^"1). Impaired glucose tolerance (IGT) is characterised by a fasting plasma glucose concentration of less than 7.0 mmolL^"1 (126 mgdL^"1) and a 2-h plasma glucose concentration during the OGTT of greater than or equal to 7.8 mmolL^'1 (140 mgdL^"1) but less than 11.1 mmolL^"1 (200 mgdL^"1). Diabetes is characterised by a fasting plasma glucose concentration of greater than or equal to 7.0 mmolL^"1 (126 mgdL^'1) or a 2-h plasma glucose concentration during the OGTT of greater than 11.1 mmolL^'1 (200 mgdL^"1).

The compounds of the invention may have utility in the therapeutic or prophylactic treatment of diseases and conditions, and /or their symptoms, in which insulin resistance in involved or implicated in a subject. Any disease or condition, or symptom thereof in which insulin resistance or impaired glucose uptake by a cell or tissue can be attributed, or play a role or is manifested is contemplated herein. Non-limiting examples include NIDDM, gestational diabetes, impaired glucose tolerance, impaired fasting glucose, Syndrome X, hyperglycemia, obesity, cardiovasular disease, atheriosclerosis, hypertriglyceridemia, dyslipidemia, hyperinsulinemia, nephropathy, neuropathy, retinopathy, ischemia, stroke and fatty liver disease.

Subjects to be treated include mammalian subjects: humans, primates, livestock animals (including cows, horses, sheep, pigs and goats), companion animals (including dogs, cats, rabbits, guinea pigs), and captive wild animals. Laboratory animals such as rabbits, mice, rats, guinea pigs and hamsters are also contemplated as they may provide a convenient test system. Non-mammalian species such as birds, amphibians and fish may also be contemplated in certain embodiments of the invention. A particularly contemplated subject is a human subject.

The compounds of the invention are administered in an amount which, when administered according to the desired dosing regimen, at least partially attains the desired therapeutic effect, e.g. modulating glucose homeostasis, modulating cellular glucose uptake, treating insulin resistance, lowering blood glucose levels or treating a disease or condition, or symptom thereof, in which insulin resistance is involved. As used herein, treatment refers to therapeutic ameliorating or prophylactic treatment, including one or more of: alleviating or ameliorating the symptoms of, preventing or delaying the onset of, inhibiting the progression of, or halting or reversing altogether the onset or progression of the particular disorder or condition, or one or more symptoms thereof, being treated.

Suitable dosage amounts and dosing regimens can be determined by the attending physician and may depend on the particular condition being treated, the severity of the condition as well as the general age, health and weight of the subject, and may be in the range of from about 0.1 mg to about 1000 mg of active per day. The active ingredient may be administered in a single dose or a series of doses. Suitable dosages may contain about 1, 2.5, 5, 10, 20, 25, 50, 75, 100, 150, 200, 250 or 500 mg of active.

While it is possible for the active ingredient to be administered alone, it is preferable to present it as a composition, preferably as a pharmaceutical composition, with one or more pharmaceutically acceptable excipients or additives. Thus, the present invention also relates to the use of a compound of formula (I) or a pharmaceutically acceptable salt or prodrug thereof in the manufacture of a medicament for modulating glucose homeostasis, modulating cellular glucose uptake, treating insulin resistance, or lowering blood glucose levels.

The formulation of such compositions is well known to those skilled in the art, see for example, Remington's Pharmaceutical Sciences, 18^th Edition, Mack Publishing, 1990. The composition may contain any suitable carriers, diluents excipients or other additives.

These include all conventional solvents, dispersion media, fillers, solid carriers, coatings, antifungal and antibacterial agents, dermal penetration agents, surfactants, isotonic and absorption agents and the like. It will be understood that the compositions of the invention may also include other supplementary physiologically active agents.

The carrier 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 (including dermal, buccal and sublingual), vaginal or parental (including subcutaneous, intramuscular, intravenous and intradermal) administration. The compositions may conveniently be presented in unit dosage form and may be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the active ingredient with the carrier which constitutes one or more accessory ingredients. In general, the compositions are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both, and then if necessary shaping the product.

Compositions of the present invention suitable for oral administration may be presented as discrete units such as capsules, sachets or tablets each containing a predetermined amount of the active ingredient; as a powder or granules; as a solution or a suspension in an aqueous or non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion.

A tablet may be made by compression or moulding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with a binder (e.g inert diluent), preservative disintegrant (e.g. sodium starch glycolate, cross-linked polyvinyl pyrrolidone, cross-linked sodium carboxymethyl cellulose) surface- active or dispersing agent. Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein using, for example, hydroxypropylmethyl cellulose in varying proportions to provide the desired release profile. Tablets may optionally be provided with an enteric coating, to provide release in parts of the gut other than the stomach.

Compositions suitable for topical administration in the mouth include lozenges comprising the active ingredient in a flavoured base, usually sucrose and acacia or tragacanth gum; pastilles comprising the active ingredient in an inert basis such as gelatin and glycerin, or sucrose and acacia gum; and mouthwashes comprising the active ingredient in a suitable liquid carrier. Compositions suitable for topical administration to the skin may comprise the compounds dissolved or suspended in any suitable carrier or base and may be in the form of lotions, gel, creams, pastes, ointments and the like. Suitable carriers include mineral oil, propylene glycol, polyoxyethylene, polyoxypropylene, emulsifying wax, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water. Transdermal patches may also be used to administer the compounds of the invention.

Compositions for rectal administration may be presented as a suppository with a suitable base comprising, for example, cocoa butter, glycerin, gelatin or polyethylene glycol.

Compositions suitable for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or spray formulations containing in addition to the active ingredient such carriers as are known in the art to be appropriate.

Compositions suitable for parenteral administration include aqueous and non-aqueous isotonic sterile injection solutions which may contain anti-oxidants, buffers, bactericides and solutes which render the composition isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents. The compositions may be presented in unit-dose or multi-dose sealed containers, for example, ampoules and vials, and may be stored in a freeze-dried (lyophilised) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.

Preferred unit dosage compositions are those containing a daily dose or unit, daily sub- dose, as herein above described, or an appropriate fraction thereof, of the active ingredient. It should be understood that in addition to the active ingredients particularly mentioned above, the compositions of this invention may include other agents conventional in the art having regard to the type of composition in question, for example, those suitable for oral administration may include such further agents as binders, sweeteners, thickeners, flavouring agents disintegrating agents, coating agents, preservatives, lubricants and/or time delay agents. Suitable sweeteners include sucrose, lactose, glucose, aspartame or saccharine. Suitable disintegrating agents include corn starch, methylcellulose, polyvinylpyrrolidone, xanthan gum, bentonite, alginic acid or agar. Suitable flavouring agents include peppermint oil, oil of wintergreen, cherry, orange or raspberry flavouring. Suitable coating agents include polymers or copolymers of acrylic acid and/or methacrylic acid and/or their esters, waxes, fatty alcohols, zein, shellac or gluten. Suitable preservatives include sodium benzoate, vitamin E, alpha-tocopherol, ascorbic acid, methyl paraben, propyl paraben or sodium bisulphite. Suitable lubricants include magnesium stearate, stearic acid, sodium oleate, sodium chloride or talc. Suitable time delay agents include glyceryl monostearate or glyceryl distearate.

The present invention also relates to prodrugs of formula (I). Any compound that is a prodrug of a compound of formula (I) is within the scope and spirit of the invention. The term "prodrug" is used in its broadest sense and encompasses those derivatives that are converted in vivo, either enzymatically or hydrolytically, to the compounds of the invention. Such derivatives would readily occur to those skilled in the art, and include, for example, compounds where a free thiol or hydroxy group is converted into an ester, such as an acetate, or thioester or where a free amino group is converted into an amide. Procedures for acylating the compounds of the invention, for example to prepare ester and amide prodrugs, are well known in the art and may include treatment of the compound with an appropriate carboxylic acid, anhydride or chloride in the presence of a suitable catalyst or base. Esters of carboxylic acid (carboxy) groups are also contemplated. Suitable esters C_1-6alkyl esters; Cμ_δalkoxymethyl esters, for example methoxymethyl or ethoxymethyl; Ci_-6alkanoyloxymethyl esters, for example, pivaloyloxymethyl; phthalidyl esters; C₃.₈cycloalkoxycarbonylC_1-6alkyl esters, for example, 1- cyclohexylcarbonyloxyethyl; l,3-dioxolen-2-onylmethyl esters, for example, 5-methyl-l,3- dioxolen-2-onylmethyl; and Ci-βalkoxycarbonyloxyethyl esters, for example, 1- methoxycarbonyloxyethyl. Prodrugs of amino functional groups include amides (see, for example, Adv. BioSci., 1979, 20, 369, Kyncl, J. et al), enamines (see, for example, J. Pharm. Sci., 1971, 60, 1810, Caldwell, H. et al), Schiff bases (see, for example, US Patent No 2,923,661 and Antimicrob. Agents Chemother., 1981, 19, 1004, Smyth, R. et al), oxazolidines (see, for example, J. Pharm. Sci, 1983, 72, 1294, Johansen, M. et al), Mannich bases (see, for example, J. Pharm. Sci. 1980, 69, 44, Bundgaard, H. et al and J. Am. Chem. Soc, 1959, 81, 1198, Gottstein, W. et al), hydroxymethyl derivatives (see, for example, J. Pharm. Sci, 1981, 70, 855, Bansal, P. et al) and N-(acyloxy)alkyl derivatives and carbamates (see, for example, J. Med. Chem., 1980, 23, 469, Bodor, N. et al, J. Med. Chem., 1984, 27, 1037, Firestone, R. et al, J. Med. Chem., 1967, 10, 960, Kreiger, M. et al, US Patent No 5,684,018 and J. Med. Chem., 1988, 31, 318-322, Alexander, J. et al). Other conventional procedures for the selection and preparation of suitable prodrugs are known in the art and are described, for example, in WO 00/23419; Design of Prodrugs, H. Bundgaard, Ed., Elsevier Science Publishers, 1985; Methods in Enzymology, 42: 309-396, K. Widder, Ed, Academic Press, 1985; A Textbook of Drug Design and Development, Krogsgaard-Larsen and H. Bundgaard, Eds, Chapter 5, pi 13-191 (1991); Advanced Drug Delivery Reviews, 8; 1-38 (1992); Journal of Pharmaceutical Sciences, 77;285 (1988), H. Bundgaard, et al; Chem Pharm Bull, 32692 (1984), N. Kakeya et al and The Organic Chemistry of Drug Desig and Drug Action, Chapter 8, pp352-401, Academic press, Inc., 1992.

Suitable pharmaceutically acceptable salts of compounds of formula (I) include, but are not limited to salts of pharmaceutically acceptable inorganic acids such as hydrochloric, sulphuric, phosphoric nitric, carbonic, boric, sulfamic, and hydrobromic acids, or salts of pharmaceutically acceptable organic acids such as acetic, propionic, butyric, tartaric, maleic, hydroxymaleic, fumarie, maleic, citric, lactic, mucic, gluconic, benzoic, succinic, oxalic, phenylacetic, methanesulphonic, toluenesulphonic, benezenesulphonic, salicyclic sulphanilic, aspartic, glutamic, edetic, stearic, palmitic, oleic, lauric, pantothenic, tannic, ascorbic and valeric acids. Base salts include, but are not limited to, those formed with pharmaceutically acceptable cations, such as sodium, potassium, lithium, calcium, magnesium, ammonium and alkylammonium. Basic nitrogen-containing groups may be quarternised with such agents as lower alkyl halide, such as methyl, ethyl, propyl, and butyl chlorides, bromides and iodides; dialkyl sulfates like dimethyl and diethyl sulfate; and others.

The compounds of the invention may be in crystalline form either as the free compounds or as solvates and it is intended that both forms are within the scope of the present invention. The term "solvate" refers to a complex or aggregate formed by one or more molecules of a solute, ie compounds contemplated by the invention, and one or more molecules of a solvent. Suitable solvents are well understood in the art and include for example, of water, ie to form hydrates, and common organic solvents such as alcohols (methanol, ethanol, isopropanol) and acetic acid. Methods of solvation are generally known within the art, for example, recrystallisation from an appropriate solvent.

The methods, compounds and compositions of the invention may be utilised in conjunction with other anti-diabetic or anti-hyperglycaemic agents and may be administered, simultaneously (separately or as a combination) or sequentially in accordance with a suitable dosing and treatment regime as determined by the attending physician. Suitable agents may include insulin sensitisers, glucose resorption/uptake inhibitors and the classes and compounds identified in US 2005/0037981 , particularly Table 2, the contents of which are incorporated herein in their entirety.

The compounds of the invention may also be presented for use in veterinary compositions. These may be prepared by any suitable means known in the art. Examples of such compositions include those adapted for:

(a) oral administration, external application (eg drenches including aqueous and nonaqueous solutions or suspensions), tablets, boluses, powders, granules, pellets for admixture with feedstuffs, pastes for application to the tongue;

(b) parenteral administration, eg subcutaneous, intramuscular or intravenous injection as a sterile solution or suspension;

(c) topical application eg creams, ointments, gels, lotions etc. The invention will now be described with reference to the following examples which are provided for the purpose of illustrating certain embodiments of the invention and are not to be construed as limiting the generality hereinbefore described.

EXAMPLES

Example 1: Effect of Plant Extract

Plant Extract Preparation

The bark and leaves of Alphitonia zizqphoides, Morinda citrifolia, Neisoperma oppositifolium, Pometia pinnata, Rhus taitensis, Syzygium malaccense, Terminalia catappa and Thespia populnea were dried and ground to form the plant material for subsequent extraction.

The plant extract used for both in vivo and in vitro studies was prepared as follows: lOOg of ground, dried plant material above was mixed thoroughly with IL of distilled water and maintained at 70°C for 24h. The mixture was then filtered and allowed to cool to room temperature. For all experiments the plant extract was prepared that day, and any unused plant extract was discarded at the end of the day.

In Vivo Studies in Psammomys obesus

The effects of the plant extract were tested in both lean, non-diabetic and obese, type 2 diabetic P. obesus. Animals from within each group (n=l 0 each) were randomly assigned to plant extract treatment or control groups. All animals used in these studies were male and 18 weeks of age. The characteristics of the animals are given in Table 1.

Table 1: Characteristics of P. obesus

For the plant extract treatment group, 500ml of plant extract was mixed thoroughly with 1.5L dH₂O and 5kg ground rodent chow. The chow, now containing 1% (w/w) plant extract, was re-pelleted and dried. After a 7-day baseline period, animals in the plant extract treatment group received ad libitum access to the chow containing 1% plant extract for 11 days, while control animals received ad libitum access to chow re-pelleted and dried as above after being mixed with άϊlzO only.

Body weight and food intake were measured daily, while blood was collected from the tail vein for the measurement of glucose and insulin at the start and end of the 11 day experimental period. Treatment with the plant extract resulted in a significant reduction in blood glucose concentration in both lean, non-diabetic (by 10.5%; p=0.001) and obese, diabetic P. obesus (by 33.7%; p=0.024; Table 2). Treatment with the plant extract tended to reduce plasma insulin concentration in the plant extract treated groups (by 22% in the lean, non-diabetic and by 31% in the obese, diabetic animals; Table 2), however these changes were not statistically significant. Treatment with the plant extract for 11 days had a significant effect on the change in body weight throughout the study in both lean, non- diabetic and obese, diabetic P. obesus (Table 2). In the lean, non-diabetic animals, the plant extract treated group lost 1.6 + 2.8 g during the study period, while the control animals gained 7.4 ± 2.2 g (p=0.035). In obese, diabetic P. obesus, the plant extract treated group lost 9.4 + 4.1 g during the study period, while the control animals gained 5.6 + 3.1 g (p=0.019). The effects of the plant extract on body weight appeared to be independent of effects on food intake, as there was no detectable difference between food intake during the baseline and experimental periods.

Table 2: Effects of plant extract on blood glucose and plasma insulin

In summary, treatment with the plant extract for 11 days reduced blood glucose concentration in both lean, non-diabetic and obese, diabetic P. obesus. Three of the 5 obese, diabetic animals treated with the plant extract became non-diabetic within the 11 day study period, while in the other two animals the severity of hyperglycemia was substantially reduced. In addition, there was a tendency for reductions in plasma insulin concentrations in the plant extract treated animals, suggesting improvement in insulin action after treatment. The plant extract treated animals also exhibited reduced body weight gain during the study period relative to control animals, which may have contributed to their improved insulin sensitivity. Interestingly, this altered body weight gain was independent of food intake, which was not affected by the treatment (which also prevents the possibility of conditioned taste aversion). It is therefore assumed that the plant extract may have affected energy expenditure and/or substrate utilisation in P. obesus.

Overall these data suggest that the plant extract used in these studies may contain an insulin sensitising agent, and has potential therefore as an antidiabetic treatment. In Vitro Studies in Cultured 3T3-L1 Adipocytes

Glucose Uptake

3T3-L1 adipocytes were differentiated and cultured in DMEM containing 5% fetal calf serum using standard procedures. For the glucose uptake assay, the cells were washed with Krebs Ringer Buffer Solution for 2h, and then insulin stimulated (0-10OnM) for 30min before addition of the plant extract at concentrations of 0.005-0.5% (v/v) and ¹⁴C-2- deoxyglucose. After 15min the reaction was stopped with ice-cold PBS containing phloretin and SDS, and radioactivity counted immediately using a beta counter.

The effects of the plant extract at concentrations of 0.005, 0.05, 0.125, 0.25 and 0.5% were tested at various concentrations of insulin (0, 0.1, 0.5, 1, 5, 10 and 10OnM). The glucose uptake in plant extract treated cells was calculated relative to the corresponding dose of insulin without plant extract, and the results presented as percentages of the effects of insulin alone. Table 3 shows the results obtained at the various insulin concentrations. At each insulin concentration, the ability of insulin to stimulate glucose uptake was tested (data not shown) and results only accepted if insulin stimulation of glucose uptake was apparent.

The plant extract clearly stimulated glucose uptake in these cells at insulin concentrations of 0-5nM (Table 3). The rate of glucose uptake was increased by 3-fold at the highest plant extract concentration (0.5%) when no insulin was present (p<0.001), and by 2-fold in the presence of InM insulin (p=0.009). At insulin concentrations of 10 and 10OnM the plant extract had no discernible effect on glucose uptake in these cells, which was likely to be maximally stimulated at these concentrations of insulin. Table 3: Effects of plant extract on glucose uptake in 3T3-L1 cells (mean ± SEM, n=4-8)

^*p<0.05, p<0.01, ^#ρ<0.001 compared with 0 plant extract.

Lipogenesis

A similar protocol was used to measure incorporation of radioactive glucose into lipid in differentiated 3T3-L1 adipocytes, a measure of lipogenesis. For this assay, the cells were serum-starved overnight, then incubated in DPBS containing insulin (0-100OnM), plant extract at concentrations of 0.005-0.5% (v/v) and ^wC-2-deoxyglucose. After 3h the reaction was stopped with ice-cold PBS and the lipid fraction extracted by addition of isopropanol:heptane:H₂SO₄ (4:1 : ;0.1) for 20 min followed by centrifugation and removal of the upper phase, in which radioactivity was counted immediately using a beta counter.

The plant extract stimulated lipogenesis in these cells at insulin concentrations of 0-1OnM (Table 4). The rate of lipogenesis was increased by more than 2-fold at the highest plant extract concentration (0.5%) when no insulin was present (p<0.001), by 1.5-fold in the presence of InM insulin (p<0.001), and by 1.2-fold at an insulin concentration of 1OnM (p=0.036). At higher insulin concentrations the plant extract had no discernible effect on lipogenesis in these cells, which was likely to be maximally stimulated at these concentrations of insulin.

^*p<0.05, ^Λp<0.01, #p<0.001 compared to 0 plant extract.

To summarise the initial in vitro data, the plant extract stimulated lipogenesis and glucose uptake in differentiated 3T3-L1 adipocytes in the presence of 0-1OnM insulin. These data support the contention that the plant extract used in these studies may contain an insulin- sensitising agent, and suggest that this agent can directly affect glucose metabolism in adipocytes.

Example 2: Fractionation, Identification and In Vitro Effect of Strictosidinic Acid

Fractionation of plant extract.

The fteeze-dried plant extract (3g) was fractionated by HPLC/ESI-MS-ELSD under the conditions described in Table 5. Table 5: Conditions for preparative chromatographic fractionation.

Preparative chromatographic (HPLC) fractionation resulted in 42 fractions of the plant extract. Each was collected in a 4-ml brown glass vial, and the water and buffer were removed online via solid phase extraction. The mass of material in each fraction ranged from 0.23 to 126.32 mg, and the total combined weight of all 42 fractions was approximately 1.7 g (accounting for ~ 56% of the 3 g used).

Table 6: Weights of fractions collected after preparative chromatographic (HPLC) fractionation

Fraction Weight (mg) Fraction Weight (mg)

FOO 126.32 F21 0.83

FOl 0.57 F22 0.59

F02 0.74 F23 1.05

F03 1.58 F24 0.59

F04 2.09 F25 0.36

F05 5.21 F26 1.42

F06 7.33 F27 0.61

F07 2.24 F28 0.38

F08 1.01 F29 0.29

F09 1.1 F30 0.23

FlO 1.09 F31 0.26

FI l Ll F32 0.24

F12 0.7 F33 0.31

F13 1.26 F34 0.41

F14 1.62 F35 0.42

F15 1.31 F36 0.45

F16 1.12 F37 0.8

F17 1.28 F38 0.4

F18 1.45 F39 0.3

F19 0.94 F40 0.25

F20 0.83 F41 1.82

Screening of fractions 1) Lipogenesis bioassay (3T3-L1 adipocytes)

The plant extract fractions were examined in the lipogenesis (glucose incorporation into lipid) bioassay, measuring the amount of glucose converted into triglyceride using radioactive ^MC-glucose. Briefly, 3T3-L1 adipocytes were incubated in starvation media overnight followed by 30 minute incubation in DPBS buffer. Next, the DPBS buffer was removed, and treatment (freeze-dried plant extract, or its reconstituted fractions) was added to each well for 90 minutes in the presence of 1 nM insulin. Then, treatments were removed, and the cells were washed twice before being lysed with a lipid-extracting reagent. The lipid extract was then transferred to 2 ml-tube containing water and heptane. Thereafter, the mixture was vigorously vortexed, and ceiitrifuged to separate the lipid and water fractions. The lipid fraction (heptane soluble layer, upper phase) was added to 4 ml scintillation fluid to count the radioactivity using the β-counter Tri-carb^® 2900TR Liquid Scintillation Analyser (Packard; Packard Bioscience B.V., Groningen, Netherlands).

In each experiment, 2 plant extract fractions were tested at 0.05, 0.5, and 1% (w/v) in triplicate, together with insulin and the IMO 14 extract as positive control. To obtain uniformity and ease of comparison, results (amount of glucose incorporated into lipid) were converted to percent change relative to control (1 nM insulin).

Fraction 18 in particular had an effect on the incorporation of glucose into lipid in 3T3-L1 cells.

Effect of Fraction 18

1. Glucose Uptake in 3T3 -Ll adipocytes Cell culture

3T3-L1 fibroblasts are grown in 10cm dishes and cultured in DMEM (4.5 g/L glucose) supplemented with 10% (v/v) heat-inactivated fetal bovine serum (FBS), 50 IU/ml pencillin and 50 μg/ml streptomycin. Cells for differentiation into adipocytes were maintained at post-confluence for 2 days, and then induced to differentiate by the addition of DMEM containing 10% FBS, 2 μg/ml insulin, 0.25 μM dexamethasone and 0.5 niM 3-isobutyl-l- methylxanthine. After 3 days, the induction medium was replaced with fresh DMEM containing 10% FBS and 2 μg/ml insulin for another 3 days. Adipocytes were maintained in DMEM with 5% FBS for 2 days thereafter. Adipocytes were used 8-12 days after initiation of differentiation, after which time greater than 80% of fibroblasts had differentiated into mature adipocytes. Treatment with Fraction 18

3T3-L1 adipocytes were re-seeded into gelatin-coated clear- wall clear-bottom 96-well plates (2 x 10cm dishes : 1 x 96-well plate) and maintained in DMEM with 5% FBS. After 24 hrs, adipocytes were treated with 0.2%, 0.5%, 1% or 2% (w/v) Fraction 18 in DMEM (4.5 g/L glucose) containing 0.2% BSA for 16 to 18 hr (overnight). Each dose of SA was added to 4 wells and the following day, [³H] -2-deoxy glucose uptake was then measured (n=4). Fraction 18 stock solution was prepared as either 10% (w/v) (54 μg/ml) or 50% (w/v) (270 μg/ml) in milliQH₂0 and stored at 4°C.

Glucose Uptake

Cells were washed twice in Dulbecco's PBS, pH 7.4 (Gibco) containing 0.2% (w/v) RIA- grade BSA, 0.5 rnM MgCl₂ and 0.5 mM CaCl₂. Insulin at 0, 0.1, 1 and 10 nM is added for 30 min at 37°C. Uptake of 50 μM 2-deoxy glucose and 0.5 μCi 2-deoxy- [U-³H] glucose (NEN, PerkinElmer Life Sciences) per well was measured over the final 10 min of insulin stimulation (0, 0.5 and 1OnM). The reaction was stopped upon addition of ice-cold 80 μg/ml phloretin in PBS, pH 7.4 and cells solublised in 0.03% (w/v) SDS. Counts per minute (cpm) were measured by scintillator counter. Each measurement was performed in quadruplet in 1 assay, and each assay was repeated independently 3 times.

Following the screening, glucose uptake assays were utilised in 3T3-L1 adipocytes to confirm the activity of the selected fractions. As shown in Table 7 below, Fraction 18 caused a dose-dependent increase in glucose uptake in 3T3-L1 cells. At a dose of 0.5% (w/v) Fraction 18 caused an increase in glucose uptake of 17%. When the dose of Fraction 18 was increased to 1% (w/v), the increase in glucose uptake was 53%. These data strongly support the proposition that Fraction 18 increased the uptake of glucose into differentiated 3T3-L1 adipocytes. Table 7: Effects of Fraction 18 on glucose uptake in 3T3-L1 adipocytes.

Treatment Dose Glucose uptake % increase (pmol/min/well)

Insulin O nM 360 ± 44 0

I nM 1201 ± 131 234

1O nM 1673 ± 87 364

Fraction 18 0.5% (w/v) 420 ± 10 17

1.0% (w/v) 550 ± 99 53

Gene expression signature analysis

In addition to the bioassays above, gene expression signatures were used as an endpoint for secondary in vitro screening studies. Specifically, microarray technology was used to characterise the effects of the plant extract fractions on gene expression in 3T3-L1 adipocytes.

Hybidisation involved indirect labelling of target cDNA and reference cDNA with fluorescent dyes Cy 5 and Cy3 respectively (Amersham Pharmacia Biotech). Total RNA was extracted using the TRIzol method. The RNA was purified using RNeasy midi columns (Qiagen). 20μg of RNA per sample was processed using the Superscript Indirect cDNA labelling system (Invitrogen) as per manufacturer's instructions. The cDNA was hybridised in a 40μl reaction containing combined target and reference fluorescently labelled cDNAs, 5X SSC (Invitrogen), 16μg PolydA (Amersham), 5X Denhardt's solution (Invitrogen), 4.8μg yeast tRNA (SIGMA) and 0.01% Sodium dodecyl sulfate (Invitrogen). The reaction was heated to 98⁰C for 2 min and maintained at 6O⁰C until used. Microarrays were placed into a hybridisation chamber containing lOOμl of 2% SSC. A coverslip was placed on top of the microarray slide and the whole reaction mixture was loaded. Hybridisation chambers were secured and placed in a waterbath within a hybridisation oven at 42⁰C for 16 hr. Following hybridisation, the array slides were washed with gentle agitation to ensure all unbound cDNA was removed. The washing procedure included gentle agitation for 2 minutes in solution 1 (0.5X SSC, 0.1% SDS) followed by 2 minutes in solution 2 (0.5X SSC, 0.01% SDS) and finally 2 minutes in solution 3 (0.06X SSC). Slides were centrifuged at 1500rpm for 1 minute to dry.

Microarrays were scanned using GenePix 4000B (Axon Instruments) scanner and microarray data was analysed using GenePix Pro 5.1 and Acuity 4.0 software (Axon Instruments). The data were analysed using linear discriminant analysis to identify a subset of genes that characterised the effects of the plant extract fractions on gene expression profiles ("gene expression signature"). Furthermore, genes were grouped according to function and assessment of trends within biological pathways was conducted.

Pathway analysis revealed that Fraction 18 (cells treated with 0.5% w/v Fraction 18 for 24h) appeared to have significant effects on a number of genes associated with glucose metabolism, including those involved with adipogenesis (C/EBPβ increased by 40%, SREBP2 increased by 22%), insulin signalling (PKCΘ increased by 54%, PIK3R2 increased by 32%, GSK3A increased by 10%, SORBSl increased by 24%). Glycogen synthesis (GYS 1 increased by 20%), glycolysis (DLAT increased by 49%, HK2 increased by TA, PDK4 increased by 10%), glucose transport (OGT increased by 13%) and mitochondrial activity (MFN2 increased by 20%). Taken together, these data provide significant support to the hypothesis that Fraction 18 had effects on processes that regulate glucose homeostasis.

Mass spectrometry analysis of Fraction 18

Low resolution LC-MS analysis of Fraction 18 was conducted. Fraction 18 appeared to contain a chemical entity with a m/z value of 517, and this was by far the most abundant entity observed following electrospray ionisation.

The fraction was then analysed by high resolution mass spectrometry and gave an ion + believed to be a protonated species [M+H ] with a mass of 517.2179090. This mass was compared with a library of possible combinations of atoms that would match this exact molecular weight. This identified a combination of 33H, 26C, 2N and 90 with a mass of

517.2180571 which differed from the fraction 18 species by only 1.921 x 10 mass units.

1 13

H and C NMR analysis of Fraction 18

The solid material was dissolved in D3OD and gave the following H and 13, C NMR data.

C₂₆H₃₂N₂O₉ Exact Mass: 516.2108 MoI. Wt: 516.5403 C, 60.46; H, 6.24; N, 5.42; 0, 27.1

Identification of strictosidinic acid

Partial linkage analysis revealed the presence of aromatic protons and a six membered aromatic carbon ring. Using proprietary computer software, a search of known structures was conducted for those that comprised a mass of -516, an empirical formula of 27C, 34H, 2N and 90 and which contain an aromatic ring. This identified strictosidinic acid (SA), a monoterpenoid indole alkaloid (empirical formula C27H34N2O9; CA Index Name 2H- Pyran-5-carboxylic acid, 3-ethenyl-2-(β-D-glucopyranosyloxy)-3,4-dihydro-4-[[(lS)- 2,3,4,9-tetrahydro-lH-pyrido[3,4-b]indol-l-yl]methyl]-, methyl ester, (2S,3R,4S)-(9C1)). It is the acid form of strictosidine, a glycosylated alkaloid intermediate that is a common precursor for some 2200 alkaloids. The ¹H and ¹³C nmr data correlated well with that reported (Arbain, D. et al, 1993). Analysis of the MS and NMR data indicated that strictosidinic acid was the only chemical entity present in Fraction 18.

Effects of strictosidinic acid in bioassays

1. Glucose Uptake in 3T3-L1 adipocytes

Cell culture

3T3-L1 fibroblasts are grown in 10cm dishes and cultured in DMEM (4.5 g/L glucose) supplemented with 10% (v/v) heat-inactivated fetal bovine serum (FBS), 50 IU/ml pencillin and 50 μg/ml streptomycin. Cells for differentiation into adipocytes were maintained at post-confluence for 2 days, and then induced to differentiate by the addition of DMEM containing 10% FBS, 2 μg/ml insulin, 0.25 μM dexamethasone and 0.5 mM 3-isobutyl-l- methylxanthine. After 3 days, the induction medium was replaced with fresh DMEM containing 10% FBS and 2 μg/ml insulin for another 3 days. Adipocytes were maintained in DMEM with 5% FBS for 2 days thereafter. Adipocytes were used 8-12 days after initiation of differentiation, after which time greater than 80% of fibroblasts had differentiated into mature adipocytes.

Strictosidinic Acid Treatments 3T3-L1 adipocytes were re-seeded into gelatin-coated clear-wall clear-bottom 96-well plates (2 x 10cm dishes : 1 x 96-well plate) and maintained in DMEM with 5% FBS. After 24 hrs, adipocytes were treated with 0.2%, 0.5%, 1% or 2% (w/v) strictosidinic acid (SA) in DMEM (4.5 g/L glucose) containing 0.2% BSA for 16 to 18 hr (overnight). Each dose of SA was added to 4 wells and the following day, [³H] -2-deoxy glucose uptake was then measured (n=4). Strictosidinic acid stock solution was prepared as either 10% (w/v) (54 μg/ml) or 50% (w/v) (270 μg/ml) in milliQH₂0 and stored at 4⁰C.

Glucose Uptake

Cells were washed twice in Dulbecco's PBS, pH 7.4 (Gibco) containing 0.2% (w/v) RIA- grade BSA, 0.5 mM MgCl₂ and 0.5 mM CaCl₂. Insulin at 0, 0.1, 1 and 10 nM is added for

30 min at 37°C. Uptake of 50 μM 2-deoxy glucose and 0.5 μCi 2-deoxy- [U-³H] glucose (NEN, PerkinElmer Life Sciences) per well was measured over the final 10 min of insulin stimulation (0, 0.5 and 1OnM). The reaction was stopped upon addition of ice-cold 80 μg/ml phloretin in PBS, pH 7.4 and cells solublised in 0.03% (w/v) SDS. Counts per minute (cpm) were measured by scintillator counter. Each measurement was performed in quadruplet in 1 assay, and each assay was repeated independently 3 times.

Results

Administration of strictosidinic acid caused a statistically significant, dose-dependent increase in glucose uptake by 3T3-L1 adipocytes. As shown in the table below, at doses of 0.5% and 1% (w/v) strictosidinic acid increased glucose uptake by up to 26%.

These results demonstrate that strictosidinic acid increases glucose uptake in 3T3-L1 adipocytes. Example 3: In Vivo Effects of Strictosidinic Acid

Compound Preparation and Administration

Dosing solutions of the test article, strictosidinic acid, were prepared fresh on each dosing day and stored at room temperature, protected from light. The compound was formulated in sterile NMP:PEG300:Saline (1:2:17, v/v). Stock solutions were prepared by dissolving the compound in NMP:PEG300 (1:2, v/v). Saline was added to the stock solution prior to administration of the compound. The solutions were mixed by vortexing immediately prior to dosing to ensure a homogeneous suspension of the compound. "Vehicle" corresponds to a sterile solution of NMP:PEG300:Saline (1:2:17, v/v).

Acute Tolerated Dose of Strictosidinic Acid in Balh/c mice

The compound was administered once only by oral gavage on Day 0 of the study. Administration was performed at a dosing volume of 10 mL/kg, at dosing concentrations of 31, 62.5, 125, and 250 mg/kg.

Each animal's body weight was measured immediately prior to dosing each day. The actual volume administered to each mouse was calculated and adjusted based on the body weight.

Starting with the lowest concentration (31 mg/kg, Group 1), one mouse was treated, followed by a 2 hour observation period. As no adverse effects were observed during this time, the remaining mice in the group were treated. Twenty four hours after the first concentration was administered, one mouse from the next dose concentration (62.5 mg/kg, Group 2) was treated. As no adverse effects were observed during a 2 hour observation period, the remaining mice in the group were treated. The process was continued until all dose concentrations were administered (125 mg/kg, Group 3 and 250 mg/kg, Group 4).

Results The acute tolerated dose (ATD) is defined as the highest dose (administered as a single dose) that does not induce a weight loss in an individual animal in excess of 15% or cause death or severe morbidity, at any time during the study period.

Sixteen female Balb/c mice were randomised into 4 treatment groups, each with 4 mice. Each group had similar mean starting body weight. The groups were allocated a different treatment regimen of strictosidinic acid, administered once only, by oral gavage. The compound was administered at a volume of 10 mL/kg, at dosing concentrations of 31, 62.5, 125 and 250 mg/kg.

Body weight measurements were made daily from Day 0 of the study and were continued until 7 days after treatment was finished, and are detailed in Table A.

None of the mice lost body weight in excess of the defined parameters of the study, and no adverse effects were observed in the mice during the study period. The ATD for strictosidinic acid administered by oral gavage was not achieved in this study, and is therefore determined to be >250 mg/kg (the highest dose concentration tested in the study).

Table A: Body weight data for ATD Study: strictosidinic acid in Balb/c mice

Effect of Strictosidinic acid on fasting blood glucose levels in diet-induced obese mice (Mus Musculus)

Animals Male C57B1/6J mice were obtained at 8 weeks of age from Animal Resources Centre (Canning Vale, Western Australia). Animals were individually housed and allowed free movement and ad libitum access to water and food. Animals were maintained on a 12 hour light (6 am-6 pm) and 12 hour dark (6 pm-6 am) cycle. After two weeks acclimatizing, mice were fed a high fat rich diet (Research Diets, Inc. [New Jersey, USA] with a total energy density of 4.7 kcal/g. Caloric distribution in the diet was 20% from protein, 35 % from carbohydrates, and 45 % from fat) for 11 weeks.

Treatment

During the last week, animals in groups of 6 were treated with doses selected from: vehicle, 25mg/kg, 50mg/kg, 100mg/kg and 175mg/kg of Strictosidinic acid in

NMP:PEG300:Saline (l:2:17, v/v) by oral gavage (5 μl per gram of body weight) using feeding tubes 18ga (1.2mm)*38mm from Instech Solomon (Plymouth, USA), while control animals were administered with vehicle only. It became clear early in the study that SA was not fully soluble at the concentration required for the 175 mg/kg dose, so this group was discontinued and not included in the analysis.

Fasting Blood Glucose Levels

Animals were fasted overnight and blood samples collected from the tail at 10am on days 0 and 9 of the study. The results are presented in Table B. Table B: Effect of Strictosidinic acid treatment on fasting blood glucose concentration in diet-induced obese mice

Data represent the mean ± SE of 4-6 animals per group.

Intraperitoneal Glucose Tolerance Test: Blood glucose concentrations during ipGTT

Animals were subjected to an intraperitoneal glucose tolerance test (ipGTT) prior to commencement of dosing, and on day 7 after final dosing. Animals were fasted overnight and then administered glucose (50%w/v in saline: Pharmalab) by intraperitoneal injection at a dose of 2g/kg. Blood glucose levels were monitored at t = Omin, 15min, 30min, 60min and 90min after the glucose dose using commercially available glucometers (Accuchek Advantage, Roche) based on the electric current generated when a blood drop is spotted on a test strip resulting in conversion of the glucose present in the sample to gluconolactone by the glucose dehydrogenase enzyme, in the presence of the coenzyme (PQQ). The results are depicted in Table C.

Table C: Effect of Strictosidinic acid treatment on area under plasma glucose curve during ipGTT in diet-induced obese mice

Data represent the mean ± SE of 5-6 animals per group. Intraperitoneal Glucose Tolerance Test: Plasma insulin concentrations during ipGTT

Plasma insulin concentrations were determined using a commercially-available insulin ELISA kit (Crystal Chem) which was used according to the manufacturer's instructions. The results are presented in Table D.

Table D: Effect of Strictosidinic acid treatment on plasma insulin concentration during ipGTT in diet-induced obese mice

Data represent the mean ± SE of 5-6 animals per group. *unpaired ttest value vs vehicle at specified timepoint.

Determination of Epididymal Fat as a Percentage of Body Weight

Animals were euthenized by cervical dislocation, and their weights were independently recorded. The animals were then dissected and their livers and epididymal fat pads were surgically excised, and their individual weights recorded. The results are presented in Table E. Table E: Effect of Strictosidinic acid treatment epididymal fat mass as % of body weight in diet-induced obese mice

Data represent the mean ± SE of 5-6 animals per group.

BIBLIOGRAPHY

Arbain, D. et al., Amt. J. Chem. 1993, 46: 977-985. Galan, M. C. et al. , Tetrahedron Lett. , 2006, 1563- 1565. Geerlings et al., J. Biol Chem., 2000, 275: 3051-3056.

Govers et al, MCB, 2004, 29: 6456-6466.

Kitajima, M. et al, Chem. Pharm. Bull. 2002, 50(10): 1376-1378.

Larock, R.E., Comprehensive Organic Transformations, VCH Publishers, 1989. The Merck Index, Thirteenth Edition, ONR-82, 310. Morita et al, Gene Therapy, 2000, 7: 1063-1066.

Patthy-Lukats, A. et al, J. Nat. Prod. 1999, 62: 1492-1499.

QvLon et aL, PNAS, 1994, 91: 5587-5591.

Stδckigt, J. et al, J. Chem. Soc. Chem. Commun. 1977, 646-648. Treimer, J. F. et al, Eur. J. Biochem. 1979, 81 : 225-233. Yamazaki et al, Phytochemistry, 2003, 62: 461-470.

United States Patent No. 6,720,331

United States Patent No. 6,350,757.

Claims

THE CLAIMS DEFINING THE INVENTION ARE AS FOLLOWS:

1. A method of increasing glucose uptake by a cell comprising contacting said cell with a compound of Formula (I):

wherein,

is a single or a double bond;

Ri-R₄ are independently selected from hydrogen, hydroxy, thiol, halo, CO₂H, carboxy ester, amino, nitro, cyano, amido, sulfoxide, sulfonamide, sulfonyl, sulfate, sulfonate, phosphate, phosphonate, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted acyl, optionally substituted alkoxy, optionally substituted alkenyloxy, optionally substituted alkynyloxy, optionally substituted aryloxy, optionally substituted heteroaryloxy, optionally substituted carbocyclyloxy, optionally substituted heterocyclyloxy, optionally substituted aralkyloxy, optionally substituted acyloxy, optionally substituted alkylthio, optionally substituted alkenylthio, optionally substituted alkynylthio, optionally substituted arylthio, optionally substituted carbocyclylthio, optionally substituted aralkylthio, optionally substituted heteroarylthio, optionally substituted heterocyclylthio, optionally substituted acylthio or any 2 adjacent R₁-R₄ together form a -0-(CHa)_n-O- or -NR-(CH₂)_n-NR-group wherein n is 1 or 2 and each R is independently H or Ci-₆alkyl;

Re is absent when is a double bond;

R₅ and R₆ (when present) independently are selected from hydrogen, optionally substituted alkyl, optionally substituted acyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aralkyl, optionally substituted heterocyclyl, optionally substituted carbocyclyl, and amido;

R₇ is selected from hydrogen, optionally substituted alkyl, CO₂H, carboxy ester, amido, optionally substituted heteroaryl and optionally substituted aryl;

Rs is selected from hydrogen, optionally substituted alkyl, optionally substituted aryl, optionally substituted heteroaryl, CO₂H, carboxy ester and SO₃H;

R_7a and R_8a are independently selected from hydrogen, optionally substituted alkyl and optionally substituted aryl.

COX forms a group selected from optionally substituted acyl, CO₂H, carboxy ester and amido,

Y is selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclyl, optionally substituted carbocyclyl, optionally substituted acyl and a saccharide group, Z is selected from optionally substituted at least C₂alkyl, optionally substituted alkenyl and optionally substituted alkynyl,

or a pharmaceutically acceptable salt or prodrug thereof.

2. A method for regulating glucose homeostasis in a subject in need thereof comprising the administration of a compound of Formula (I) as defined in claim 1 or a pharmaceutically acceptable salt or prodrug thereof.

3. A method of treating insulin resistance in a subject in need thereof comprising the administration of a compound of Formula (I) as defined in claim 1 or a pharmaceutically acceptable salt or prodrug thereof.

4. A method of lowering blood glucose levels in a subject in need thereof comprising the administration of a compound of Formula (I) as defined in claim 1 or a pharmaceutically acceptable salt or prodrug thereof.

5. A method for treating a disease or condition, or symptom thereof, in which insulin resistance is involved comprising the administration of a compound of Formula (I) as. defined in claim 1 or a pharmaceutically acceptable salt or prodrug thereof to subject in need thereof.

6. A method according to claim 5 wherein the disease or condition is NIDDM, gestational diabetes, impaired glucose tolerance, impaired fasting glucose or syndrome X.

7. A method according to any one of claims 1 to 6, wherein:

R_1-4 are independently selected from hydrogen, alkyl, alkoxyalkyl, alkoxy, alkylthio, halo, trifluoromethyl, trichloromethyl, tribromomethyl, hydroxy, phenyl (which itself may be further substituted), benzyl (wherein benzyl itself may be further substituted), phenoxy (wherein phenyl itself may be further substituted), benzyloxy (wherein benzyl itself may be further substituted), amino, alkylamino, dialkylamino, acylamino, phenylamino (wherein phenyl itself may be further substituted), nitro, formyl, -C(O)-alkyl, O-C(O)-alkyl, benzoyl (wherein benzyl itself may be further substituted, benzoyloxy (wherein benzyl itself may be further substituted, COjH₅ CCtølkyl, Cθ₂phenyl, C0₂benzyl (wherein benzyl itself may be further substituted, CONH₂, CONHphenyl (wherein phenyl itself may be further substituted, CONHbenzyl (wherein benzyl itself may be further substituted, CONHalkyl, CONdialkyl, aminoalkyl, thioalkyl, carboxyalkyl, carboxyesteralkyl, amidoalkyl, formylalkyl, acylalkyl, nitroalkyl, sulfoxidealkyl, sulfonylalkyl, sulfonamidoalkyl, and where 2 adjacent carbon atoms are substituted by one end each of a -O-(CH₂)_n-O- or -NH-(CH₂)_n-NH- group, wherein n is 1 or 2;

R₅ and R₆ are selected from hydrogen, optionally substituted C_1-6alkyl, optionally substituted C(O)C_1-6alkyl, optionally substituted C(O)phenyl, and optionally substituted C(O)benzyl;

R₇ is selected from hydrogen, CO₂H, CO₂C_1-6alkyl, CONH₂, CONHC_1-6alkyl and CONHaryl;

Rg is hydrogen;

R_7a and R_8a are hydrogen;

X is selected from OH, OCi_-I0alkyl, NHC_1-loalkyl, NHphenyl and NHbenzyl;

Y is H or a saccharide;

Z is CH₂=CH₂ or C_2-10alkyl

8. A method according to any one of claims 1 to 6 wherein the compound is strictosidinic acid.

9. A method according to claim 8 wherein the compound is in substantially pure or isolated form.

10. A compound of Formula (I):

wherein,

is a single or a double bond;

R₁-R₄ are independently selected from hydrogen, hydroxy, thiol, halo, CO₂H, carboxy ester, amino, nitro, cyano, amido, sulfoxide, sulfonamide, sulfonyl, sulfate, sulfonate, phosphate, phosphonate, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted acyl, optionally substituted alkoxy, optionally substituted alkenyloxy, optionally substituted alkynyloxy, optionally substituted aryloxy, optionally substituted heteroaryloxy, optionally substituted carbocyclyloxy, optionally substituted heterocyclyloxy, optionally substituted aralkyloxy, optionally substituted acyloxy, optionally substituted alkylthio, optionally substituted alkenylthio, optionally substituted alkynylthio, optionally substituted arylthio, optionally substituted carbocyclylthio, optionally substituted aralkylthio, optionally substituted heteroarylthio, optionally substituted heterocyclylthio, optionally substituted acylthio or any 2 adjacent R₁-R₄ together form a -O-(CH₂)_n-O- or -NR-(CH₂)_n-NR-group wherein n is 1 or 2 and each R is independently H or C_1-6alkyl;

R₆ is absent when "■ is a double bond;

R₅ and R₆ (when present) independently are selected from hydrogen, optionally substituted alkyl, optionally substituted acyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aralkyl, optionally substituted heterocyclyl, optionally substituted carbocyclyl, and amido;

R₇ is selected from hydrogen, optionally substituted alkyl, CO₂H, carboxy ester, amido, optionally substituted heteroaryl and optionally substituted aryl;

R₈ is selected from hydrogen, optionally substituted alkyl, optionally substituted aryl, optionally substituted heteroaryl, CO₂H, carboxy ester and SO₃H;

R_7a and R_8a are independently selected from hydrogen, optionally substituted alkyl and optionally substituted aryl.

COX forms a group selected from optionally substituted acyl, CO₂H, carboxy ester and amido,

Y is selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclyl, optionally substituted carbocyclyl, optionally substituted acyl and a saccharide group, Z is selected from optionally substituted at least C₂alkyl, optionally substituted alkenyl and optionally substituted alkynyl,

or a pharmaceutically acceptable salt or ester thereof, provided that the compound is not (3R) or (3S) strictosidine, strictosidine ethyl ester or allyl ester, strictosidinic acid, 5- carboxystrictosidine, 3,4-dehydro-5-carboxystrictosidine, 3,4-dehydro5- carbomethoxystrictosidine or gluco-acetylated forms thereof.

11. A composition comprising a compound of Formula (I), as defined in claim 1, or a pharmaceutically acceptable salt or prodrug thereof, together with a pharmaceutically acceptable excipient.