JP2002529409A - Glucose and lipid lowering compounds - Google Patents

Abstract

  (57) [Summary] Aims and methods for lowering lipid or glucose, especially lipid lowering or glucose lowering compounds having an AB ring structure of formula (I) are provided. In the formula (I), the dotted lines between carbons 5 and 6, carbons 4 and R2, and carbons 11 and R2 may each have a double bond at the carbon bonded from the numeral carbon. Where X is hydroxy or oxy which, together with Y, is bonded to carbons 8 and 12.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

[Industrial applications]

The present invention relates to methods for reducing hyperlipidemia and hyperglycemia, and certain compounds and compositions. This application claims priority to US patent application Ser. No. 09 / 190,507, filed Nov. 12, 1998 and US Ser. No. 09 / 197,352, filed Nov. 20, 1998.

[0002] The worse complications of hyperlipidemia are widespread throughout the world and are manifested by a high incidence of serious atherosclerotic diseases, including heart attacks and strokes. Cardiovascular and cerebrovascular diseases are often slowly evolving symptoms that are not detected in the early stages of pathophysiology, the symptoms typically exhibit high circulating lipid levels and can last for years. Demonstrated by deposits of fatty substances accumulating in large vessels over decades. Only when there is enough blood vessel damage and accumulated thrombotic material to occlude the major blood vessels, or only when they dislodge and cause vessel closure
The disease causes its high morbidity or mortality.

[0003] Causal factors that are within and beyond the control of an individual contribute to the disease process, among other factors are genetic features, diabetes mellitus, etc. Or lifestyle factors, including diet, smoking, and physical activity. Therapeutic interference has only recently emerged as a direct target for genetic factors, and lifestyle changes are difficult and often difficult to achieve. Therapies directed at pharmacologically lowering circulating lipid levels have been clinically successful in reducing the incidence of cardiovascular and cerebral vascular disease. Markers indicative of high circulating lipids and their pathological condition include free fatty acids, glycerol, triglycerides, cholesterol, and low density lipoprotein. Reduction or suppression of each of these components can have a positive effect on other abnormal circulating analytes, including glucose and insulin.

[0004] Diabetes mellitus is a disease of a number of factors, including metabolic diseases associated with insulin-producing tissues as well as insulin-regulated tissues. In addition to being a disease of hyperglycemia, certain forms of the disease are associated with abnormally high circulating lipid levels. Thus, lowering circulating levels of lipids and glucose is beneficial in treating patients with diabetes mellitus. In type II diabetes, genetic mutations can produce a small group of certain diseases (eg, diabetes or MODY that begins with young adult maturity), but the most common forms of the disease are: It involves resistance to insulin action on insulin target tissues and is associated with obesity, primarily obesity of abdominal origin. In addition to high glucose levels, circulating levels of triglycerides, cholesterol, and non-esterified fatty acids (NEFA) increase during the fasting period. People with diabetes tend to develop cardiovascular disease, which is a major cause of the morbidity and mortality of the disease. Resistance to the action of insulin occurs not only in hyperglycemic conditions but also in Syndrome X, an insulin resistant disease characterized by hyperinsulinemia and dyslipidemia (Reaven GM, Syndrome X). , Clinical Diabetes, March / April 19
94: 32-36). In this condition, normoglycemia (nor
moglycemia). However, when insulin resistance cannot be maintained due to the compensatory activity of the pancreas, there is a concomitant lack of glucose tolerance. Lipid abnormalities are therefore believed to contribute to the severity of diabetic complications, such as cardiovascular complications, when combined with either excessive hyperglycemia or euglycemic syndrome X phenotype (Olewski, JM). , Current approaches to managing type 2 diabetes: A practical monograph, National Diabetes Education I
nitiative, 1997).

[0005] Oral antihyperglycemic agents such as sulfonylureas and biguanizides, as in the case of sulfonylureas, by promoting the endogenous release of insulin from β-cells of the pancreas, or in the case of biguanides. Promotes the improvement of high lipid levels by overcoming the associated insulin resistance state by enhancing glucose processing and reducing gluconeogenesis, as in Other methods of controlling high lipids and glucose have involved the use of widely acting oral antihyperglycemic agents, such as the thiazolidinedione class of drugs, which lower fatty acids and improve the insulin resistance state of insulin responsive tissues. Compound 4,7,8αH-eudesma-5 (6), 11 (13) -diene-8,12-olide (4,7,8αH-eudesma-5 (6), 11 (13) -di
en-8,12-olide) can induce hyperglycemia in rabbits at high doses to induce hypoglycemia at moderate doses and to suppress food-induced hyperglycemia. It has been reported (1986, Handbook of Effec
tive Ingradients of Medicinal Plants), Beijing, China).

[0006] A number of different therapeutic strategies have been developed to treat hyperlipidemia and related hyperglycemia in type 2 diabetes. The purpose of lowering circulating lipids was to reduce cardiovascular morbidity and / or improve overall diabetes status. Examples of drugs that directly affect the triglyceride and cholesterol content of plasma include H
Includes MG-CoA reductase inhibitors, fibric acids, and bile salt resins. These classes are effective in lowering triglyceride and cholesterol content, but have little effect on plasma fatty acids.

[0007] Among the lipids, non-esterified fatty acids appear to play a role in promoting a diabetic and / or hyperlipidemic, insulin resistant state. High free fatty acids result from either or both excessive body loading of lipid tissue in obese conditions or uncontrolled degradation of triglycerides in adipose tissue, the primary insulin target tissue. Furthermore, high levels of free fatty acids have been shown to actually induce insulin resistance in muscle, the main tissue that uses body glucose, where direct effects on glucose transport in muscle were observed. Have been. In addition, fatty acids affect hepatic metabolism and increase hepatic glucose output. Furthermore,
High fatty acids induce impaired α-cell function by reducing insulin secretion from α-cells in response to glucose stimulation. Attempts to reduce fatty acid levels have involved the development of pyrogenic drugs (3 agonists) that function by stimulating brown adipose tissue to oxidize fatty acids, and are therefore believed to clean them from the circulatory system. . Alternatively, removing fatty acids from the effects of the liver can be achieved by using the enzyme creatinine palmitoyltransferase I (C
Attempts have been made to develop inhibitors of fatty acid oxidation by PTI).

[0008] Resistance to the action of insulin, particularly in adipose tissue, appears to play a significant role in fatty acid gain. Drugs that inhibit the breakdown of adipose tissue of triglycerides
It is known as a lipolytic inhibitor. The present invention relates to reducing circulating lipids to treat diseases associated with hyperlipidemia, or hyperglycemia,
It is concerned with developing new methods and drugs to lower circulating glucose to treat diabetes and related diseases.

[0009]

Summary of the Invention

The present invention is directed to lipid-lowering or glucose-lowering methods and compounds, in particular A of the formula I
Lipid lowering or glucose lowering compounds having a B ring structure.

Embedded image Wherein, between up carbon dotted between carbons 5 and 6, the dotted line between carbons 4 and R ^2, and the dotted line between carbons 11 and R ^3, which are attached from the carbon of independently to each of the numbers Represents that a double bond may be present.

The present invention provides, among others, the following: Methods for lowering lipid levels in mammals to treat or prevent the development of pathologies associated with high lipid levels, compounds that inhibit enzymatic hormone-sensitive lipases and inhibit lipolysis, insulin resistance and syndrome Drugs and methods for treating X, methods for lowering glucose levels in mammals to treat or prevent the development of pathologies associated with hyperglycemia, eg, delaying or preventing the onset of non-insulin dependent diabetes mellitus Therefore, a method of treating impaired glucose tolerance that is associated with fasting or dietary hyperglycemia, euglycemia, or insulin resistance.

[0011]

[Detailed description]

In accordance with the present invention, there is provided a method for reducing the glucose level of a mammal (particularly a hyperglycemic mammal) or reducing the lipid level of a mammal, comprising administering to the animal an effective amount of a composition comprising a compound of the above formula. A method is provided.

The present invention provides for administering to a mammal an effective lipid-lowering or glucose-lowering amount of one or more compounds having an AB ring structure, ie, compounds of Formula I, or a pharmaceutically acceptable salt thereof. becomes, decreases blood lipid levels of a mammal, or a method for lowering blood glucose, in the above formula I, the dotted line between carbons 5 and 6, the dotted line between carbons 4 and R ^2, And the dashed line between carbon 11 and R ³ indicates that a double bond may exist independently from each numbered carbon to the carbon to which it is attached, where R ¹ is A hydrogen, hydroxy, lower alkanoyloxy or alloyoxy group, wherein the aryloxy is one or more lower alkyl, lower alkoxy, halo, trifluoromethyl, di (lower) alkylamido; , Hydroxy, nitro, or C1-C3 alkylenedioxy groups), wherein X is hydroxy or, together with the Y group, bonded to carbon 8 and carbon 12 to form oxy Wherein Y is hydroxy or an amino group which may be mono- or di-substituted with lower alkyl, wherein said lower alkyl may together form a 5- or 6-membered ring, Wherein R ² and R ³ are each independently a lower alkyl or alkenyl group, wherein the lower alkyl of R ³ can be mono- or di-substituted with a lower alkyl at the carbon bonded to C11. Can be substituted with an amino group, wherein the lower alkyl can also form a 5- or 6-membered ring, provided that when R ¹ is hydrogen, (a) X is hydroxy or (b ) R ³ is either substituted with amino. The invention further relates to pharmaceutical compositions to be administered.

The present invention also relates to compounds of formula I (and pharmaceutical compositions thereof) or pharmaceutically acceptable salts thereof, wherein the dashed line between carbons 5 and 6, the dashed line between carbons 4 and R ² , and The dashed line between carbon 11 and R ³ indicates that a double bond may exist independently from each numbered carbon to the carbon to which it is attached, and R ¹ is hydrogen, hydroxy, lower An alkanoyloxy or an aryloxy group, wherein the aryloxy is substituted by one or more lower alkyl, lower alkoxy, halo, trifluoromethyl, di (lower) alkylamino, hydroxy, nitro, or C1-C3 alkylenedioxy groups. Wherein X is hydroxy or oxy bonded to carbon 8 and carbon 12 together with the Y group, wherein Y is hydroxy Or an amino group which can be mono- or di-substituted with lower alkyl, wherein the lower alkyl can also form a 5- or 6-membered ring together, wherein R ² and R ³ are Each is independently a lower alkyl or alkenyl group, wherein the lower alkyl of R ³ can be substituted with an amino group which can be mono- or di-substituted with lower alkyl, wherein the lower alkyl is 5 members or 6
(1) when X is hydroxy, R ¹ is not hydrogen; and (2) when R ¹ is hydroxy, X is (a) whether X is hydroxy. (B) R ³ is substituted with amino; (3) R ² is a methylene linked to C 4 by a double bond, methylene linked to C 11 by a R ³ double bond, and X is oxy The condition is that R ¹ is not ethanolyloxy. Preferably R ¹ is not propanoyloxy.

In one embodiment, the hydrogen substituent at positions 5, 7, and 8 is α to the AB ring structure. In another embodiment, X is hydroxy. In another embodiment,
R ¹ is selected from lower alkanoyloxy groups such as acetoxy, propanoyloxy, isobutyroxy and cyclopropanoyloxy. The compound is, for example, 2α-acetoxy-5,7,8αH-eudesmer 4 (15), 11 (13) diene-8,12-olide; 2α-propanoyloxy5,7,8αH-eudesma-4 (15), 11 (13 ) Diene-8,12-oride; 2α-cyclopropanoyloxy 5,7,8αH-eudesmer 4 (15), 11 (13) -diene-8,12-oride; 2
α-acetoxy 4,5,7,8,11αH-eudesmer 8,12-oride; selected from the group consisting of 2α-cyclopropanoyloxy 5,7,8,11αH-eudesm-4 (15) -ene-8,12-oride Is done. In another embodiment, R ¹ is an alloyoxy, such as floyloxy and benzyloxy, such as 2α-benzoyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,1.
2-Olide; 2α-Flouroxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-
Oride; 2α-benzoyloxy 4,5,7,8,11αH-eudesman-8,12-olide; or 2
α-Benzyroxy-5,7,8,11αH-eudesm-4 (15) -ene-8,12-olide.

The compound can be administered in a glucose lowering effective amount or a lipid lowering effective amount. In another embodiment, (1) X is oxy, (2) R ¹ is lower alkanoyloxy or alloyoxy, provided that R ² is a methylene bonded to C4 by a double bond, and R ³ is a double bond. R ¹ is not ethanoyl (ethanoyloxy) when is methylene linked to C 11 at, and (3) R ² and R ³ are each independently a lower alkyl group or unsaturation at the bond to carbon 4 or carbon 11 It is a limited lower alkenyl group. In another embodiment, (1) X is oxy, (2) R ¹ is lower alkanoyloxy or alloyoxy, and (3) R ² and R ³ are each independently a lower alkyl group or unsaturated carbon 4 or A lower alkenyl group limited to the bond to carbon 11;

[0016] Lower alkyl and alkenyl groups contain 1 to 6 carbon atoms. A lower alkenyl group is defined in connection with the backbone structure of Formula 1 such that the methylene group has unsaturation, for example, in connection with carbon 4 or carbon 11. In one embodiment, at least one or both of R ¹ and R ² is methyl or methylene (both are bonded to carbon 4 or carbon 11 by a double bond). The lower alkanoyloxy groups mentioned above contain 1 or 2 to 6 carbon atoms and include methanoyloxy, ethanoyloxy or acetoxy, propanoyloxy, butyryloxy, pentanoyloxy, hexanoyloxy and the corresponding branched isomers thereof. Also included herein in the alkanoyloxy groups are substituted and unsubstituted cycloalkyl groups such as cyclopropanoyloxy, cyclobutanoyloxy, cyclopentanoyloxy, cyclohexanoyloxy, 1-methylcyclohexanoyloxy, And trans-4-isopropylcyclohexanoyloxy. The aryloxy groups mentioned herein include substituted and unsubstituted aromatic and heterocyclic groups such as furanoyloxy, benzoyloxy, 4-fluorobenzoyloxy, 2-chlorobenzoyloxy, 2-methoxybenzoyloxy, 3-ethyl. Benzoyloxy, 4-ethylbenzoyloxy, 4-isopropylbenzoyloxy, 4-chlorobenzoyloxy, 3,5-di-t-butyl-4-hydroxybenzoyloxy, 2-methoxy-5-chlorobenzoyloxy, 3-fluoro-4'-methoxybenzoyloxy, 2-trifluoromethylbenzoyloxy, 4-trifluoromethylbenzoyloxy, 2-naphthoyloxy, 2-thiophenecarboniloxy, 3-pyridinecarbonyloxy, and 5-methyl-2 ' -Pyrazinecarbonyloxy and the like. It includes both 2α and 2β isomers, with 2α being preferred. Lower alkyl groups include methyl, ethyl, propyl, butyl, pentyl, hexyl and the corresponding branched isomers thereof. Preferably the aryl group is C6
A heteroaromatic group having a C10 aromatic group or 5 to 10 ring elements, of which up to two are preferably heteroatoms selected from nitrogen, oxygen or sulfur.

The lower alkyl groups mentioned above are methyl, ethyl, propyl, butyl, pentyl,
It includes hexyl and the corresponding branched isomers. As noted above, the linking groups R ² and R ³ can be single or double bonds, and the alkyl groups described herein can be single or double bonded to carbons 4 and 11, respectively.

Non-limiting examples of compounds where R ¹ is hydrogen include salts such as the sodium salts of the following: 8β-hydroxy-4,7,8αH-eudesma-5,11 (13) -diene-12-oic acid; 8β-hydroxy-5,7,8,11αH-eudesm-4 (15) -en-12-oic acid ; 8β-hydroxy-4,5,7,8,11αH-eudesman-12-oic acid; 2α-
Hydroxy-4 (15), 11 (13) -eudesmadiene-12,8-olide. Non-limiting examples of compounds where R ¹ is hydroxy include salts such as the sodium or hydrochloride salts of: 2α, 8β-dihydroxy-2β, 4,5,7,8,11αH-eudesmann-12-oic acid; 2α, 8β-dihydroxy-4 (15), 11 (13) -eudesmadiene-12-oic Acid; 2α-benzoyl-8β-hydroxy-2β, 4,5,7,8,11αH-eudesmann-12-oic acid; 1- [2α-hydroxy-11,12 dihydro-5,7,8αH-eudesm-4 (15
) -Ene-8,12-oridyl] -pyrrolidine; and 1- [2α-benzoyloxy-11,12-dihydro-5,7,8αH-eudesma-4 (15) -ene-8,12-oridyl] -pyrrolidine.

Compounds in which R ¹ is a lower alkanoyloxy group include, but are not limited to, 2α-acetoxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; -
Propanoyloxy 5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; 2α
-Isobutylyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; α
-Cyclopropanoyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; 2α-acetoxy-4,5,7,8,11αH-eudesmann-8,12- And 2α-cyclopropanoyloxy 5,7,8,11α H-eudesm-4 (15) -ene-8,12-olide. Compounds in which R ¹ is an aryloxy group include, for example, 2α-benzoyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; 2α-fluoroyloxy 5,7,8αH -Eudesma-4 (15), 11 (13) -diene-8,12-orito ゛; 2α-benzoyloxy 4,5,7,8,11αH
-Eudesman-8,12-olide; and 2α-benzoyloxy-5,7,8,11αH-eudesm-
Includes 4 (15) -ene-8,12-olide.

The compounds of the invention exert a positive effect on circulating lipid levels when administered to hyperlipidemic mammals. Furthermore, in vivo, this compound specifically inhibits the hormone-sensitive lipase (HSL) enzyme and inhibits the following lipolysis in a whole cell model. In vivo, the compounds described herein suppress the appearance of an increase in circulating free fatty acids in normal mice fasted overnight. Obesity / type
This compound reduces elevated circulating lipid levels in a genetic model of type II diabetes-related hyperlipidemia.

The compounds of the present invention are made from synthetic starting materials or those isolated from natural sources. 2α-hydroxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene, a particular starting material having a double bond between carbons 4 and 15 and between carbons 11 and 13 -8,12-Olide can be synthesized by the procedure described by Tomioka et al. (1984, Tetrahedron Letters 25: 333-336) or described by Hearts et al. (1962, J. Org.Chem. 27: 905-910). And isolated and purified from plants, Iva microcephala or other plants. Alkanoyloxy and alloyoxy derivatives of 2α-hydroxy compounds, such as 2α-hydroxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide, can be made by synthetic procedures known to those skilled in the art. Can be, for example, 2
α-propanoyloxy 5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide and 2α-benzyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -Diene-8,12-olide is derived from 2α-hydroxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide by propanecarbonyl chloride and benzoyl chloride, respectively. It is made by the reaction with To make compounds of the invention having a single bond between R ² and R ³ and each ring carbon, a diene such as 2α-hydroxy-5,7,8αH-eudesma-4 (15), 1
1 (13) -Diene-8,12-olide is first converted to, for example, Hertz et al. (1962, J. Org.Chem. 27: 905-910)
Reduction to 2α-hydroxy-4,5,7,8,11αH-eudesmann-8,12-olide and then reaction with, for example, an acyl chloride or benzoyl chloride to give 2α-
Acetoxy 4,5,7,8,11αH-eudesmann-8,12-olide and 2α-benzoyloxy
Produces 4,5,7,8,11αH-eudesmann-8,12-olide. Compounds in which only one of R ² or R ³ is double bonded to the ring may be isolated or prepared synthetically or from natural sources, followed by analogous synthetic procedures as described above. For example, compound 5,7,8αH-
Eudesma-4 (15), 11 (13) -diene-8,12-olide; 4,7,8αH-eudesma-5 (6), 11 (1
3) -Diene-8,12-olide and 5,7,8,11αH-eudesm-4 (15) -ene-8,12-olide can be chromatographed on silica gel (for example, by filling with petroleum ether and increasing concentrations of ethyl ether). (Eluting) from 80% isohelenin (available from Sigma Chemical Company, St. Louis, Mo., USA). 4,5,7,8,11αH-Eudesman-8,12-Oride is 4,7,8αH-Eudesma-5 (6), 11 (13
) -Diene-8,12-Olide.

Useful sources or starting materials for the compounds are plant isolates described by numerous books. Some of the authors include Bohlmann et al.
New sesquiterpene lactones from (Inula Species) 1978, Phytochem. 17: 1
165-1172; Topcu et al. `` Cytotoxic and antibacterial sesquiterpenes from Inula Graveolens '' 1993, Phytochem. 33:
J. Org. Chem. 29: 1022-1026.

Compounds in which R ³ contains an amino are added by Michael addition, for example, Hejeckmann et al. (1995
, 38: 3407-3410), and the like. Ring-opened compounds can be synthesized by hydrolysis of the corresponding lactone.

A typical synthetic procedure is described in the examples below. Both 2α and 2β isomers are included. 2α is preferred. The 2β isomers can be produced from the corresponding 2α isomers using the Mitsunobu reaction (Mitsunobu et al., 1967, Bull. Chem. Soc. Japan 40: 935).

Representative compounds of the present invention include the following: 2α-cyclohexanecarbonyloxy 4,5,7,8,11αH-eudesmann-8,12-oride; 2α-cyclopentanecarbonyloxy 4,5,7,8,11αH-eudesmann-8,12-oride; 2α -Cyclobutanecarbonyloxy-4,5,7,8,11αH-eudesmann-8,12-olide; 2α- (2'naphthoyloxy) -5,7,8,11αH-eudesm-4 (15) -ene-8,12 -Olide; 2α- (1'-methylcyclohexanecarbonyloxy) -5,7,8αH-eudesma-4 (15), 11
(13) -diene-8,12-olide; 2α- (4′-trifluoromethylbenzoyloxy) -4,5,7,8,11αH-eudesmann-8,1
2-Olide; 2α- (2'naphthoyloxy) -4,5,7,8,11αH-eudesman-8,12-oride; 2α-cyclohexanecarbonyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -Diene
-8,12-oride; 2α- (3 ′, 5′-di-tert-butyl-4′-hydroxybenzoyloxy) -5,7,8,11αH-eudesm-4 (15) -ene-8,12-olide; 2α- (2'-methoxy-5'-chlorobenzoyloxy) -4,5,7,8,11αH-eudesmann-8,1
2-Olide; 2α- (trans-4'-isopropylcyclohexanecarbonyloxy) -5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; 2α-cyclopentanecarbonyl Roxy 5,7,8αH-eudesma-4 (15), 11 (13) -diene
-8,12-Olide; 2α- (4'-Ethylbenzoyloxy) -5,7,8,11αH-Eudesm-4 (15) -ene-8,12-Olide; 2α- (3'-Ethylbenzoyl) (Roxy) -5,7,8,11αH-eudesm-4 (15) -ene-8,12-olide; 2α-cyclobutanecarbonyloxy 5,7,8αH-eudesma-4 (15), 11 (13) -diene -8
, 12-Olide; 2α- (2'-methoxybenzoyloxy) -5,7,8,11αH-eudesm-4 (15) -ene-8,12-
Orido;

2α- (2′-trifluoromethylbenzoyloxy) -5,7,8,11αH-eudesm-4 (15)-
Ene-8,12-olide; 2α- (3′-fluoro-4′-methoxybenzoyloxy) -5,7,8,11αH-eudesm-4 (15)
-Ene-8,12-olide; 2α- (2'-methoxy-5'-chlorobenzoyloxy) -5,7,8,11αH-eudesm-4 (15)-
Ene-8,12-olide; 2α- (2α- (2′-chlorobenzoyloxy) -5,7,8,11αH-eudesm-4 (15) -ene-8,1
2-olide; 2α- (4′-fluorobenzoyloxy) -5,7,8,11αH-eudesm-4 (15) -ene-8,12-
Oride; 2α- (trans-4′-isopropylcyclohexanecarbonyloxy) -5,7,8,11αH-
Eudesma-4 (15), 11 (13) -diene-8,12-olide; 2α- (1′-methylcyclohexanecarbonyloxy) -5,7,8αH-eudesma-4 (15), 11
(13) -diene-8,12-olide; 2α- (4′-fluorobenzoyloxy) -4,5,7,8,11αH-eudesmann-8,12-olide; 2α- (2α- (2 ′ -Chlorobenzoyloxy) -4,5,7,8,11αH-eudesmann-8,12-olide; 2α- (2'-methoxybenzoyloxy) -4,5,7,8,11αH-eudesmann-8 , 12-Olide; 2α- (3'-Ethylbenzoyloxy) -4,5,7,8,11αH-Eudesman-8,12-Olide; 2α- (4'-Ethylbenzoyloxy) -4,5 , 7,8,11αH-eudesmann-8,12-oride; 2α- (4′-isopropylbenzoyloxy) -4,5,7,8,11αH-eudesmann-8,12-oride; 2α- (4 ′ -Chlorobenzoyloxy) -4,5,7,8,11αH-eudesman-8,12-olide; 2α- (3 ′, 5′-t-butyl-4′-hydroxybenzoyloxy) -4,5, 7,8,11αH-eudesman-8,12-oride; 2α- (2'naphthoyloxy) -5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-oride; 2α- (3'-Fluoro-4'-methoxybenzoyloxy) -4,5,7,8,11 H- Yudesuman -8
, 12-Olide;

2α- (2′-trifluoromethylbenzoyloxy) -4,5,7,8,11αH-eudesmann-8,1
2-olide; 2α- (4′-isopropylbenzoyloxy) -5,7,8,11αH-eudesm-4 (15) -ene-8,
12-Olide; 2α- (4'-chlorobenzoyloxy) -5,7,8,11αH-eudesm-4 (15) -ene-8,12-olide; 2α-cyclohexanecarbonyloxy-5,7,8αH- Eudesma-4 (15), 11 (13) -diene
-8,12-Olide; 2α-cyclopentanecarbonyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene
-8,12-Olide; 2α- (4'-trifluoromethylbenzoyloxy) -5,7,8,11αH-eudesm-4 (15)-
Ene-8,12-olide; 2α-cyclobutanecarbonyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8
, 12-Olide; 2α- (1'-methylcyclohexanecarbonyloxy) -4,5,7,8,11αH-eudesmann-8
, 12-Olide; 2α- (trans-4'-isopropylcyclohexanecarbonyloxy) -4,5,7,8,11αH
-Eudesman-8,12-olide; 2α- (4'-Fluorobenzoyloxy) -5,7,8αH-Eudesma-4 (15), 11 (13) -diene-
8,12-Olide; 2α- (2α- (2′-chlorobenzoyloxy) -5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; 2α- ( 2'-Methylbenzoyloxy) -5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,
12-Olide; 2α- (3′-ethylbenzoyloxy) -5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,
12-Olide; 2α- (4′-ethylbenzoyloxy) -5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,
12-Olide;

2α- (4′-isopropylbenzoyloxy) -5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; 2α- (4′-chlorobenzoy Roxy) -5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,
12-Olide; 2α- (3 ′, 5′-tert-butyl-4′hydroxybenzoyloxy) -5,7,8αH-eudesma-4
(15), 11 (13) -diene-8,12-olide; 2α- (2′-methoxy-5′-chlorobenzoyloxy) -5,7,8αH-eudesma-4 (15), 11 (1
3) -diene-8,12-olide; 2α- (3′-fluoro-4′-methoxybenzoyloxy) -5,7,8αH-eudesma-4 (15), 11
(13) -Diene-8,12-olide; 2α- (2′-trifluoromethylbenzoyloxy) -5,7,8αH-eudesma-4 (15), 11 (1
3) -diene-8,12-olide; 2α- (4′-trifluoromethylbenzoyloxy) -5,7,8αH-eudesma-4 (15), 11 (1
3) -Diene-8,12-olide; 2α-benzoyloxy 4,5,7,8,11αH-eudesmann-8,12-olide; 2α-benzoyloxy 5,7,8αH-eudesma-4 (15 ), 11 (13) -Diene-8,12-olide; 2α-benzoyloxy-5,7,8,11αH-eudesm-4 (15) -ene-8,12-olide; 2β-benzoyloxy 4,5 2,7,8,11αH-eudesman-8,12-oride; 2β-benzoyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; 2β-benzoi Roxy 5,7,8,11αH-eudesm-4 (15) -ene-8,12-oride; 2α-cyclopropanoyloxy 4,5,7,8,11αH-eudesmann-8,12-oride; 2α-cyclo Propanoyloxy 5,7,8,11αH-eudesm-4 (15) -ene-8,12-olide
; 2α-cyclopropanoyloxy 5,7,8,11αH-eudesma-4 (15), 11 (13) -diene-8,
12-Olide; 2β-cyclopropanoyloxy 4,5,7,8,11αH-eudesman-8,12-olide; 2β-cyclopropanoyloxy 5,7,8,11αH-eudesm-4 (15) -ene- 8,12-Olide
;

2β-cyclopropanoyloxy 5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-
Oride; 2α-fluorooxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; 2α-fluorooxy-5,7,8αH-eudesm-4 (15) -ene- 8,12-Olide; 2α-Flouroxy 4,5,7,8,11αH-Eudesman-8,12-Olide; 2β-Floyloxy 5,7,8αH-Eudesma-4 (15), 11 (13)- Diene-8,12-olide; 2β-flouroxy-5,7,8,11αH-eudesm-4 (15) -ene-8,12-olide; 2β-flouroxy-4,5,7,8,11αH-eudesmann- 8,12-Olide; 2α-isobutylyloxy-5,7,8,11αH-eudesm-4 (15) -ene-8,12-olide; 2α-isobutylyloxy 4,5,7,8,11αH-eudesmann- 8,12-Olide; 2α-isobutylyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-oride
; 2β-isobutylyloxy-5,7,8,11αH-eudesm-4 (15) -ene-8,12-oride; 2β-isobutylyloxy-4,5,7,8,11αH-eudesmann-8,12-oride ; 2β-isobutylyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide
; 2α-propanoyloxy 4,5,7,8,11αH-eudesmann-8,12-oride; 2α-propanoyloxy 5,7,8,11αH-eudesm-4 (15) -en-8,12-oride ; 2α-propanoyloxy 5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide
; 2β-propanoyloxy 4,5,7,8,11αH-eudesman-8,12-oride; 2β-propanoyloxy 5,7,8,11αH-eudesm-4 (15) -en-8,12-oride ; 2β-propanoyloxy 5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide
; 2α-acetoxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; 2α-acetoxy-5,7,8,11αH-eudesm-4 (15) -ene-8 , 12-Oride; 2α-acetoxy-4,5,7,8,11αH-eudesman-8,12-oride; 2β-acetoxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8 , 12-Olide;

2β-acetoxy-5,7,8,11αH-eudesm-4 (15) -ene-8,12-oride; 2β-acetoxy-4,5,7,8,11αH-eudesmann-8,12-oride; 4 , 5,7,8,11αH-Eudesman-8,12-Olide; 4,7,8αH-Eudesma-5 (6), 11 (13) -Diene-8,12-Olide; 5,7,8αH-Eudesma -4 (15), 11 (13) -diene-8,12-olide; 5,7,8αH-eudesm-4 (15) -ene-8,12-olide; 2α-hydroxy-4 (5), 11 (13 ) Eudesmadiene-12,8-olide; 1- [sodium 2α-benzoyloxy-8β-hydroxy-11,13-dihydro-5,7,8,11α
H-eudesm-4 (13) -en-12-oateyl] -pyrrolidine hydrochloride; sodium 8β-hydroxy-4,5,7,8,11αH-eudesmann-12-oate; sodium 8β-hydroxy-5,7,8αH-eudesma- 4 (15), 11 (13) -diene-12-oate; sodium 8β-hydroxy-4,7,8αH-eudesma-5 (6), 11 (13) -diene-12-oate; sodium 8β-hydroxy-5, 7,8αH-eudesm-4 (15) -ene-12-oate; sodium 2α, 8β-dihydroxy-2β-4,5,7,8,11αH-eudesmann-12-oate; sodium 2α, 8β-dihydroxy-4 ( 15), 11 (13) -eudesmadiene-12-oate; sodium 2α-benzoyl-8β-hydroxy-2β, 4,5,7,8,11αH-eudesmann-1
2-Oate; 8β-hydroxy-4,7,8αH-eudesmer 5,11 (13) -diene-12-oic acid; 8β-hydroxy-5,7,8αH-eudesm-4 (15) -en-12-oic acid 8β-hydroxy-4,5,7,8,11αH-eudesmann-12-oic acid; 2α, 8β-dihydroxy-2β-4,5,7,8,11αH-eudesmann-12-oic acid; 2α, 8β-dihydroxy-4 (15), 11 (13) eudesmadiene-12-oic acid; 2α-benzoyl-8β-hydroxy-2β, 4,5,7,8,11αH-eudesman-12-oic acid; 1- [2α-hydroxy-11,12-dihydro-5,7,8αH-eudesm-4 (15) -ene-8,12-orisyl] -pyrrolidine;

Embedded image 1- [2α-Benzoyloxy 11,12 dihydro-5,7,8αH-eudesm-4 (15) -ene-8,12
-Oridyl] -pyrrolidine.

After synthesis, the compounds of the present invention can be isolated and purified by established methods to yield pharmaceutically acceptable substances for administration to mammals. The compounds used in the methods of the present invention can be administered as a medicament, preferably as a medicament, ie as a pharmaceutical composition. Preferred are oral and intraperitoneal dosage forms of the compound.

The pharmaceutical compositions used in the methods of the present invention for administration to mammals and humans comprise a compound of the present invention in combination with a pharmaceutical carrier or excipient. The medicament may be in the form of a tablet (including lozenges and granules) druggy (sugar), capsule, pill, ampoule or suppository containing the compound of the invention.

As used herein, the term “drug” refers to a separate integral part suitable for pharmaceutical administration. As used herein, "dosage unit form" refers to a daily dose or multiple (up to 4 times) or less than the daily dose (up to 1/40) of the daily dose of an active compound of the present invention. Is a physically separate, unitary unit suitable for pharmaceutical administration, wherein each unit contains in combination with a carrier and / or enveloped.
Whether a drug contains a daily dose or half, one-third or one-fourth of a daily dose can be determined, for example, by administering the drug twice, three or four times daily, respectively. Depends on whether or not.

Advantageously, the compositions are formulated as dosage units, with each unit providing a fixed dosage of the active ingredient. Tablets, coated tablets, capsules, ampoules and suppositories are examples of preferred dosage forms according to the present invention. It is only necessary that the active ingredient constitute an effective amount; that is, a suitable effective amount is consistent with the proper dosage form to be employed as single or multiple unit dosages. The exact individual dosage as well as the daily dosage will, of course, be determined according to standard pharmaceutical principles under the direction of the clinician or pharmacist.

The compounds of the present invention can be administered as suspensions, solutions and emulsions of the active compounds in aqueous or non-aqueous diluents, as syrups, as granules or as powders.

The diluents that can be used in pharmaceutical compositions (eg, granules) containing the active compound to be formed into tablets, dragees, capsules, and pills are not limited. Not including the following: (A) fillers and extenders;
Corn, sugar, mannitol and silicic acid; (b) binders, such as carboxymethylcellulose and other cellulose derivatives, alginates, gelatins and polyvinylpyrrolidone, (c) humectants, such as glycerol (d) disintegrants, For example, agar-agar, calcium and potassium carbonate and sodium bicarbonate, (e) agents that retard dissolution, such as paraffin, (f) sorption enhancers, such as quaternary ammonium compounds, (g) surface activity Agents such as cetyl alcohol, glycerol monostearate, (h) adsorbent carriers such as kaolin and bentonite, (i) lubricants such as talc, calcium and magnesium stearate and solid polyethylene glycols.

Tablets, draggies, capsules and pills containing the active compounds can have conventional coatings, envelopes and protective matrices, which can contain opacifying agents. They can be configured to release the active ingredient, possibly only in certain parts of the intestinal tract or, preferably, in certain parts over a period of time. The coatings, envelopes and protective matrices can be made, for example, from polymeric substances or waxes. The compounds of the present invention can also be made in microencapsulated form with one or more of the above-mentioned diluents. The diluents used in the pharmaceutical compositions adapted to be formed into suppositories are, for example, customary water-soluble diluents, such as polyethylene glycols and fats (eg coconut oil and higher esters, such as C14 alcohols and C16 fatty acids). Or it can be a mixture of these diluents.

Pharmaceutical compositions, which are solutions and emulsions, may contain, for example, conventional diluents such as solvents,
It may contain solubilizers and emulsifiers. Specific non-limiting examples of such diluents include water, ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oil , Peanut oil, glycerol, tetrahydrofurfuryl alcohol, polyethylene glycol and fatty acid esters of sorbitol or mixtures thereof. For parenteral administration, solutions and suppositories should be sterile, for example, water or peanut oil and, if appropriate, should be isotonic with blood. Pharmaceutical compositions that are suspensions may be prepared with conventional diluents such as liquid diluents such as water, ethyl alcohol, propylene glycol, surfactants (eg, ethoxylated isostearyl alcohols, polyoxyethylene sorbitols and sorbitan esters), It can contain microcrystalline cellulose, aluminum metahydroxide, bentonite, agar and tragacanth or mixtures thereof.

The pharmaceutical compositions may also contain coloring and preservative agents as well as flavoring and flavoring agents such as peppermint and eucalyptus oil and sweetening agents such as saccharin and aspartam.
Can be contained. Pharmaceutical compositions will generally contain 0.5-90% of the active ingredient by weight of the total composition. In addition to the compounds of the present invention, the pharmaceutical compositions and medicaments may also contain other pharmaceutically active compounds. Any diluent for the agents of the present invention can be any of those described above for the pharmaceutical composition. Such drugs have a molecular weight of 2 as a single diluent.
It can contain less than 00 solvents. The compounds of the invention may be administered orally, parenterally (eg, intramuscularly, intraperitoneally, subcutaneously, transdermally or intravenously), rectally or topically, but preferably are orally, parenterally. It is administered specifically, especially perlingually or intravenously. Dosage rates (e.g., 0.5-500 mg / kg body weight) will depend on the nature and weight of the person or animal being treated, the individual response of the subject to the treatment, the type of formulation in which the active ingredient is to be administered, and the administration. Method and time of disease progression or dosing interval (
Frequency). Thus, in some cases it may be sufficient to use less than the minimum dose rate, while in other cases the upper limit may have to be exceeded to achieve the desired result. When large doses are administered, it is advisable to divide them up into several individual doses over the course of the day.

Without intending to be bound by any particular theory, in one aspect, the compounds of the present invention can inhibit the lipolysis of stored triglycerides and inhibit their metabolism to fatty acids and glycerol by inhibiting hormone-sensitive lipase activity. Interfere with. High glucose levels can be reduced as a result of its metabolism. Fat metabolism provides three times more calories of energy than carbohydrates or proteins. Thus, it is a fuel source whose use is limited only by its relative abundance. Free fatty acids are alpha in the liver
Undergoes oxidation, which in turn inhibits glycolysis by a negative feedback mechanism while simultaneously promoting gluconeogenesis. In addition, the released glycerol is used as a three carbon substrate in gluconeogenesis and thus contributes to increased levels of hepatic glucose output. Under normal circumstances of metabolic control, the enhanced lipolysis of stored triglycerides in adipose tissue occurs after a long fasting period and is quickly suppressed by the action of insulin after eating a meal. However, in the insulin resistant state of type II diabetes, the inhibition of lipolysis by insulin is lost even after eating. Under these conditions, the liver remains in a steady state of gluconeogenesis,
Glucose transport to muscle tissue is reduced, resulting in overt hyperglycemia.

The anti-lipolytic effect of the compounds of the present invention in adipose tissue results in a reduction in plasma free fatty acid and glycerol concentrations. In addition, the reversal of hepatic gluconeogenesis to glycolysis is achieved due to the reduced availability of fatty acids as a fuel source and is re-established as a major energy source, thus resulting in a final drop in plasma glucose levels. Accompanied by Reduced free fatty acids have an additional effect on pancreatic function. Elevation of free fatty acids in β-cells results in impaired insulin secretion mechanisms, which is due to the fact that compounds demonstrating NEFA-lowering properties by UNGER (Diabetes 44: 863-870, 1995) have shown As demonstrated by preventing and recovering. Thus, the compounds of the present invention achieve a reduction or normalization of elevated blood glucose and insulin levels, achieving an improved state of muscle insulin resistance in addition to their beneficial effects on circulating lipids.

In summary, the enzyme inhibition studies, lipolysis assays and animal studies described herein show that the compounds of the present invention have a unique mechanism of action and that the toxicity observed with the prior art thiazolidinediones. Is expected to show no cell transformation effect. Accordingly, they are highly beneficial and desirable therapeutic agents because lowering blood glucose levels and / or inhibiting lipolysis has utility in a variety of disease states where treatment of patients is necessary. Such disease states include diabetes, especially onset or type 2 diabetes, insulin resistance associated with dyslipidemia called dyslipidemia and euglycemia, hypertension and atherosclerosis. Is included.

The hormone-sensitive lipase (HSL) enzyme is characterized in that its activity is governed by various steroid and non-steroid hormones and secretes neurotransmitter substances via a second messenger pathway mediated by the receptor. To distinguish it from other endogenous lipase enzyme systems. Examples of such substances include insulin, corticosterone and norepinephrine. The main location of hormone-sensitive lipase is in adipose tissue, where it is involved in the coordinated control of fat metabolism. As mentioned above, the activity of this enzyme is responsible for the metabolism of triglycerides stored to provide a fuel source during fasting. Recent expression of the gene for mammalian hormone-sensitive lipase in transfected bacterial vectors has provided a means of isolating pure hormone-sensitive lipase protein. This has led to the development of in vitro assays to evaluate the direct effects of potential therapeutics on the activity of the enzyme itself, without disturbing other signals and regulators such as those found in whole cell systems.

A suitable model for assessing adipocyte function in tissue culture conditions is the transformed NI
H-3T3 cells. Culture of the 3T3 cell line with dexamethasone and 3-isobutyl-1-methylxanthine (IBMX) is capable of performing insulin-regulated lipogenesis and norepinephrene-mediated lipolysis from fibroblastic lipoblasts (adipoblasts) Differentiation into adipocytes (adipocytes) occurred.
Thus, inhibition of lipolytic activity can evaluate potential drugs under whole cell and permeabilizing conditions. This type of assay has been used to correlate the potential of a therapeutic agent to be evaluated in a purified enzyme system under appropriate intact biological conditions. Was.

Under the control of normal metabolic control, the degradation of stored triglycerides in mammalian adipose tissue is tightly controlled by the hormone insulin. When insulin levels are high, such as after release from pancreatic β-cells in response to high circulating glucose blood levels, the activity of hormone-sensitive lipase is suppressed.
In parallel, insulin levels decrease as glucose levels decrease. At some point during the prolonged fasting period, insulin levels decrease below that required for suppression of hormone-sensitive lipase. Thus, the catalytic activity of the enzyme is enhanced resulting in the release of non-esterified free fatty acids (NEFA) and glycerol from the stored triglyceride reservoir.

Thus, non-diabetic animals can be used to measure inhibition of hormone-sensitive lipase by administering a test compound prior to the onset of a fasting period when insulin levels are high. Decreased levels of NEFA in plasma after an extended 18-hour fasting period would be indicative of an inhibition of migration of triglyceride stores from adipose sites through the hormone-sensitive lipase pathway. In type II (secondary) diabetes, the insulin regulatory mechanism for lipolysis does not function properly. Inhibition of hormone-sensitive lipase activity is severely reduced and stored triglycerides have enhanced lipolytic activity due to the state of insulin resistance and, in some cases, elevated insulin levels beyond the normal range. Resulting in very elevated fasting plasma NEFA levels. More recent experimental evidence suggests that a transient state of insulin resistance can be achieved in non-diabetic individuals following infusion of NEFA directly into the circulatory system. This data would support the pathological role of elevated plasma NEFA in type II diabetes.

At this time, the exact etiology of the development of type 2 diabetes beginning in adulthood is unknown. However, this is clearly associated with obesity, as 70% of this group of patients can be classified as obese under the general criteria set forth by the American Diabetes Association National Institute of Health Guidelines. Thus, non-human mammals exhibiting similar pathophysiological profiles can serve as suitable models for evaluating potential drugs and anti-diabetic properties.

Animal models showing these overall characteristics include mouse ob / ob and K
K / Ay strains and Zucker fa / fa (ZDF) rats are included.
Ob / ob mice develop a diabetic profile at 4 weeks of age. Plasma glucose levels increase from 9 to 10 weeks, at which point the levels level off. In addition to being hyperglycemic, ob / ob mice show high levels of NEFA, triglyceride production and an insulin resistant state. Furthermore, a parallel increase in plasma insulin levels results from hyperglycemia. Hyperglycemia is improved as plasma insulin levels peak. However, the amount of insulin present is not sufficient to completely normalize the state of insulin resistance during the oral glucose tolerance test. Overall, KK / Ay is similar to ob / ob but with some differences. The onset of the type 2 profile occurs much later, at an age of approximately 3-4 months, and lasts for a year. Both mouse models show hyperinsulinemia, hyperglycemia, insulin resistance and dyslipidemia similar to those seen in the clinical setting.

[0049] ZDF rats are similar to ob / ob and KK / Ay mice in all common type II-like profile attributes, with one exception. In contrast to a mouse model showing severe hyperinsulinemia, ZDF rats show a progressive decrease in the ability of insulin to secrete insulin from alpha cells starting at 10 weeks of age, resulting in a hypoinsulinemia state. Thus, this condition resembles the subtype profile of type II diabetics who are unable to respond to certain drugs such as sulfonylureas and require insulin therapy.

The present invention can be better understood by reference to non-limiting examples, which are provided to illustrate the invention. The following examples are provided to more fully illustrate the preferred embodiments of the present invention. They should in no way be construed as limiting the broad scope of the invention.

Example 1 Preparation of Intermediates and Compounds Isolation and Purification of 2α-Hydroxy-5,7,8αH-eudesm-4 (15), 11 (13) -diene-8,12-olide Prepared as an intermediate in the preparation of some of the compounds of the invention. 2α-hydroxy-5,7,8αH-eudesm-4 (15), 11 (13) -diene-8,12-olide is I
va microcephaia Nutt was isolated by a modification of the method reported by Hertz et al., J. Org. Chem., 27: 905-910, 1962. Dry, grind and dry the flower heads, leaves and soft trunks of the plants with a mixture of petroleum ether and ether (2: 1 v / v) in a Soxhlet extractor.
Extracted with stirring for hours. Crude 2-hydroxy 5,7,8αH-eudesm-4 (15), 11 (1
3) -Diene-8,12-olide began to precipitate within 2 hours. The extract was cooled overnight at room temperature, then filtered and washed with the same mixture of solvents, and crude 2-hydroxy-5,7,8α-eudesm-4 (15), 11 (13) -diene-8,12-olide (2.93 % Yield). Crude 2-hydroxy-5,7,8α-eudesm-4 (15), 11 (13) -diene-8,12-olide crystallizes from a mixture of CH ₂ Cl ₂ , ether and petroleum ether and is 2-hydroxy-5,7,8α-eudesm -Four(
15), 11 (13) -Diene-8,12-olide, mp 127-129 ° C, 2.86% yield. 1H-N
4. MR (CDCl ₃ , 400 MHz) d6.13 (1H, d, H-13α);
59 (1H, d, H-13α), 4.87 (1H, d, H-15α), 4.54
(1H, d, H-15α), 4.49 (1H, ddd, H-8α), 3.82 (
1H, m, H-2α), 2.98 (1H, ddd, H-7α), 1.88 (1H
, Dd, H-5α), and 0.82 (1H, s, C10- CH 3). ¹³ C-NM
R (CDCl ₃ , 100 MHz) d170.5 (C-12), 145.9 (C-
4), 141.9 (C-11), 120.5 (C-13), 109.3 (C-1)
5), 76.6 (C-8), 67.1 (C-2), 50.9 (C-5), 46.
3 (C-7), 45.5 (C-9), 41.1 (C-1), 40.5 (C-3)
, 34.0 (C-10), 27.3 (C-6), and 18.7 (C-14).

Preparation of 2α-benzyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide: The following reaction scheme was used:

Embedded image2α-Hydroxy-eudesma-4 (15), 11 (13) -diene-8,12-olide (4.0 g, 16.1 mmol
) To CH_TwoCl_Two(13 ml) and stirred at 0 ° C. for 2 hours to remove excess benzoyl chloride.
(4.0 g, 28.4 mmol) and pyridine (3.1 ml) were added, and stirring was continued at room temperature overnight.
. The reaction mixture was poured into ice water with stirring. The precipitate was collected by filtration and washed with ice water
. CH to solid_TwoCl_Two(30 ml) and anhydrous Na_TwoSO_FourDried on. CH _Twoー Cl_TwoThe solution was evaporated in vacuo to a crude product (7.0 g)_TwoCl_TwoAnd 1%
MeOH-CH_TwoCl_TwoColumn chromatography eluting continuously with silica gel
Was purified. The product (5.08 g) was CH_TwoCl_TwoOf ether, petroleum and petroleum
2α-benzoyloxy 5,7,8α H-eudesma-4 (15), 11 (13) -die
Melting point: 127-129 ° C to obtain -8,12-Olide (4.5 g, 79.3%). HPL
Purity estimated by C: 99.9%.¹H-NMR (CDCl_Three, 400 MHz) δ 7.9
9 (2H, d, H-2 ", 6"), 7.55 (1H, dd, H-4 "), 7.43
(2H, dd, H-3 ", 5"), 6.14 (1H, d, H-13-α), 5.6
1 (1H, d, H-13α), 5.13 (1H, m, H-2β), 4.97 (1
H, d, H-15β), 4.63 (1H, d, H-15α), 4.51 (1H,
ddd, H-8α), 3.00 (1H, ddd, H-7α), 2.05 (1H,
dd, H-5α), and 0.94 (3H, s, C_Ten-CH_Three).¹³C-NMR (C
DCI_Three, 100MHZ) δ 170.1 (C-12), 165.8 (C-1 '),
144.9 (C-4), 141.8 (C-11), 132.9 (C-1 "), 1
30.5 (C-4 "), 129.6 (C-2", 6 "), 128.4 (C-3", 5
"), 120.5 (C-13), 110.4 (C-15), 76.4 (C-8)
, 70.3 (C-2), 46.9 (C-5), 45.7 (C-7), 44.9 (
C-9), 40.9 (C-1), 40.5 (C-3), 34.2 (C-10),
27.2 (C-6) and 18.5 (C-14). C_{twenty two}H_{twenty four}O_FourAnalyzer for
Calculated value, C% 74.90%, H 6.81%, measured value, C% 75.06%, H% 6.
95%.

Preparation of 2α-hydroxy-4,5,7,8,11αH-eudesman-8,12-olide: It was used as an intermediate in the preparation of certain compounds. 2α-hydroxy-5,7,8αH-eudesma-4 (15), 11 (13)-in methanol (150 ml) containing acetic acid (1.5 ml)
A solution of diene-8,12-olide (4.0 g) was added to platinum oxide (0.12 g).
The reaction mixture was hydrogenated on a pearl shaker at 40 psi for 17 hours at room temperature. The reaction mixture was filtered and concentrated in vacuo. The residue was washed with ice water to obtain a white powder product, 2α-hydroxy-4,5,7,8,11αH-eudesmann-8,12-olide (3.
36g, 82.7%). Mp 154-155 ° C. ¹ H-NMR (CDCl ₃ , 40
0 MHz) δ 4.42 (1H, ddd, H-8α), 3.97 (1H, m, H−
2α), 2.40 (1H, m, H-4α), 2.1 (1H, dd, H-7α),
_{1.17 (3H, d, C 11} -CH 3), 1.01 (3H, s, C 10 -CH 3) and _{0.91 (3H, d, C 4} -CH 3). ¹³ C-NMR (CDCl ₃ , 100 MHZ
) Δ 179.5 (C-12), 78.0 (C-8), 63.6 (C-2), 51
. 1 (C-5), 45.0 (C-7), 43.5 (C-11), 42.9 (C-
9) 41.6 (C-1), 41.4 (C-4), 34.5 (C-3), 34.2
(C-10), 23.8 (C-6), 22.1 (C-14), 15.6 (C-1)
5) and 9.3 (C-13).

Preparation of 2α-benzoyloxy 4,5,7,8,11αH-eudesmann-8,12-olide: 2α-benzoyloxy 4,5,7,8,11αH-eudesmann-8,12-olide Was prepared according to the following reaction scheme.

Embedded image 2α-hydroxy-4,5,7,8,11αH-eudesmann-8,12-olide (3.36 g, 13.
3 mmol) was dissolved in CH ₂ Cl ₂ (15 ml) and excess benzoyl chloride (4.0 g,
28.4 mmol) and pyridine (7.6 ml) were added with stirring at 0 ° C. for 2 hours, and stirring was continued at room temperature overnight. The reaction solution was evaporated and poured into ice water (700 ml) with stirring. The precipitate was collected by filtration and washed with ice water. The solid was dissolved in CH ₂ Cl ₂ (50ml), dried over anhydrous Na ₂ S0 _4. The CH ₂ Cl ₂ solution was evaporated in vacuo to give 7.51 g of crude product. This product was purified by silica gel column chromatography (CH ₂ Cl ₂ as eluent). The product was recrystallized from a mixture of CH ₂ Cl ₂ , ether and petroleum ether to give pure 2α-benzoyloxy 4,5,7,8,11αH-eudesman-8,12-olide (3.38 grams, 70 .8%); mp 181-
183 ° C. Purity 99.1% estimated by HPLC. ¹ H-NMR (CDCl ₃ , 4
00 MHz) δ 8.00 (2H, d, H-2 ", 6"), 7.53 (1H, dd,
H-4 "), 7.41 (2H, dd, H-3", 5 "), 5.32 (1H, m, H
-2β), 4.44 (1H, ddd, H-8α), 2.04 (1H, dd, H-
7α), 1.16 (3H, d, C ₁₁ —CH ₃ ), 1.10 (3H, s, C ₁₀ —C
_{H 3), 0.99 (3H,} d, C 4 -CH 3), 13 C-NMR (CDCl 3, 100
MHz) δ 179.4 (C-12), 166.1 (C-1 ′), 132.8 (C
-1 "), 130.7 (C-4"), 129.5 (C-2 ", 6"), 128.3 (
C-3 ", 5"), 77.6 (C-8), 68.0 (C-2), 47.1 (C-5
), 44.9 (C-9), 43.7 (C-7), 41.5 (C-1), 41.3.
(C-4), 39.1 (C-11), 34.5 (C-3), 34.0 (C-10)
), 23.8 (C-6), 21.9 (C-14), 15.5 (C-15), and 9.3 (C-13). Analysis calculated value for C ₂₂ H ₂₈ O ₄ C% 74.15%, H%
7.86. Found C 74.17%, H 8.00%.

Compound 2α-acetoxy 4,5,7,8,11αH-eudesman-8,12-olide was prepared according to the above procedure.

2α-cyclopropanoyloxy 5,7,8,11αH-eudesm-4 (15) -ene-8,12-olide and 2α-benzoyloxy 5,7,8,11αH-eudesm-4 (15)- Preparation of ene-8,12-olide: These compounds are used for the preparation of 2α-benzoyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide. 2α-Hydroxy-5,7 using cyclopropanecarbonyl chloride and benzoyl chloride, respectively, according to the procedure described in the specification.
, 8,11αH-eudesm-4 (15) -ene-8,12-oride.

5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; 4,7,8αH-eudesma
-5 (6), 11 (13) -diene-8,12-olide; and 5,7,8,11αH-eudesm-4 (15) -ene-8,
Isolation and Purification of 12-Olide: Crude 5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide, a commercial source of isofernin (Sigma, Cat. 1-1639) , Purity 80%). 5,7,8αH-
In order to purify Eudesma-4 (15), 11 (13) -diene-8,12-olide, this material (20
. 0 g) as eluent was chromatographed from silica gel with petroleum ether containing increasing amounts of ether, 100-150 ml of 60 ml each.
Fraction was obtained. Based on TLC similarities, these fractions were combined to give three major fractions, which were further purified again by chromatography on a silica gel column with the same eluent. Fractions 2 to 4 contain 4,7,8αH-eudesma-5 (6), 11 (13) -diene-8,12-olide (20.6%), and fractions 11 to 19 have 5,7,8αH -Eudesma-4 (15), 11 (13) -Diene-8,12-Olide (5
7.5%), and fractions 41 to 51 are 5,7,8,11αH-eudesma-4 (15)
-En-8,12-olide (13.0%). The structures of these compounds were derived by high field NMR, including mass spectroscopy and 2D NMR studies.

4,7,8αH-eudesma-5 (6), 11 (13) -diene-8,12-olide: columnar (from a mixture of CH ₂ Cl ₂ , Et ₂ O and petroleum ether), melting point 73 ~ 75 ° C. Estimated purity of 99.8% by HPLC (conditions: column-Dynamax C-18, detection group-U
V203 / 210 nm, flow rate -1.6 ml / min, mobile phase -65% acetonitrile)
. ¹ H-NMR (CDCl ₃ , 400 MHz): δ 6.15 (1 H, d, H-13)
β), 5.59 (1H, d, H-13α), 5.12 (1H, dd, H-6α)
, 4.78 (1H, dd, H-8α), 3.55 (1H, dd, H-7α),
2.41 (1H, m, H-4α), 2.08 (1H, dd, H-9β), 1.5
3 (1H, dd, H- 9α), 1.16 (3H, s, C 10 -CH 3), and 1.
_{06 (3H, d, C 4} -CH 3). ¹³ C-NMR (CDCl ₃ , 100 MHZ): δ
170.06 (C-12), 149.15 (C-11), 139.91 (C-5
), 121.74 (C-13), 118.85 (C-6), 76.52 (C-8).
), 42.75 (C-7), 41.81 (C-9), 39.58 (C-1), 3
7.68 (C-4), 32.80 (C-3), 32.75 (C-10), 28.
66 (C-2), 22.63 (C-14) and 16.84 (C-15).

5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide: needle-like (CH_TwoCl _Two , Et_TwoO, and mixtures of petroleum ethers). 108-110 ° C. HPL
Estimated purity 99.8% by C (conditions: column-Dynamax C-18, detector
-UV 203/210 nm, flow 1.6 ml / min, mobile phase -65% ACN). EI
MS m / z 232 [M] + (21), 217 (17), 190 (60), 17
6 (18), 164 (19), 161 (8), 145 (20), 131 (27)
, 121 (40), 107 (32), 105 (42), 93 (70), 91 (7)
7), 79 (100), 67 (76) and 53 (97).¹H-NMR (CDC
l_Three, 400 MHZ): δ 6.11 (1H, d, H-13β), 5.57 (1H
, D, H-13α), 4.75 (1H, d, H-15β), 4.48 (1H, d
dd, H-8α), 4.42 (1H, d, H-15α), 2.95 (1H, dd)
d, H-7α), 2.16 (1H, dd, H-9β), 1.82 (1H, dd,
H-5α), 1.37 (1H, dd, H-9α), 1.55 (1H, dd, H-
6α), 1.21 (1H, ddd, H-6β) and 0.81 s, C_Ten-CH_Three?
? ?¹³C-NMR (CDCl_Three, 100 MHz): δ170.71 (C-12)
, 149.03 (C-4), 142.28 (C-11), 120.12 (C-1
3). 106.68 (C-15), 76.90 (C-8), 46.28 (C-5
), 42.26 (C-7), 41.44 (C-9), 40.61 (C-1), 3
6.89 (C-3), 34.36 (C-10), 27.55 (C-6), 22.
76 (C-2), and 17.74 (C-14).

5,7,8,11αH-eudesma-4 (15) -ene-8,12-olide: needle-like (CH ₂ Cl ₂ , E
t ₂ O, and mixtures petroleum ether). Melting point 171-173C. Estimated purity by HPLC of 99.4% (conditions: column-Dynamax C-18, detector-U
V203 / 210 nm, flow 1.6 ml / min, mobile phase 65% ACN). ¹ H-NM
R (CDCl ₃ , 400 MHz): δ 4.76 (1H, d, H-15β), 4.
47 (1H, d, H-15α), 4.46 (1H, ddd, H-8α), 2.7
9 (1H, dq, H-11α), 2.33 (1H, ddd, H-7α), 2.1
5 (1H, dd, H-9β), 1.76 (1H, dd, H-5α), 1.56 (
1H, dd, H-6α), 1.45 (1H, dd, H-9α), 1.21 (3H
, D, C ₁₁ -CH ₃ ), 1.12 (1H, ddd, H-6β) and 0.79 (3
_{H, s, C 10 -CH 3} ). ¹³ C-NMR (CDCl ₃ , 100 MHz): δ170
. 90 (C-12), 149.49 (C-4), 106.48 (C-15), 7
7.92 (C-8), 46.62 (C-5), 42.33 (C-7), 41.8
5 (C-9), 41.68 (C-11), 40.41 (C-1), 36.86 (
C-3), 34.92 (C-10), 22.77 (C-2), 21.35 (C-
6), 17.86 (C-14) and 9.36 (C-13).

Preparation of 4,5,7,8,11αH-eudesman-8,12-olide: 5,7,8αH-eudesma-4 (15) in methanol (135 ml) containing acetic acid (1.0 ml) A solution of, 11 (13) -diene-8,12-olide (5.0 grams, 0.021 mole) was added to platinum (IV) oxide (0.12 grams). The reaction mixture was hydrogenated at room temperature at 40 psi overnight. The reaction mixture was filtered and evaporated in vacuo. The residue was dissolved in methylene chloride (100ml) and dried over anhydrous sodium sulfate. The methylene chloride solution was evaporated to give 4.95 grams of crude product. The product was recrystallized from a mixture of methylene chloride and petroleum ether to yield 4.52 grams of 4,5,7,8,11αH-eudesman-8,12-olide. 144-146 ° C, 88.9% yield. ¹
H-NMR (CDCl ₃ , 400 MHz) δ 4.42 (1H, ddd, H-8α)
), 2.74 (1H, m, H-4α), 2.36 (1H, m, H-7α), 1.
_{17 (3H, d, C 11} -CH 3), 0.96 (3H, s, C 10 -CH 3) and 0.
_{86 (3H, d, C 4} -CH 3). ¹³ C NMR (CDCl ₃ , 100 MHz) δ1
79.7 (C-12), 78.4 (C-8), 45.3 (C-5), 44.2 (
C-7), 42.3 (C-9), 41.7 (C-1), 41.4 (C-11),
33.7 (C-4), 33.3 (C-3), 33.1 (C-10), 24.6 (
C-2), 21.1 (C-6), 16.8 (C-14), 15.0 (C-15)
And 9.3 (C-13).

Example 2A: Additional Compounds The following additional compounds were prepared in a manner analogous to that described in Example 1 above. 2α-acetoxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide, melting point 125-127 ° C. 2α-isobutylyloxy-5,7,8αH-eudesma-4 (15 ), 11 (13) -Diene-8,12-oride, melting point 76-78 ° C 2α-propanoyloxy 5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-oride Mp 98-100 ° C 2α-Flouroxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide, mp 98-100 ° C 2α-cyclopropanoyloxy 5,7 , 8αH-eudesma-4 (15), 11 (13) -diene-8,12-
Oride, melting point 160-163 ° C 2α-acetoxy 4,5,7,8,11αH-eudesmann-8,12-olide, melting point 183-18
5 ° C 2α-cyclopropanoyloxy 5,7,8,11αH-eudesm-4 (15) -ene-8,12-olide, melting point 157-160 ° C 2α-benzoyloxy 5,7,8,11αH-eudesm-4 (15) -Ene-8,12-Olide, melting point 1
50-153 ° C

Example 2B: Further Example Preparation of 1- [2α-hydroxy-11,12-dihydro-5,7,8αH-eudesm-4 (15) -ene-8,12-oridyl] -pyrrolidine: 1- [2α-hydroxy-11 , 12 Dihydro-5,7,8αH-eudesm-4 (15) -ene-8,12-
Oridyl] -pyrrolidine was synthesized using the Michael addition method described in Hejchman et al. (1995, 38: 3407-3410). Ivalin (2α-hydroxy-4 (15), 11
(13) Eudesmadiene-12,8-olide) (1.5 mmol) was dissolved in THF (5 ml). The solution was cooled in the freezer for 15 minutes. Pyrrolidine (3 mmol) in THF (5 ml) was added to the solution of ivaline. The mixture was kept in the refrigerator at 5 ° C. for 20 hours. The solvent was evaporated. The residue was dissolved in ether and filtered through a pad of silica gel. The solvent was evaporated and the residue was kept under vacuum for 24 hours to remove traces of pyrrolidine. The product crystallized from a mixture of ethyl acetate and hexane in about 55% yield. 121-124 ° C.

In the same manner, 1- [11,12-dihydro-5,7,8αH-eudesm-4 (15) -ene-8,12-oridyl] -pyrrolidine is converted to isohelenin (5,7,8αH-eudesma-4 (15) , 11 (13) -diene-
8,12-Olide) and 1- [2α-benzoyloxy 11,12 dihydro-5,7,
8αH-eudesm-4 (15) -ene-8,12-oridyl] -pyrrolidine is synthesized from 2α-benzoyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide did.

Example 2C: Further examples Preparation of ring-opened structures: The starting compound (8 mol) is added to a solution of NaOH (8 mol) in 90% EtOH (10-20 ml if necessary) and the mixture is treated with 60-90. 2 at ℃
Stirred for ~ 3 hours.

Embedded image Carboxylate salt of the solvent the product were evaporated at reduced pressure was recrystallized from CH ₃ CN and EtOH or MeOH and ethyl acetate.

Example 3 Dosage Form The compound of the present invention can be formulated as an oral formulation for administration to patients in need of blood glucose lowering as follows. Ingredients milligrams / tablet Compound of the invention 50 Starch 50 Manitul 75 Magnesium stearate 2 Stearic acid 5 The compound, a portion of the starch and lactose are combined and wet granulated with a starch paste. The wet granules are placed on a tray and dried overnight at a temperature of 45 ° C. The dried granules are crushed in a crusher to a particle size of about 20 mesh. Add magnesium stearate, stearic acid and the remaining starch and blend the entire mixture prior to compression on a suitable tablet press. The tablets are compressed at a weight of 232 mg using an 11/32 inch punch at a hardness of 4 kg. These tablets will disintegrate within 30 minutes according to the method described in USP XVI.

Example 4 Hormone Sensitive Lipase The ability to directly inhibit the activity of hormone sensitive lipase is measured in an in vitro purified enzyme system. Recombinant hormone-sensitive lipase protein isolated from a bacterial expression vector is pre-cultured with a compound of the invention (eg, 100 μM) prior to substrate conversion,
Triolein phosphatidylcholine / phosphatidylinositol (PC / PI
) To start the reaction. Aliquots of the reaction are removed at various time points and glycerol levels are used as an indicator of enzyme activity. Pharmacological specificity is determined by culturing test compounds with other mammalian and non-mammalian lipase enzyme systems to assess inhibition of activity. The assay is between 42 μM and 835 μM.
Performed with triolein concentrations up to.

Example 5 Inhibition of Hormone-Sensitive Lipase In a similar manner as described in Example 4 above, 2α-benzyloxy-5,7,8αH-eudesma-4 (15), 11 (100 μM) 13) -Diene-8,12-olide was evaluated for its ability to inhibit the activity of hormone-sensitive lipase on cholesterol oleate PC / PI. The assay was performed using cholesterol oleate concentrations from 12.5 μM to 225 μM.

Example 6 Hormone-Sensitive Lipase and 3T3-L1 Hormone-Sensitive Lipase 2α-Benzoyloxy-4,5,7,8,11αH-eudesmann at 100 μM in a similar manner as described in Example 4 above. -8,12-Olide was evaluated for its ability to inhibit the activity of recombinant hormone-sensitive lipase and hormone-sensitive lipase from preparations of 3T3-cell lysate (lysate) on glycerol-stabilized triolein. The results are shown in Table 1.

[0070]

[Table 1]

Example 7: Additional enzyme evaluation To demonstrate the specificity of the compounds of the present invention for hormone sensitive lipase, 2α-
Inhibition of Candida rugosa lipase on triolein substrate PC / PI emulsified with benzyloxy 5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide was evaluated. . As shown in Table 2 below, 2α-benzoyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide did not inhibit enzyme activity.

[0072]

[Table 2]

Example 8: Further enzyme evaluation The specificity of the compounds according to the invention was further determined by the glycerol-stabilized triolein by a 100 μM concentration of 2α-benzoyloxy-4,5,7,8,11αH-eudesman-8,12-olide. This was demonstrated by assessing the inhibition of liver lipase and lipoprotein lipase catalyzed by substrate hydrolysis. Table 3 shows the results.

[0074]

[Table 3]

As can be seen in Table 5, the compound 2α-benzoyloxy-4,5,7,8,11αH-eudesman-8,12-olide failed to significantly inhibit the activity of the two evaluated enzymes. Was.

Example 9 Effect of the Compounds of the Invention on Lipolysis An optional in vitro assay for lipolysis was developed using 3T3-L1 cells. In this assay protocol, 3T3-L1 adipocytes were permeabilized with digitonin and pre-cultured with compounds of the present invention. This was followed by activation of hormone-sensitive lipase with cAMP, which resulted in the hydrolysis of triglycerides to fatty acids and glycerol. Released glycerol in the supernatant was quantified by a fluorescence assay performed in a microfluor B flat bottom plate.

The protocol for this lipolysis assay is as follows. 3T3-L1
The adipocytes used were 14 days after the start of differentiation. The medium was carefully removed from the adipocytes. Cells were gently washed twice with 300 μl of PBS. Cells were cultured with 0.2 ml of a metabolic buffer consisting of 10 mM sodium phosphate, 150 mM sodium chloride, pH 7.4, 1% fatty acid free BSA and digitonin (10 mg / ml). After incubation at 37 ° C. for 30 minutes, an additional 0.2 ml of permeabilizing buffer containing the test compound was added and the cells were incubated for an additional 30 minutes. The buffer was then completely removed, 200 μl of permeabilizing buffer containing the test compound, N ⁶ -benzoyloxy-cAMP (0.6 mM), 8-thimethyl-cA.
Replaced with 200 μl of permabilizing buffer containing MP (0.6 mM), 2 mM ATP and 4 mM MgCl ₂ . The addition of cAMP activates cAMP-dependent protein kinase, which further phosphorylates and activates hormone-sensitive lipase. The cells were then cultured at 37 ° C. for 30-60 minutes. A 350 μl aliquot of buffer was removed and mixed with 100 μl of a 10% charcoal solution. The suspension was centrifuged (3000 RPM, 20 minutes) and 50 μl of the supernatant was transferred to a 96-well flat bottom plate (Microflor B) and frozen at −20 ° C. until assayed for glycerol by the following procedure.

Glycerol Assay: The amount of glycerol produced during lipolysis was measured in the following two-step reaction.

Embedded image The reaction buffer was a kinase buffer, pH 7.0 (2 mM MgCl ₂ , 100 μl triethanolamine, 2 mg / ml fatty acid-free BSA), 130 μM ATP, 2 units /
Consists of ml glycerol kinase and 1 unit / ml glycerol phosphate oxidase. In the first step of the reaction, 50 μl of reaction buffer is added to 50 μl of the supernatant from the lipolysis assay. Standard curves ranging from 0 to 12000 pmol are generated with standard glycerol diluted in permeabilization buffer. Seal the plate and incubate at 37 ° C. for 30 minutes. The reaction results in the conversion of glycerol to dihydroxyacetone phosphate and hydrogen peroxide.

In the second step of the reaction, the released hydrogen peroxide is converted to a quantitable fluorogenic substrate (
(Pierce, Rockford, Ill.) And HRP. 50
Add 1 μl of the Quantable Working Solution to the sample and the standard, followed by 50 μl of 250 units / ml of HRP. The Quantable Working Solution is a Quantable Substrate Solution of Cube (Pierce, Quantable Fluorescent Paoxidase Substrate Kit, cat. # 15169) and some Kinase Buffer (2 mM MgC
l ₂ , 100 mM triethanolamine, 2 mg / ml BSA without fatty acid), p
Made by mixing H7.0. The plate was sealed and incubated at 37 ° C. for 30 minutes. The reaction was stopped by adding 50 μl / well of a Quantable Stop Solution (Pierce, Quantable Fluorescent Paoxidase Kit, Cat. # 15169). The relative fluorescence units (RFU) of the product were 320 nm and 405 n, respectively.
m were measured for excitation and emission, respectively, and corrected for background fluorescence.
The final data was expressed as% inhibition of lipolysis. CAMP of lipolysis in calculation
Stimulation was considered as 100% stimulation or conversely 0% inhibition. The final data was expressed as% inhibition listed below.

(Equation 1) The RFU values have been corrected, ie, the background has been subtracted.

Example 10: Circulating Free Fatty Acid Levels in Normal Fasted Normal Mice Male C57 / B16J from Jackson Laboratories, Bar Harbor, ME
Purchased a thin mouse. These animals were housed and maintained according to NIH guidelines and had free access to water and food. Test compounds of the invention were suspended in 0.25% mytilcellulose (w / v in distilled water) and stored at -20 ° C as single use aliquots. Animals were dosed daily via the intraperitoneal route of administration for two weeks. After the last dose, the animals were fasted overnight. The following morning, a 250 μl blood sample was dispensed via the retro-orbital sinus into the EDTA-
It was withdrawn into a test tube containing an aqueous salt solution and centrifuged at 1200 × g for 15 minutes. The plasma fraction was analyzed using the NEFA C diagnostic kit (Wako Chemical USA, Inc.).
Unesterified Free Fatty Acid Content (NEF, Richmond, VA)
A) was analyzed and glycerol was analyzed with a triglyceride GPO-Trinder diagnostic kit (Sigma) modified for use on a BM-Hitachi 911 clinical chemistry analyzer. P values from the Student's t-test were calculated using the Prism software data analysis program (Grafado, Sorrento, CA).
The data is shown in Table 4 below. As you can see, the animal is 2α-benzoyloxy
Treatment with -5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide at a dose of 50 mg / kg resulted in a 27% reduction in fasted NEFA plasma levels. .

[Table 4]

Example 11 Effect on Fasted KK / Ay Mice Male KK / Ay mice were purchased from Clare Japan USA of Pennington, NJ. At the age of 4 months, the animals received daily intraperitoneal administration of 2α-benzoyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide for 2 weeks. NEFA values in fasted plasma were obtained as described in Example 10. In addition, cholesterol and triglycerides were measured on a BM Hitachi 911 clinical chemistry analyzer using a BM reagent assay kit. Data show significant significance of NEFA, triglycerides, cholesterol and glycerol in mice receiving 2α-benzyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide Showed a decrease. Glucose levels in plasma were measured on a Boehringer Mannheim Hitachi 911 clinical chemistry analyzer using the Boehringer Mannheim reagent assay kit. Control mice exhibited fasting plasma glucose levels of 184 ± 24 mg / dL, while animals receiving the test compound exhibited plasma glucose levels of 90 ± 29 mg / dL (p <0.01
).

Example 12: Effect on fasted ob / ob mice Male C57 / B16J ob / ob mice were purchased from Jackson Laboratories (Bar Harbor, ME). At 8-9 weeks of age, the animals received daily administration of the compound at the indicated doses for 2 weeks by injection via the intraperitoneal route. Test compounds of the present invention were prepared as described previously in Example 10. 2 weeks or 4 of treatment
Animals were fasted overnight during any of the weeks and plasma NEFA levels were measured as described in Example 11. Data are expressed as% reduction compared to animals receiving vehicle alone. Test compounds induced a decrease in NEFA levels in the treated animals. Glucose levels are measured in retro-orb sinus (retro-orbital) in heparinized microcapillaries.
Ital was measured using a Johnson & Johnson one-touch glucometer from a 20 μl blood sample collected via sinalus. Fasting glucose values for the vehicle group ranged between 250-350 mg / Dl as compared to 100-120 mg / Dl for normal non-diabetic mice. This data was expressed as% normalized using the following algorithm. a) Range = vehicle control value-normal value b)% normalized = (vehicle control value-compound treated value) / range

Example 13 Effect on Circulating Free Fatty Acid Levels in Fasted ZDF Rats Genetic Model Similar in male ZDF rat model obtained from Inc. (Indianapolis, IN) with type II diabetes The test was performed. Animals received daily intraperitoneal administration of compound as previously described. NEFA levels in plasma were obtained in the fasted state 6 weeks after treatment. As seen in the KK / Ay mouse model described in Example 11, a similar effect on lowering fasted NEFA levels occurred in ZDF rats.

Example 14: Effect on insulin resistance in KK / Ay mice As described in the previous example, KK / Ay mice received the compound. Two weeks after treatment, animals fasted overnight are dosed with a 2 g / kg glucose solution made in deionized water and, at regularly scheduled times, venous glucose is measured from the tip of the tail. Oral glucose tolerance test (OG)
TT) was performed. For further comparison, data obtained from young non-diabetic C57 / B16J mice were included. Calculations of the area under the curve (AUC) shown in Table 2 were performed using the rectangle addition procedure built into the Prism Data Software Analysis Program (Grafad, Sorrento, CA). The% normalized AUC was derived using the following algorithm. a) Range = vehicle control value-normal value b)% normalization = (vehicle control value-compound treatment value) / range Treatment of mice with test compound resulted in significant normalization of glucose treatment by challenge occured.

Example 15: Effect on obese / ob mice on normoglycemic insulin resistance Female ob / ob mice were administered the compounds as described above. OGTT was performed 5-6 weeks post-dose, as described previously in Example 14. Initial fasting glucose values were obtained from non-diabetic mice. In the absence of predominant fasting hyperglycemia as defined by the American Diabetes Association at levels> 140 mg / Dl, along with an insulin resistance condition clinically referred to as Syndrome X, the compounds of this invention show this condition. Induced a significant normalization of the treatment of glucose.

All publications and references, including but not limited to patents and patent applications, cited in this specification are incorporated by reference in their entirety as if each individual publication and reference were incorporated herein by reference. Are individually incorporated herein by reference as if fully set forth. All patent applications for which this application claims priority are also incorporated by reference herein in their entirety in the manner described above for publications and cited references. Although the present invention has been described with a particular emphasis on preferred embodiments, those skilled in the art will recognize that changes may be made in the preferred apparatus and methods and that the present invention may be practiced otherwise than as specifically described. It is clear that what is possible is also intended. Accordingly, the invention is intended to cover all modifications falling within the spirit and scope of the invention as defined by the appended claims.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI theme coat ゛ (Reference) A61P 3/06 A61P 3/06 3/10 3/10 C07C 59/11 C07C 59/11 69/003 69 / 003 D 69/013 69/013 E 69/16 69/16 69/743 69/743 69/78 69/78 C07D 405/08 C07D 405/08 (81) Designated country EP (AT, BE, CH, CY, DE, DK, ES, FI, FR, GB, GR, IE, IT, LU, MC, NL, PT, SE), OA (BF, BJ, CF, CG, CI, CM, GA, GN, GW, ML, MR, NE, SN, TD, TG), AP (GH, GM, KE, LS, MW, SD, SL, SZ, TZ, UG, ZW), EA (AM, AZ, BY, KG, KZ) , MD, RU, TJ, TM), AE, AL AM, AT, AU, AZ, BA, BB, BG, BR, BY, CA, CH, CN, CU, CZ, DE, DK, EE, ES, FI, GB, GD, GE, GH, GM, HR , HU, ID, IL, IN, IS, JP, KE, KG, KP, KR, KZ, LC, LK, LR, LS, LT, LU, LV, MD, MG, MK, MN, MW, MX, NO, NZ, PL, PT, RO, RU, SD, SE, SG, SI, SK, SL, TJ, TM, TR, TT, UA, UG, UZ, VN, YU, ZA, ZW (72) Invention Terrekgey, Ihol United States 07450 Ridgewood, New Jersey East Ridgewood Avenue 333 (72) Inventor Lesek, John United States 11210 Brooklyn Henry Street, New York 248 (72) Inventor Eagan, Jack United States 10021 New York, New York East 69 Street 169 Apartment 6-day F-term (reference) 4C037 XA01 4C063 AA01 BB03 CC76 DD03 EE01 4C086 AA03 BA05 GA07 NA14 ZC33 ZC35 4C206 AA02 AA03 DA11 GA06 KA01 KA04 NA14 ZC33A AB23 BN20 BS10

Claims (17)

[Claims] 1. A compound of the formula having an AB ring structure. Wherein the dotted line between carbons 5 and 6, the dotted line between carbons 4 and R ^2, and carbon 11
The dashed line between and R ³ indicates that a double bond may exist independently from each numbered carbon to the carbon to which it is attached, wherein R ¹ is hydrogen, A hydroxy, lower alkanoyloxy or alloyoxy group, wherein the aryloxy is one or more lower alkyl, lower alkoxy, halo, trifluoromethyl, di (lower) alkylamino, hydroxy, nitro, or C1-C3 alkylenedioxy group Wherein X is hydroxy or oxy bonded to carbon 8 and carbon 12 together with the Y group, wherein Y is hydroxy or lower alkyl. it is a amino group which may be mono- or di-substituted lower alkyl wherein the can also form a 5- or 6-membered ring together, wherein R ² and ³ is an independently lower alkyl or alkenyl group, wherein the lower alkyl R ³ is at the carbon bonded to C11, substituted with mono- or di-substituted by also be an amino group with lower alkyl Where the lower alkyls can also together form a 5- or 6-membered ring, provided that when R ¹ is hydrogen, (a) X is hydroxy or (b) R ³ is amino. Is replaced by A method of lowering blood lipid levels or lowering blood glucose in a mammal, comprising administering to the mammal one or more effective lipid-lowering or glucose-lowering amounts of the compound or a pharmaceutically acceptable salt thereof. 2. The method according to claim 1, wherein the hydrogen substituent at the 5, 7, and 8 positions relative to the AB ring structure is α. 3. The method according to claim 1, wherein R ¹ is a lower alkyloxy group. 4. The method of claim 3, wherein said alkanoyloxy group is selected from the group consisting of acetoxy, propanoyloxy, isobutylyloxy, and cyclopropanoyloxy. 5. The compound according to claim 1, wherein the compound is 2α-acetoxy-5,7,8αH-eudesmer 4 (15), 11 (13) diene-8,12-olide; 2α-propanoyloxy-5,7,8αH-eudesma-4 (15 ), 11 (13) diene-8,12-oride; 2α-cyclopropanoyloxy5,7,8αH-eudesmer 4 (15), 11 (13) -diene-8,1
2-Olide; 2α-acetoxy 4,5,7,8,11αH-eudesman-8,12-oride; 2α-cyclopropanoyloxy 5,7,8,11αH-eudesm-4 (15) -ene-8,12-oride 4. The method according to claim 3, wherein the method is selected from: 6. The method of claim 1, wherein R ¹ is alloyoxy. 7. The method according to claim 6, wherein said alloyoxy group is selected from the group consisting of floyloxy and benzoyloxy. 8. The compound according to claim 1, wherein the compound is 2α-benzoyloxy-5,7,8αH-eudesma-4 (15), 11 (13) -diene-8,12-olide; -4 (15), 11 (13) -diene-8,12-olide; 2α-benzoyloxy 4,5,7,8,11αH-eudesman-8,12-olide; and 2α-benzoyloxy 5, The method according to claim 7, wherein the method is selected from the group consisting of 7,8,11αH-eudesm-4 (15) -ene-8,12-olide. 9. The method of claim 1, wherein the compound is administered in a lipid lowering effective amount. 10. The method of claim 1, wherein the compound is administered in a glucose lowering effective amount. (1) X is oxy; (2) R ¹ is lower alkanoyloxy or alloyoxy, provided that R ² is methylene bonded to C 4 by a double bond, and R ³ double bond Wherein R ¹ is not ethanoyl (ethanoyloxy), and (3) R ² and R ³ are each independently a lower alkyl group or an unsaturated group having 4 or 4 carbon atoms.
2. The method of claim 1, wherein the alkenyl is a lower alkenyl restricted to a bond to 11. 12. (1) X is oxy; (2) R ¹ is lower alkanoyloxy or alloyoxy; (3) R ² and R ³ are each independently a lower alkyl group or unsaturated carbon 4 or carbon
2. The method of claim 1, wherein the alkenyl is a lower alkenyl restricted to a bond to 11. 13. A compound of the formula Wherein the dotted line between carbons 5 and 6, the dotted line between carbons 4 and R ^2, and the dotted line between carbons 11 and R ^3, until carbon bonded carbon of the respective numbers independently Wherein R ¹ is a hydrogen, hydroxy, lower alkanoyloxy or alloyoxy group (where the aryloxy is one or more lower alkyl, lower alkoxy, , Halo, trifluoromethyl, di (lower) alkylamino, hydroxy, nitro, or C1-C3 alkylenedioxy groups, wherein X is hydroxy or together with the Y group Bonded to carbon 8 and carbon 12 is oxy, wherein Y is hydroxy or an amino group which may be mono- or di-substituted with lower alkyl, wherein Lower alkyl can also form a 5- or 6-membered ring together, mono a respectively in R ² and R ³ wherein independently lower alkyl or alkenyl group, wherein the lower alkyl R ³ is a lower alkyl Or the lower alkyl may be substituted with an amino group which may be disubstituted, wherein the lower alkyl may form a 5- or 6-membered ring together, provided that (1) when X is hydroxy, , R ¹ is not hydrogen, (2) when R ¹ is hydroxy, X is (a) X is hydroxy or (b)
R ³ is substituted by amino; (3) when R ² is a methylene bonded to C 4 by a double bond, a methylene bonded to C 11 by an R ³ double bond, and when X is oxy, , R ¹ is not an ethanoloxy. ] The compound of. 14. The compound according to claim 13, wherein X is hydroxy. 15. The compound according to claim 14, wherein R ¹ is lower alkanoyloxy or alloyoxy. 16. The compound according to claim 13, wherein R ¹ is lower alkanoyloxy. 17. The compound according to claim 13, wherein R ¹ is alloyoxy.