Polyhydronaphthylethyl tetrahydro-2h-pyran-2-one derivatives and hydroxy acids and esters thereof,their preparation and pharmaceutical compositions containing them

62044/3 1Ίκ-2-ΐΝΊ>3-2Η-ητ’ΠκΊϋο > ·ίικ5’no a’op m’η’5 το ΐιπΐηη ΠΊηρίΊ >Ί>ϋΡηι onaph ,οη5ϋ ο’Ίοονί ΌρΊΊΐ’η πίυοίπί ηηικ tnb’OQh

Polyhydronaphthyl ethyl tetrahydro-2H-pyran-2-one derivatives and hydroxy acids and esters thereof, their preparation and pharmaceutical compositions containing them MERCK & CO. INC. C. 57124 62044/2

SUMMARY OF THE INVENTION

This invention relates to a group of 6(R)-[2- (8'-acyloxy-2*-methyl-6'-methyl(or hydrogen)-polyhydronaphthyl-1’ )-ethyl]-4(R)-hydroxy-3,4,5,6-tetrahydro-2H-pyran-2-ones and to the hydroxy acid form of said pyranones, the pharmaceutically acceptable salts of said hydroxy acids and to the lower alkyl and phenyl, dimethylamino or acetylamino substituted lower alkyl esters of said hydroxy acid.

More specifically,'this invention relates to compounds of the formula

<img img-format="tif" img-content="drawing" file="IL62044AD00021.tif" id="idf0001" />

wherein R' is H or CK^; A is hydrogen or the group R-CO-, provided that when A is hydrogen R' is CH^, R is 1,1-dimethylpropyl ; R^ is OH and R^ is H or R^ and R^ together form a bond, the dotted lines at X, Y and Z represent possible double bonds, said double bonds, when any are present, being either X and Z in combination or X, Y or Z alone; and pharmaceutically acceptable salts, C^^alkyl esters, phenyldimethylamino-, or acetylamino-substituted-C^_4 alkyl esters of those compounds in which R^ is OH and R2 is H. 62044/2

The above formula also covers the 6(R)-[2-(8'-hydroxy-2',6 1 -dimethylpolyhydronaphthy1-1')ethyl]-4(R)-hydroxy-3,4,5,6-tetrahydro-2H-pyran-2-ones as intermediates for the above-mentioned 8'-acyloxy compounds.

BACKGROUND OF THE INVENTION

It is known that certain mevalonate derivatives inhibit the biosynthesis of cholesterol, cf. F. M. Singer et al, Proc. Soc. Exper. Biol. Med.,.102, 370 (1959) and F.H. Hulcher, Arch. Biochem. Biophys., 146, 422 (1971). Nevertheless, the activity of these known compounds has not always been found to be satisfactory, i.e. to have practical application.

Recently, Endo et al, reported (U.S. Letters Patent 4,049,495, Patent 4,137,322 and Patent 3,983,140) the production of fermentation products which were quite active in the inhibition of cholesterol biosynthesis.

The most active member of this group of natural products, now called compactin, IIIa(R*=H) was reported by Brown et al [J.Chem.Soc.Perkin I 1165 (1976)] to have a complex mevalonolactone structure.

More recently, Monaghan et al in U.S. Patent 4,231,938, which is incorporated herein by reference, reported an inhibitor, designated MK-803 and having the structure IIIa (R’=CH3) in Table I, which was isolated from an entirely different fermentation. Albers-Schonberg et al ( U.S. Patent No. 4,294,846) d described a dihydro MK-803, designated III^ (R'=CH3) in Table I, of equal potency to MK-803 isolated from the same fermentation as was MK-803. Patchett et al ( U.S. Patent No. 4,35 h, 8444- : describe dihydro and tetrahydro derivatives of MK-803 of different structures (111^ c and θ (R'=CH3) in Table I) prepared by the catalytic hydrogenation of MK-803. 62044/2 - 3 - - ---- A tetrahydro analog IIIe(R’=H), of compactin was reported in published Japanese Application (Kokai) 55009-024.

Very recently a dihydro-analog of compactin of structure III^ (R=H) was isolated from compactin fermentation broths as reported by Gullo et al, (U.S. Patent No. 4,343,814) . '

The preparation of the starting materials, III

CL and IIId, (R’=CH3) are the products of fermentation with a strain of Aspergillus terreus, ATCC No. 20542, designated MF-4845 in the culture-collection of MERCK &amp;amp; CO., Inc., Rahway, New Jersey.

Preparation of Compounds III = and III. (R'=CH,) Q__Q □ A. Fermentation A tube of lyophilized culture MF-4845 was opened aseptically and the contents suspended in an unbaffled 250 ml Erlenmeyer flask (seed flask) containing approximately 10 ml of the Medium which has the following composition:

Medium

Corn steep liquor 5 g

Tomato paste 40 g

Oatmeal 10 g

Glucose 10 g

Trace Element Solution 10 g

Distilled water 1000 ml

pH 6.8 with NaOH 4

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Trace Element Solution:

FeSO4.7H2O 1000 mg MnSO4·4H2O 1000 mg CuC12.2H2O 25 mg CaCl2.2H2O 100 mg H3BO3 56 mg (NH4)6Mo7024.4H20 19 mg ZnSO4.7H2O 200 mg Distilled Deionized Water 1000 ml

The inoculated flask was incubated for 24 hours at 28°C on a 220 rpm shaker (2 inch throw). An unbaffled 2 liter Erlenmeyer flask containing 500 ml of the medium was then inoculated with 10 ml of the first stage fermentation growth from the seed mixture. This too was shaken 24 hours at 28°C. A 200 gallon stainless steel fermentation vat was then charged with 485 liters of a medium comprising:

Cerelose 4.5% wt/vol

Peptonized Milk 2.5% wt/vol

Autolyzed yeast 0.25% wt/vol

Polyglycol P2000 0.25% vol/vol whose pH was adjusted to 7.0. This was sterilized 15 minutes at 121°C. One liter of the second stage above was then charged and the mixture was incubated at 85 rpm for 12 hours then at 130 rpm for 84 hours at 28°C with an air flow of 5 cfm for 12 hours then 10 cfm for 84 hours. B. Isolation 1. Extraction

Two batches of one hundred gallons of whole broth were combined, acidified with stirring to pH 4.1 by careful addition of 800 ml of concentrated hydrochloric acid, and extracted by addition of 75 gal of ethyl acetate and further stirring for two hours. 5

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About 25 lbs of a silicaceous filter aid was then added and the total slurry was pumped through a 24-inch filter press. An additional 75 gal of ethyl acetate was used to wash the press cake and continue the extraction, by reversing the direction of pumping through the press four times. Then all of the wash solvent was discharged from the press and combined with the first filtrate. The two-phase filtrate was allowed to settle, and the water layer removed. The ethyl acetate layer was washed with 10 gal of deionized water, the phases were allowed to separate and the ethyl acetate extracts were concentrated under vacuum to a residue of about 10 gal. 2. Lactonization

Ethyl acetate extracts from an additional three hundred gal of broth were added to the above extract and the volume was reduced to about thirty gal by vacuum distillation. About fifty gal of toluene was added, and the batch was concentrated under vacuum to 32 gal; this step was repeated; then sufficient new toluene was added to bring the volume to 75 gal. Without vacuum, the batch was brought to reflux and maintained there for two hours, with a temperature over 106°C.

This solution was then concentrated under vacuum to a small volume, which was further concentrated to an oily residue in a large rotary evaporator under vacuum. 3. Chromatography on Silica Gel

The extract obtained above was flushed free of other solvents by addition of 2 gal of methylene chloride and reconcentration to an oil.

The oily residue was dissolved in about 5 gal of ethyl acetate-methylene chloride (30/70; v/v) mixture, and a slurry was made by addition of 2.8 kg of silica gel. 6

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The slurry was loaded as a level layer on the top of a 12 in. x 50 in. silica gel column packed in the same solvent mixture.

Elution was with ethyl acetate-methylene chloride (40/60; v/v) at 800 ml/min. A forerun of 10 gal, then further fractions of 4 gal each were collected.

Fractions 6-10 inclusive were concentrated under vacuum to an oily residue which was dissolved in hot ethyl acetate, treated with decolorizing carbon, filtered hot, and cooled. Crystals of Compound III_ (R*=CH,) were

Cl «J filtered off and the mother liquors were concentrated to an oil for further chromatography. Pure III (R'=CH,) cL ό has m.p. 170-171°C. 4. Rechromatography on Silica Gel

Mother liquor residues from similar broth extract work-ups equivalent to an additional 600 gal of fermentation production were combined with the above in methylene chloride solution. One-half of this solution was taken for further silica gel chromatography. A small aliquot showed a total solids content of 325 g. The solution was treated with 40 g of decolorizing carbon, filtered, and the cake rinsed with methylene chloride.

The combined filtrate and washings were concentrated under vacuum to an oily residue. This was redissolved in 800 ml of ethyl acetate/methylene chloride (30/70; v/v) and slurried with 225 g of silica gel. The slurry was loaded on top of a 14 x 36 cm column bed of silica gel packed in the same solvent mixture. Development was with ethyl acetate/methylene chloride (40/60; v/v). A forecut of three liters was set aside; then fractions of 800 ml each were collected. 5. Chromatography on Reverse-phase Packing

Forty ml from fraction 12 of the above chroma tography were concentrated to an oil weighing 500 mg and 7

16449Y the oil redissolved in 5 ml acetonitrile. This acetonitrile solution was charged to a 5/8" OD by 6 ft long stainless steel chromatography column packed with preparative reverse-phase liquid chromatography column packing material "Bondapak Cl8/PorasilB" (Waters Associates, Inc., Milford, Mass. 01757). The column was eluted with a mixture consisting of (v/v) 55% acetonitrile and 45% 0.05 M ammonium phosphate pH3. The elution volume between 1360 ml and 1700 ml was combined on the basis of refractive index detection. The organic solvent was removed in vacuo and the residual aqueous solution extracted with ethyl acetate. In vacuo removal of the ethyl acetate left 120 mg of compound which crystallized from a concentrated acetonitrile solution yielding crystals of Compound II (R* =CH3), m.p. 129-131°C.

Preparation of Compounds III, d f c f e

Starting materials IIIb, IHC and ΙΙΙθ (R'=CH3) are prepared in accordance with the following Flow Sheet and preparative methods.

The desmethyl analogs, III^, IIIC an&lt;^ IJIe are obtained substantially as described by Patchett et al. but starting with III (R'=H) in each case, cl

For the preparation of ΙΙΙθ it is advantageous to reduce III^ inasmuch as the desired trans fusion of the perhydronaphthalene ring, present in the starting materials, is retained in the final product, and the need to separate isomers is avoided.

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FLOW SHEET

<img img-format="tif" img-content="drawing" file="IL62044AD00022.tif" id="idf0002" />

(4)

III 9

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Reactions and Reagents 1. Hydrogenation at about 20-75°C and about atmospheric pressure to about 4 atmospheres over tris-(triphenylphosphine)chlororhodium in an aromatic 5 solvent such as benzene, toluene or xylene, preferably toluene. Preferred conditions are about 4 0°C and about 2-7 atmospheres in toluene. 2. Hydrogenation at about 20-25°C and about atmospheric pressure over 5% palladium on calcium carbonate in a lower alkanol such as a Cj_3 alkanol, especially ethanol. 3. Hydrogenation at about 20-25°C and atmospheric pressure over platinum oxide in ethyl acetate. 4. Hydrogenation at 20-25°C and atmospheric 15 pressure over 10% Palladium on charcoal in ethyl acetate.

V 10 16449γ •

Preparation of 6α, 12-(8^-2-(S) -methylbutyryloxy-2 ' β , 6 ' α-di-methyl-1' , 2 ’ , 3 ' , 4 1 ,6 ' , 7 ' , 8 ' , 8 'a-octahydronaphthyl-1) ethyl]-4(3-hydroxy-3,4,5,6-tetrahydro-2H-pyran-2-one, III^ (R'-CH^) A mixture of 50 mg (0.1236 mmol) of Compound III = cl 5 (r’=CH3) and an equal molar amount (114.35 mg, 0.1236 mmol) of tris(triphenylphosphine)chlororhodium in 10 ml of dry toluene was hydrogenated at room temperature for 6 days, with a total uptake of 14.6 ml of hydrogen. The mixture was evaporated in vacuo to dryness. The 10 red residue was subjected to preparative thin-layer chromatography on silver nitrate impregnated silica plates and was developed twice in the 10% ethyl acetate-ether system. The yield of Compound III^ (R'=CH3) was 22.3 mg. 15 Mass spectrum (M/^ ) 406 (m+) 304 (m-102) 286 (m-102-18) 20 nmr (CDC13, 300MHz) J 4.37 (m,lH) 4.60 (m,lH) 5.34 (d of t, J=2.5 Hz, 1H) 5.41 (m,lH) - 11 - 16449Υ

Preparation of 6α [2-(δ'β-2- (S) -metn.ylbutyryloxy-2 ' β , 6 'α-dimethyl-l’, 2#, 3', 5', 6', 7', 8', 8a-octahydronaphthyl-l) ethyl] -4 β-hydroxy-3,4,5,6-tetrahydro-2H-pyran-2-one, I.II (R*=CH3) A solution of 80.91 mg (0.2 mmol) of Compound IIIa 5 (R*=CH3) in 10 ml of absolute ethanol, in the presence of an equal weight of 5% Pd on CaCO3 was hydrogenated at 1 atmosphere until an uptake of one mole equivalent of hydrogen was observed. The catalyst was then removed by filtration and the filtrate was evaporated to dryness (81 mg). After 1° a purification by preparative thin-layer chromatography to remove a small amount of by-product tetrahydro compound, 72 mg of the 1,4 reduction product ΠΙ (R’-CH.) was c o isolated.

Mass spectrum (M/x.) 406 (m+) 15 304 (m-102) 286 (304-H2O) 20 nmr (CDCl3, 300MHz) &amp;amp; 4.38 (m,lH) 4.64 (m,lH) 5.28 (d of t, J=3.5Hz, IH) 5.48 (m,lH) 12

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Preparation of 6α- 12- (Ό'β-2 (S) -methylbutyryloxy-20,, 6^-dimethyl-l', 2Z, 3', 4', 4aa, 5', 6Z, 7', 8', 8a-decahydronaphthyl-l) -ethyl]-4 3-hydroxy-3,4,5,6-tetrahydro-2H-pyran-2-one, ΙΙΙθ (R’=CH3)

A solution of 80.91 mg (0.2 mmol) of Compound III u (R*=CH3) in 10 ml of ethyl acetate was hydrogenated in the presence of an equal weight of platinum oxide at one atmosphere. An exact 2 mole equivalent of hydrogen was consumed within 1 hour. The catalyst was removed by filtration and the filtrate was concentrated to dryness to give an oil. The cis and trans isomers were separated by preparative thin-layer chromatography on silica gel plates (10% ethyl acetate - ether system, bands detected by water spray). The trans isomer ΙΙΙθ (R,=CH3) appears as the more polar spot, compared to the cis isomer, and 60 mg was isolated.

Mass spectrum (M/^ ) 408 (m+) 323 (m-85) 306 (m-102) 20 nmr (CDC13, 300MHz) S 4.36 (broad singlet, 1H) 4.59 (m,lH) 5.19 (d of t, J=2.5Hz, 1H) 13

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Fermentative Production of Compound ΠΙ^ (R'=H) A. Fermentation: A natural isolate of Penicillium citrinum, NRRL 8082 was used to prepare a yeast-malt extract (YME) slant which was incubated for 2 weeks at 28°C. A portion (1/5) of the slant (MF-4870a) was used to inoculate each of 5 unbaffled seed flasks (250 ml) containing 44 ml of KF seed medium with CaCl2· They were incubated for 3 days at 28°C, and 220 rpm. A portion of the seed growth (about 1.5 ml) was used to inoculate each of 100 production medium flasks (250 ml unbaffled) containing 40 ml of LM Production Medium Without Malt Extract. The production flasks were incubated for 4 days at 25 °C.

Another group of production medium flasks (140), each containing 40 ml of LM Production Medium Without Modification were inoculated and incubated under the same conditions as previously described.

The broths from both fermentations were combined.

The various media employed in the foregoing fermentations are : YME Slant

Dextrose Malt Extract Yeast Extract Agar

Dist. Water PH 4 g./l. 10 g./l. 4 g./i. 20 g./l/ to 1 liter 7.0 14

16449Y KF Seed Medium with CaCl2

CaCl2 Corn steep liquor 10 g. 5 9· Tomatoe Paste 40 g. Oatmeal 10 g. Cerelose 10 g. Trace Element Mix 10 ml. Distilled Water 1000 ml. PH 6.8

Trace Element Mix

FeSO.. 7H~O 4 2 1 g· MnSO4.4H2O 1 g· CuCl2.2H2O 25 mg CaCl2 100 mg H3BO3 56 mg (NH4)6Mg7O24.4H2O 19 mg ZnSO4.7H2O 200 mg Distilled Water 1000 ml LM Production Medium Without Malt Extract

Dextrose 20 g. Glycerol 20 ml. Ardamine pH 10 g. CoCl2.6H2O 8 mg. Polyglycol p 2000 0.25% Distilled Water 1000 ml. PH 7.0 20 g. 2 0 ml. 10 g. 20 g. 8 mg. 0.25% 1000 ml. 7.0 LM Production Medium Without Modification

Dextrose Glycerol Ardamine pH Malt Extract CoCl2.6H2O Polyglycol p 2000 Distilled Water PH 15

16449Y B. Isolation

The combined whole broth (10.3 liters) was filtered and the mycelia cake was washed with 2.5 liters of deionized water. The combined filtrate and wash was adjusted to pH 4.0 with IN hydrochloric acid. The aqueous solution was extracted with 7 liters of ethyl acetate and the extract was back-extracted with 3x2 liters of aqueous sodium hydroxide solution. The combined sodium hydroxide extract was adjusted to pH 3.8 with IN hydrochloric acid and extracted with 2 liters and 1 liter of ethyl acetate. The combined ethyl acetate solution was dried over anhydrous Na2SO4, filtered and concentrated to dryness. The oily residue was dissolved in toluene and refluxed for 1 hour. The toluene solution was concentrated to dryness and the residue was dissolved in 18 ml of a mixture of n-hexane/toluene/methanol (4/1/1 by volume). This solution was loaded onto a 30 mm (ID) x 40 cm.

Sephadex LH-20 column equilibrated in the same solvent system. After eluting with 300 ml of solvent, a 10 ml fraction was obtained which was concentrated to an oil. High performance liquid p chromatography (HPLC) on an ES Industries Chromega column (9 mm x 50 cm) using a mixture of acetonitrile/water (60/40 by volume) as the eluting solvent yielded 45 mg of dihydrocompactin (Compound III^, R'=H), m.w. 392.2560 by mass spectrum (calculated for C23H36°5' 392.2558).

In KBr, the major IR peaks obtained from a Fourier Transform -IR (FTIR, Nicolet, Model 7199) are at 1724, 1704, 1258, 1078 and 1070 cm 1. Of significance is a peak at 3005 cm-^ and the absence of a peak at 3030 cm'1.

- 16 - 16449Y A nuclear magnetic resonance spectrum was obtained in CDCl^, (zvl mg/0.5 ml) on a Varian SC-300 superconducting nmr spectrometer. The following are the peak positions given in ppm (J- ) relative to internal tetramethylsilane (TMS). 10 15

Assignment 5.62 d,d,d (2.17, 4. 5, 10.0) H3,(or 4') 5.43 d (10) H4,(or 3') 5.20 m Hg ι 4.63 m H6 4.39 m H4 2.75 d,d (17.5, 5. 5) 2.63 d,d,d (17.5, 4. o, 1.5) 3-CH2 2.39 m CH3HCC^° 2.29 m H4a’ + H2· 1.14 d CH3CHC^° 0.90 t CH3CH2 0.84 d 2’-CH, 20 d: doublet; m: multiplet; t: triplet

The evidence indicates the structure to be:

<img img-format="tif" img-content="drawing" file="IL62044AD00023.tif" id="idf0003" />

H* 17

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DESCRIPTION OF THE INVENTION

We have found that the α-methylbutyryl group in Compound IIIa (R'=CH3) and hydro-derivatives, IIIb_e, can be removed cleanly to produce a family of 6(R)—[2-(8-hydroxy-2,6-dimethylpolyhydronaphthyl-l)ethy4(R)-hydroxy-3,4,5,6-tetrahydro-2H-pyran-2-ones which are themselves hypocholesterolemic agents and which are extremely useful as intermediates for the preparation of novel esters which are even more potent in this use.

The preparation of the novel alcohols of this invention is carried out by heating the esters III _ (R* =CH3) with an alkali metal hydroxide such as lithium hydroxide, potassium hydroxide or sodium hydroxide in a protic solvent such as water or alcohols for extended periods. Preferred is lithium hydroxide in water at reflux for about 50-72 hours or under pressure at higher temperatures of 120-180°C for shorter times of 8-24 hours

The pyranone ring readily opens but the removal of the side chain acyl group is not easily effected. The heating must be prolonged and/or pressure must be used.

An inert atmosphere is also helpful. It is quite unexpected that molecules with so many highly sensitive functional centers can withstand the harsh conditions necessary for removal of the highly hindered a-methyl-butyryl ester. It is especially unexpected to find the yields high.

In the case of Compounds III (R'=H) the saponification of the 8'-esters is much more facile proceeding to completion in about 20 hours to give IVa_e (R'=H).

The Compound IV^ (R'=H) is known as ML-236A

Cl as reported by Endo et al. in U.S. Patent 3,983,140. 18

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The products are isolated by acidification and extraction with organic solvents which provides the trihydroxy acid form of compounds ΐν&amp;amp;_θ. These trihydroxy acids can be relactonized by heating a solution of the acid in an appropriate organic solvent such as toluene or benzene in an apparatus permitting continuous separation of the water formed.

The alcohols which form part of this invention comprise Structures IV Ω (R’=CH^) as well as the trihydroxy acids resulting from opening of the lactone rings.

An alternate synthetic route to the Compounds IV, / comprises the steps of hydrolysis of III to IV as described herein followed by hydrogenation of IV under the conditions described previously for the

preparation of III. „ , to produce IV, , IV or IV D f C f 6 JO C θ depending on those reaction conditions. 20 19

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Preparation of Compounds IVa_e

The starting materials, the 8a-hydroxy -compounds IV (R'=CH^) described by Willard (Serial No. 118,04 9) are prepared from the various e^-esters described by Monaghan et al (IIIa, R'=CH3), Albers-Schonberg et al (IHd, R'=CH3) and Patchett et al (IIIb c θ, R’=CH3) by heating them with lithium hydroxide solution for extended periods. The pyranone ring readily opens but the removal of the side chain acyl group is not easily effected. The heating must be prolonged and/or pressure must be used. An inert atmosphere is also helpful.

In the case of the Compounds III (R’=H) the saponification of the 8-esters is much more facile proceeding to completion in about 20 hours.

The 8-hydroxy products are isolated by acidification and extraction with organic solvents which provides the hydroxy acid form, in which the pyranone ring is still opened. These hydroxy acids are relactonized by heating a solution of the acid in an appropriate organic solvent such as benzene or toluene in an apparatus permitting continuous separation of the water formed.

The Compound IV (R*=H) is known as ML-236A as &amp;amp; reported by Endo et al in U.S. Patent 3,983,140.

In their lactone form, these alcohols are the compounds of Formula ΐν&amp;amp;_θ in Table I and are prepared as described in the following preparations. 20

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Preparation of 6(R):- [2-(81 (S)-hydroxy-2' (_S) ,6' (R)-dimethyl-1* ,2* ,6 ' ,7' ,8* ,8 ' a(,R)-hexahydronaphthyl-11 (S))- ethyl]-4(R)-hydroxy-3,4,5,6-tetrahydro-2H-pyran-2-one, iva (r'=ch3) A mixture of 8.0 g. (19.78 mmole) of MK-803 (ma, R'=CH3) and 8.31 g (197.8 mmole) of LiOH.H2O in 600 ml of water was stirred at reflux under a nitrogen atmosphere for 56 hours. The reaction mixture was cooled to 0° and treated, with stirring, with 20 ml of concentrated hydrochloric acid. The mixture was then extracted with three 250-ml portions of ether and the combined extracts were washed successively with three 200-ml portions of water and then 200 ml of saturated brine. After drying over MgSO^, this organic solution was filtered and the solvent evaporated in vacuo to give an oily residue. This residue was dissolved in 200 ml of toluene and heated at reflux under a nitrogen atmosphere for 2 hours with continuous separation of water to effect relactonization. Evaporation of the toluene and trituration of the residue with hexane gave 5.15 g (81%) of the title compound IV (R'=CH-) as a white solid which did not require further purification.

An analytical sample was prepared by recrystallization of a portion of this material from butyl chloride to give white clusters: m.p. 128-131° (vacuum); NMR(CDC13) &amp;amp; θ·θ7 (d,3,J=7Hz, CH3), 1.16 (d,3,J=7Hz, CH3), 2.64 (m,2,pyran C3H’s), 4.27 (brm,l, naphthalene CgH), 4.37 (m,l,pyran C4H) , 4.71 (m,l,pyran CgH) , 5.56 (m,l, naphthalene CgH), 5.79 (dd,l, J=6,1O Hz, naphthalene C3H), 6.03 (d,l,J=10 Hz, naphthalene C4H); IR (CHC13) 3400 (OH), 1725 (C=O), 1240, 1120, 1080cm"1.

Anal. Calcd for ε19Η28θ4.0.lC4HgCl C, 70.67; H, 8.84. Found: C,70.77; H, 8.75. 21

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Alternative preparation of 6 CR) -12-18' (S)-hydroxy-2' (_S) , 6' (R)-dimethy1-1',2' ,6',7',8' r8'a CR)-hexahydron'aphthyl- 1'(S)]ethyl]-4(R)-hydroxy-3,4,5,6-tetrahydro-2H-pyran- 2-one, IV CR'=CH3) 5 A suspension of 188 mg (0.463 mmol) of MK-803

(III , R*=CH-) in 5 ml (5 mmol) of aqueous IN LiOH solution &amp;amp; J is shaken for 12 hours at 135° in a 30 ml stainless steel pressure vessel. The cooled reaction mixture is acidified with IM H3PO4 and extracted with ethyl 10 acetate. The ethyl acetate solution is dried (MgSO4) and filtered and the solvent is evaporated. The residue is dissolved in 20 ml of toluene which is heated to reflux for 4 hours in a Dean-Stark apparatus to effect relactonization. Evaporation of the toluene gives the 15 title compound.

Preparation of alcohols IV&amp;amp; (R’=H) and IVb, IV,, IV^, and IV (R,=H or CH3)

Following essentially either procedure described above but substituting an equivalent amount of esters 20 IIIa(R'=H) or IHb, IIIC, IIId, or ΙΙΙθ (R'=H or CH-j) , for III, (R'=CH-) used therein the corresponding alcohols a. 3 IVa (R'=H), IVb, IV , IVd and ΐνθ (R'=H or CH3) are respectively obtained.

We have found that the 8'-hydroxy compounds 25' of Structure IV can be acylated to give a new class of 8-acyloxy compounds of the structure defined by Formulas I and II and the definitions thereunder. These new compounds are inhibitors of cholesterol synthesis in vivo. 30 22

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The absolute configuration of these compounds is known from X-ray diffraction. Table I provides a convenient tabulation of these structures and their stereochemical relationship. The reference numerals to the various compounds, including those of the various series of polyhydronaphthyl structures, remain the same throughout these specifications and are so used. Each of the esters I (R1=CH,), of this invention contains seven or eight chiral centers. The relative and absolute configuration of these asymmetric centers is as depicted in Table I. More specifically, for ester I (R’=CH,), the Cahn, Ingold, Prelog designations for the absolute configurations are 4(R), 6(R), l'(S), 2'(S), 6'(R), 8'(S) and 8a' (R) [R.S.

Cahn, C.Ingold and V. Prelog, Angew.Chem.Int.Ed.,5, 385 (1966)].

<img img-format="tif" img-content="drawing" file="IL62044AD00024.tif" id="idf0004" />

As is indicated in the formulas I , all of these a—e compounds have the same spatial orientation of groups at each chiral carbon atom and therefore belong to the same stereochemical series. The R-S designation for each center may not be identical to that found for the ester I (R’=CH,) because of the details of the sequence rules used for determining that designation. In the two esters 1^ and I which have an additional chiral carbon atom not present in ester I , the hydrogen atom at 4a' s,

_23 _ 16449Y ' is in the down (or a) orientation as depicted in Table I, giving a trans ring junction.

TABLE I

THE COMPOUNDS OF THIS INVENTION AND THEIR

STEREO-RELATIONSHIP

<img img-format="tif" img-content="drawing" file="IL62044AD00025.tif" id="idf0005" />

<img img-format="tif" img-content="drawing" file="IL62044AD00026.tif" id="idf0006" />

<img img-format="tif" img-content="drawing" file="IL62044AD00027.tif" id="idf0007" />

R’=H or CH3

STEREOCHEMISTRY OF THE HYDRONAPHTHYL SERIES

Series

Double Bonds Present X and Z

R

R

R

R

None

Structure

TO

.TO ,330 xp xp fi 10 24 -

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The 8'-acyloxy compounds of this invention are useful as antihypercholesterolemic agents for the treatment of atherosclerosis, hyperlipemia and like diseases in humans. They may be administered orally or parenterally in the form of a capsule, a tablet, an injectable preparation or the like. It is usually desirable to use the oral route. Doses may be varied, depending on the age, severity, body weight and other conditions of human patients but daily dosage for adults is within a range of from about 2 mg to 2000 mg (preferably 10 to 100 mg) given in three or four divided doses. Higher doses may be favorably applied as required.

The compounds of this invention also have useful anti-fungal activities. For example, they may be used to control strains of Penicillium sp.,

Aspergillus niger, Cladosporium sp., Cochliobolus miyabeanus and Helminthosporium cynodnotis. For those utilities they are admixed with suitable formulating agents, powders, emulsifying agents or solvents such as aqueous ethanol and sprayed or dusted on the plants to be protected.

The preparation of the compounds of this invention is described in Flow Sheet A. 25

16449Y

FLOW SHEET A

<img img-format="tif" img-content="drawing" file="IL62044AD00028.tif" id="idf0008" />

VI a-e &lt;5&gt;

II a-e a-e

Definitions

Reactions 1) 2) 3) X,Y,Z,R and R' as defined in specification and series a-e as defined in Table I.

Lithium hydroxide, heat, acidify and lactonize t-Butyldimethylchlorsilane and imidazole in DMF at ambient temperatures in an inert atmosphere.

Treatment with RCOCl and 4-dimethylamino-pyridine in pyridine solution preferably under inert atmosphere.

Treatment with RCOOH and N,Ν’-dicyclohexyl-carbodiimide and 4-pyrrolidinopyridine in dichloromethane, preferably under an inert atmosphere. 4) 26

16449Y 5) Three equivalents of tetrabutylammonium fluoride and four equivalents of acetic acid per equivalent of ester in THF, preferably in an inert atmosphere. 6) Aqueous alkali followed by careful acidification with dilute acid. 7) See Reactions and Reagents and Flow Sheet for synthesis of III, ~ O f C f c

In the novel process of this invention the 4-hydroxyl on the pyranone ring, of alcohols IVa_e is first protected with a t-butyldimethylsilyl group by reaction with t-butyldimethylchlorosilane in an inert atmosphere at ambient temperatures in the presence of an acid acceptor such as imidazole to provide the protected alcohols V . The 8-hydroxyl on the polyhydronaphthyl ring is then acylated in one of two ways. The first comprises treatment with the acid chloride of the desired acyl group in pyridine in the presence of 4-dimethylaminopyridine as a catalyst. The second comprises treatment of the 8'-polyhydronaphthol with the free acid of the desired acyl group and a carbodiimide such as N,N*-dicyclohexylcarbodiimide with 4-pyrrolidinopyridine as a catalyst in dichloromethane. These procedures give the protected esters VIa_e· The removal of the silyl protecting group from the 4-hydroxyl of the pyranone ring is then carried out, using three equivalents of tetrabutylammonium fluoride and four equivalents of acetic acid per equivalent of esters VIa_e, to give the desired compounds 1&amp;amp;_θ. The ratio of reagents in this last reaction is critical to the yield of the process and the purity of the products.

The acyl groups thus put on the 8’-hydroxyl are those in which R in I is; a-e - 27 - 62044/2

It is preferred that R' be CH^ and it is especially preferred that none of X, Y or Z is a double bond.

Compounds I can be hydrolyzed with bases such as NaOH to yield the salts such as the sodium salt of Compounds II . The use of bases with other pharmaceutically acceptable cations affords salts of those cations. Careful acidification of the salts affords the hydroxy acids Ha_e which revert to Compounds Ia_e at acidic pH. Treating Compound I under acidic or basic catalysis with methanol, ethanol, propanol, or butanol or with phenyl-, dimethylamino-, or acetylamino-alkanols 28

16449Y yields the corresponding esters of Compounds IIa_e which also form a part of this invention.

The pharmaceutically acceptable salts of this invention include those formed from cations such as sodium, potassium, aluminum, calcium, lithium, magnesium, zinc and tetramethylammonium as well as those salts formed from amines such as ammonia, ethylenediamine, N-methylglucamine, lysine, arginine, ornithine, choline, N,N’-dibenzylethylenediamine, chloroprocaine, diethanolamine, procaine, N-benzylphenethylamine, l-p-chlorobenzyl-2-pyrrolidine-l'-yl-methylbenzimidazole, diethylamine, piperazine, and tris(hydroxymethyl)aminomethane. EXAMPLE 1 6(R)- [2-(8'(S)-2",2"-dimethylpropanoyloxy-2'(S),6'(R)-dimethyl-1*,2’,6',7’,8',8'a(R)-hexahydronaphthyl-1'(S))-ethyl]-4(R)-hydroxy-3,4,5,6-tetrahydro-2H-pyran-2-one

Step A: Preparation of 6(R)-[2-(8’ (S)-hydroxy-21 (S)-6*(R)-dimethyl-1',2',6',7',8', 8 1 a (R)-hexahydronaphthyl-1' (S)) ethyl]-4(R)-(dimethyl-tert-butylsilyloxy)- 3,4,5,6-tetrahydro-2H-pyran-2-one, V (R'=CH-) _____ cl j A mixture of the alcohol IV (R'=CH-) (18.3 g, 57.1 mmol), 21.5 g (142.8 mmol) of tert-butyldimethylchloro-silane and 19.4 g (285.6 mmol) of imidazole in 200 ml of Ν,Ν-dimethylformamide was stirred at 20° under a nitrogen atmosphere for 18 hours. The reaction mixture was then diluted with 1500 ml of ether and washed successively with water, 2% aqueous hydrochloric acid, water and saturated sodium bicarbonate. The ether solution was dried over MgSO4, filtered and reduced to a volume of 1 L. After addition of 600 ml of hexane, the volume was reduced to 600 ml on a steam bath. The product crystallized at room temperature; after isolation and air drying this provided 13.7 g of a white cottony solid. The mother liquors were reduced to 250 ml and a second crop of crystals was 29

16449Y isolated after this solution stood at 0° overnight.

The combined yield was 17.13 g (69%) of the title compound as a white cottony solid: mp 142-1440(vac); NMR (CDC13) £ °·1θ (s,6,(CH3)2Si),0.90 (s , 9, (CH3) gCSi) , 1.19 (d,3,J=7Hz, CH3), 2.58 (d, 2,J=4Hz,pyran C3H's), 4.3 (m,2,pyran C^H and naphthalene CgH) 4.70 (m, 1, pyran CgH), 5.57 (m,1,naphthalene CgH), 5.58 (dd,l, J=6,10Hz, naphthalene CgH),6.03 (d,l, J=10Hz, naphthalene C^H).

Anal. Calcd. for C25H42°4Si: c/ 69.08, H, 9.74. Found: C, 69.46; H, 9.83.

Step B: Preparation of 6 (R) — [2- (8* (S)-2" ,2’’-dimethyl- propanoyloxy-2'(S),6'(R)-dimethyl-1',2',6',7’, 8',8 ' a(R)-hexahydronaphthyl-1’(S)) ethyl]-4(R)- (dimethyl-tert-butylsilyloxy)-3,4,5,6-tetrahydro 2H-pyran-2-one, VI (R’=CHO) a j A solution of 6.0 g (13.8 mmol) of the alcohol

Va (Rl=CH3) from Step A and 200 mg of 4-dimethylamino-pyridine in 50 ml of pyridine was cooled to 0° under a nitrogen atmosphere. To this stirred solution was added 6.8 ml (6.65 g, 55.2 mmol) of pivaloyl chloride over 15 minutes. The reaction mixture was stirred at 0° for 1 hour and then at 20° for 4 days. The reaction mixture was diluted with 750 ml of ether and washed with 2% aqueous hydrochloric acid until the wash was acidic and then with saturated NaHCOg solution. After drying over MgSO^ the solution was filtered and evaporated to give 7.81 g of the title compound as a light orange oil: NMR (CDClg) ξ 0.09 (s,6(CHg)2Si), 0.88 (s,9,(CHg)3CSi), 1.28 (s,9, (CHg)gCCOg-), 2.57 (d,2,J=4Hz pyran CgH's), 4.32 (m, 1, pyran C4H) , 4.63 (m,l, pyran CgH), 5.34 (m, 1, naphthalene CgH), 5.54 (m,l, naphthalene CgH), 5.78 (dd, 1, J=6, 10Hz, naphthalene CgH), 6.03 (d,l,J=10Hz, naphthalene C4H). - 30 - 62044/2

Step C: Preparation of 6(.R)-[2-(8,(S)-2",2"-dimethyl- propanoyloxy-2'(S),6’(R)-dimethyl-1’,2',6',7’,8', 8'a(R)-hexahydronaphthyl-1’(S)) ethyl]-4(R)-hydroxy-3,4, 5,6-tetrahydro-2H-pyran-2-one, I_ (R* =CH-,)

Cl □

To a solution of 10.0 g (31.7 mmol) of

Bu4N+F .3h2O and 2.4 ml (2.5 g, 42.3 mmol) of acetic acid in 50 ml of tetrahydrofuran was added 7.81 g (13.8 mmol) of the silyl ether VI, (R'=CH^) from Step B in 50 ml tetra-hydrofuran. This mixture was stirred at 20° under a nitrogen atmosphere for 18 hours. The reaction mixture was diluted with 700 ml of ether and washed successively with 2% aqueous hydrochloric acid, water and saturated aqueous NaHCOy The organic solution was dried (MgSO^) and filtered. Evaporation of the solvent left 6.45 g of an off-white solid. This material was crystallized from 100 ml of butyl chloride and the isolated crystals were dried at 35°/0.01 mm for four hours to give 4.0 g (72%) of the title compound as nearly white needles: mp 167.5-170.5° (vac); NMR (CDC13) £ 0.88 (d,3,J=7Hz,CH3), 1.08 (d,3,J=7Hz, CH3), 1.19 (s,9,(CH3)3C), 2.67 (d,2,J=4Hz, pyran C-jH’s), 4.39 (m,l,pyran C4H) , 4.65 (m,l,pyran CgH) , 5.36 (m,l, naphthalene CgH) 5.55 (m,1,naphthalene CgH), 5.80 (dd,l,J=6,10Hz, naphthalene C3H), 6.04 (d,l,J=10Hz, naphthalene C4H); HPLC (4.6 mm. x 25 cm Partisil 10 PAC, 10% isopropanol/hexane, 4 ml/min) retention time 4.4 min.

Anal. Calcd. for C24H36°5: C,71.25; H, 8.97. Found: C,71.40; H, 8.93. 62044/2 - 31 -

Employing the procedures of Example 1, Step A, followed by Example 1, Steps B and C, but substituting for the diol of structure IV (R'=CH_.) in Example 1, cl □

Step A, the corresponding diol of structure IV_ (R'=H) cl or IV, . or (R'=H, or CH-) , there are produced D f C f Cl / θ in sequence the silyl ethers of structures Va (R'=H) or VI, ,, and (R'=H, or CH-,) , and the novel O f C | Cl © «5 esters of structures I (R’=H) or I, -, and cl D ι C f Cl Θ

(R'=H or CH^) in accordance with Flow Sheet A EXAMPLE 2

Typical formulations for filling a size 0 hard gelatin capsule comprise 3.125, 6.25, 12.5, 25 or 50 mg of one of the novel compounds of this invention such as the product of Example 1, Step C, and sufficient finely divided lactose to provide a total capsule content of about 580-580 mg.