Substituted glutaric acid lactones in the treatment of hyperlipidemia

Compounds of formula I ##STR1## are described which are useful to inhibit the formation of serum cholesterol by virtue of their ability to inhibit .beta.-hydroxy-.beta.-methylglutaryl-CoA(HMG CoA), the rate-controlling substance in the synthesis of serum cholesterol.

BACKGROUND OF THE INVENTION 
The invention relates to novel substituted glutaric acid lactone 
derivatives of formula I which are useful to inhibit the formation of 
serum lipids, and especially cholesterol. The novel compounds exhibit this 
utility by virtue of their ability to inhibit the activity of 
.beta.-hydroxy-.beta.-methyl-glutaryl coenzyme A (HMG CoA reductase) and 
thus inhibit the formation of serum cholesterol. HMG CoA is a substance 
which controls the rate at which cholesterol is synthesized in mammalian 
liver (one of the two principal in vivo sources of serum cholesterol). 
Thus the compounds of the present invention are useful to inhibit sterol 
biosynthesis in individuals predisposed to familial type 
hypercholesterolemia and hyperlipoproteinemia. The significance of such 
compounds is widely recognized, e.g. Breslow et al. Biochim. Biophys. 
Acta, 398, 10 (1975); Betheridge et al., Brit. Med. J., 4,500 (1975); 
Brown et al., Scientific American, 58 Nov. (1984). 
PRIOR ART 
The use of agents which lower serum cholesterol is widely described in the 
art as described above. 3-hydroxy-3-substituted glutaric acid derivatives 
are described in U.S. Pat. No. 3,818,080 as being useful for their 
antiulcerogenic activity. 
Pentanedioic acid derivatives are described as antihyperlipidemic agents in 
commonly assigned co-pending application Ser. No. 577,411 filed Feb. 6, 
1984, now abandoned. 3-Substituted pentanedioic hemiesters and anhydrides 
are described as elastase inhibitors in commonly assigned co-pending 
application Ser. No. 569,007, filed Jan. 9, 1984, now U.S. Pat. No. 
4,554,359. In contrast, the compounds of the present invention are 
glutaric acid lactone derivatives which have not been heretofore suggested 
for lowering serum cholesterol and which are structurally unrelated to the 
aforementioned prior derivatives by reason of their cyclic (i.e., lactone) 
structures. The compounds of the invention are prepared from intermediates 
having a hydroxy group .delta. to the carboxylic acid moieties which 
intermediates have not been described previously. 
SUMMARY OF THE INVENTION 
The present invention particularly provides compounds of formula I 
##STR2## 
wherein 
R represents lower alkyl, cycloalkyl aryl, or a group of the formula 
##STR3## 
wherein R.sup.1 represents cycloalkyl or phenyl; 
m is an integer from 8 to 15, inclusive; and the pharmaceutically 
acceptable salts or esters thereof. 
DETAILED DESCRIPTION OF THE INVENTION 
As used herein, the expression "lower alkyl" includes straight or branched 
chain alkyl of 1 to 6 carbon atoms. Exemplary of suitable lower alkyl 
groups are methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, n-hexyl 
and the like. 
Examples of suitable "cycloalkyl" groups are groups of 3 to 7 carbon atoms, 
including, for instance, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 
cycloheptyl. Especially preferred in the practice of the present invention 
is cyclohexyl. 
Suitable "aryl" groups are those of 6 to 10 carbon atoms, including phenyl 
and naphthyl with phenyl being especially preferred. 
Appropriate pharmaceutically acceptable "salts" include Na+, K+, Ca++, 
NH.sub.4 + and any other cation capable of reacting with the carboxylic 
acid moiety provided same does not adversely affect the pharmacological 
properties of the resulting compounds. Likewise, appropriate lower alkyl 
esters are encompassed within the scope of formula I. 
Especially preferred compounds of formula I are those wherein R is a group 
selected from 
##STR4## 
and m is an integer between 8 to 13, inclusive. 
Particularly preferred compounds in accordance with the present invention 
are of the formulae: 
##STR5## 
It will be appreciated by those skilled in the art that the compounds of 
the present invention contain asymmetric carbon atoms, (i.e., centers of 
chirality) and, therefore, the compounds depicted may exist as individual 
diastereomers or mixtures thereof and such diastereomers and mixtures are 
included within the scope of the defined structural formulas herein. 
The utility of the instant compounds and their inhibition of the formation 
of serum cholesterol can be demonstrated via the following standarized 
test procedures: 
The in vitro inhibitory activity of the present compounds is evaluated 
using rat liver microsomal HMGCoA reductase as described by Edwards, et 
al. J. Biol. Chem. 249, 2891 (1974). The IC.sub.50 is defined as the 
concentration required to inhibit the enzyme by 50% of control. 
HMG derivaties are added to a preincubation mixture consisting of 0.1 M K 
PO.sub.4, PH7.2, 0.02 M glucose-6-PO.sub.4, 2.5 mM NADP, 0.7 units of 
glucose-6-PO.sub.4 dehydrogenase, 5 mM dithiothreitol, 50 mM mevalonic 
acid and approximately 50 .mu.g of microsomal protein. Triplicate samples 
are preincubated for 15 min. at 37.degree. C. in a volume of 1 ml. 
Incubation is started with 40 .mu.M 14C-HMGCoA (0.1 ml) run for 15 min. at 
37.degree. C. and stopped with 5 N HCl (0.1 ml). Assay tubes are allowed 
to set for at least 30 min., then approximately 50,000 dpm of .sup.3 
H-mevalonic acid are added to provide for extraction efficiency. Mevalonic 
acid is extracted with ether and the % .sup.14 C-HMGCoA incorporation 
determined for concurrent control and test reaction systems. Testing is 
done (first) with a range finding assay followed by a 4 or 5 point assay 
to find the IC.sub.50 value. Coefficient of variation ranged from 5 to 20% 
with an average value of 14%. Compactin in this test had an IC.sub.50 
value of 1 .mu.M under these conditions. 
In vivo activity is tested as follows: 
Initial serum total cholesterol, triglycerides, and lipoprotein cholesterol 
values are determined 3 times for each male Rhesus monkey used before 
treatment with a test compound begins. The test compound is administered 
in an initial dose of 60 mg/kg for 2 weeks and blood samples are taken to 
determine if rebound occurs. A dose is rated active if the 14-day mean 
values are significantly reduced from the pretreatment values (p 0.05). 
Statistical comparisons are made using the two tailed student's t test. 
By virtue of their activity in the foregoing tests, the compounds of 
formula I are useful in treating type 2 hypercholesterolemia (TTH-2) in 
humans and animals. A physician or veterinarian of ordinary skill could 
readily determine a subject who has TTH-2 symptoms. Regardless of the 
route of administration selected, the compounds of the present invention 
are formulated into pharmaceutically acceptable dosage forms by 
conventional methods known to the pharmaceutical art. 
The compounds can be administered in such oral unit dosage forms as 
tablets, capsules, pills, powders, or granules. They also may be 
administered rectally, intraparenterally, subcutaneously, or 
intramuscularly, using forms known to the pharmaceutical art. In general, 
the preferred form of administration is orally. 
An effective but non-toxic quantity of the compound is employed in 
treatment. The dosage regimen for preventing or treating TTH-2 or other 
hyperlipidemic conditions by compounds of this invention is selected in 
accordance with a variety of factors including the type, age, weight, sex, 
and medical condition of the mammal, the severity of the condition, the 
route of administration and the particular compound employed. An 
ordinarily skilled physician or veterinarian will readily determine and 
prescribe the effective amount of the active agent to prevent or arrest 
the progress of the condition. In so proceeding, the physician or 
veterinarian could employ relatively low dosages at first, subsequently 
increasing the dose until a maximum response is obtained. 
Initial dosages of the compounds of the invention are ordinarily in the 
area of 10 mg/kg up to 200 mg/kg orally. When other forms of 
administration are employed equivalent or adjusted doses are administered 
depending on the route of administration. 
The general procedure for producing the compounds of the present invention 
is outlined in the following reaction scheme, which is similar to the 
general procedures used to produce the compounds of U.S. Pat. No. 
3,818,080 which is incorporated herein by reference. 
##STR6## 
The hydroxyacid esters 3 and the 4-allyl derivatives 4 in the above 
reaction scheme are novel intermediates for the synthesis of the active 
lactones of the present invention of formula I. In the above formulas, R' 
is a suitable protecting group selected in accordance with conventional 
practices in the art. Alkylsilyl groups, e.g. trimethylsilyl, are 
preferred in the present invention. 
The starting esters 1 as well as the aldehydes 2 may be readily synthesized 
in accordance with methods known in the art or may be purchased from 
available sources for conversion to the novel intermediates 3. 
The compounds of the present invention by reason of their serum lipid (e.g. 
cholesterol) lowering properties are useful alone or in the pharmaceutical 
compositions and methods of the invention as antihyperlipidemic agents. 
Ultimately the compounds of the invention find applications in the 
treatment, prevention or mitigation of atherosclerosis, arteriosclerosis, 
myocardial infarction, hypertension, and related conditions in which 
elevated serum lipid/cholesterol levels are a causative component.

The following non-limiting examples further illustrate details for the 
preparation of compounds of the present invention. The invention as a 
whole is not to be construed or limited either in spirit or in scope by 
the following examples. Those skilled in the art will readily understand 
that variations in the conditions and processes exemplified in the 
following preparative procedures can be utilized to prepare these 
compounds. All temperatures are degrees Celcius unless otherwise 
specified. Chemical shifts for NMR spectra are reported in parts per 
million (.delta.). Splitting patterns are designated as s, singlet; d, 
doublet; t, triplet; q, quartet, and m, multiplet. Elemental analyses were 
performed by microanalytical procedures. 
DESCRIPTION OF THE PREFERRED EMBODIMENTS 
Example 1 
3-Carboxymethyl-3,5-Dihydroxyoctadecanoic Acid-1,5-Lactone 
##STR7## 
(A) 3-Hydroxyhexadecanoic acid Ethylester, (3, R=CH.sub.3, m=12) To a 
freshly prepared solution of 250 ml of dry THF containing 4.41 g (0.05 
mole) of lithioethyl acetate cooled to -78.degree., standing for one hour, 
was added, over ten minutes, a solution of 10 g (0.047 mole) of 
tetradecanal (2, R=CH.sub.3, m=12) in 90 ml of THF. Then 15 ml of a 20% 
aqueous solution of HCl was added to the mixture, followed by water and 
ether. The organic layer was separated, washed with aqueous sodium 
bicarbonate solution, water, dried over MgSO.sub.4, and distilled to 
dryness at reduced pressure to yield 9.4 g of product. The crude product, 
when purified by chromatography on silica gel utilizing ethyl 
acetate-hexane (15:85) as an eluant, yielded 3.7 g of pure 
3-hydroxyhexadecanoic acid ethylester; .delta. (CDCl.sub.3) 
1.28(24CH.sub.2), 3.98-4.33 (quartet,OCH.sub.2 --). 
(B) 4-allyl-4,6-dihydroxynonadec-1-ene (4,R=CH.sub.3, R'=H m=12) To a small 
amount of freshly prepared allyl magnesium bromide and 1.7 g of Mg (0.07 
mole) covered with THF was added a solution of 3.7 g of 
3-hydroxyhexadecanoic acid ethyl ester (0.0123 mole) and 6.05 g (0.05 
mole) of allyl bromide in 100 ml of THF. The addition proceeded so as to 
maintain a reflux temperature. The mixture was heated at refluxing 
temperature for 1 hr., cooled, and poured into saturated aq. ammonium 
chloride solution. The mixture was extracted with ether and the ether 
solution was dried over MgSO.sub.4 and distilled to dryness. The crude 
product weighed 5.0 g and could be used without further purification. The 
product could be purified by column chromatography on silica gel using 
ethyl acetate-hexane (15:85) as the eluant; .delta. (CDCl.sub.3), 
1.28(CH.sub.2), 4.90-5.25(m, =CH.sub.2) and 5.4-6.2(m,=CH). 
(C) 3-Carboxymethyl-3,5-dihydroxyoctadecanoic Acid 1,5-Lactone (5, 
R=CH.sub.3, m=12). To a solution of 7.7 g (0.023 mole) of 
4-allyl-4,6-dihydroxynonadec-1-ene, (4, R=CH.sub.3, R'=H, m=12) and 2.32 g 
(0.034 mole) of imidazole in 100 ml. of methylene chloride, cooled to 
0.degree., was added, dropwise with stirring, 3.69 g (0.34 mole) of 
trimethylsilylchloride. A white precipitate formed. The mixture was 
allowed to warm to 20.degree. and 100 ml. of water was added. The 
methylene chloride solution was separated, dried over MgSO.sub.4, and 
distilled to dryness to yield 8.7 g of the crude trimethylsilyloxy 
derivative (4, R=CH.sub.3, R'=Si(CH.sub.3).sub.3, m=12) which was used 
without further purification. 
A solution of 14.0 g of the trimethylsilyloxydiene (4, R=CH.sub.3, 
R'=Si(CH.sub.3).sub.3, m=12) in 600 ml of ethylacetate/methylene chloride 
(1:1) was cooled to -25.degree.. Ozone was passed into the solution until 
it turned blue, and then the excess ozone was purged with oxygen. This 
cold solution was then added dropwise, with stirring, to 100 ml of glacial 
acetic acid. This solution was concentrated to about 1/3-1/2 of its 
original volume and heated to reflux on a steam bath. A solution of 30 ml 
water, 80 ml glacial acetic acid, and 32 ml of 30% hydrogen peroxide, was 
added dropwise to the refluxing solution. The resulting reaction mixture 
was then refluxed for 2 hours and poured onto 400 ml of ice. 
The aqueous mixture was extracted into ether, which was washed with aqueous 
sodium bisulfite, dried over MgSO.sub.4 and distilled to dryness under 
reduced pressure. Trituration of the crude product with hexane yielded a 
crystalline material which on crystallization from methylene chloride and 
hexane gave 5 (R=CH.sub.3, m=12) R.sub.F =0.2 [Merck silica gel and 
elution with ethyl acetate-hexane (1:1), containing a trace of acetic 
acid]; mass spectral analysis, molecular ion=356. 
Anal. Calcd. for C.sub.20 H.sub.36 O.sub.5 C: 67.41; H: 10.11; Found C: 
66.99; H: 10.16. 
.sup.13 CMR spectrum exhibited maxima at .delta. 14.1 (CH.sub.3); 22.7, 
24.9, 29.7, 32.0, and 39.4 [(CH.sub.2).sub.12 ]; 39.9 and 42.2 (CH.sub.2 
in lactone), 45.0 (CH.sub.2 attached to carboxylic acid), 68.4 (C--OH), 
77.2 (C--O), 172.0 (lactone C.dbd.O), and 174.9 (COOH). 
Example 2 
3-Carboxymethyl-3,5-Dihydroxy-14-Phenylpentadecananoic Acid-1,5-Lactone 
##STR8## 
(A) 10-Phenylundecanal 
##STR9## 
To a solution of 100 g (0.362 mole) of 11-phenylundecanoic acid methyl 
ester in 250 ml of toluene cooled to -78.degree. C. was added dropwise 
with stirring 67 g (405 ml solution of toluene) of diisobutylaluminum 
hydride. The reaction mixture was stirred at -78.degree. for 11/2 hours 
and then quenched by adding 50 ml of water dropwise slowly and carefully, 
so as to prevent foaming. Then a saturated solution of 1.3 liters sodium 
potassium tartrate was added with stirring and the mixture was extracted 
with ether. The ether solution was separated, washed with water, dried 
over MgSO.sub.4 and distilled to dryness to yield 78.5 g of the aldehyde 
as an oil. The compound exhibited NMR maxima at .delta.(CDCl.sub.3) 1.28 
(aliphatic H's), 2.,31 (-CH.sub.2 adjacent to aldehyde), 7.18 (m, phenyl), 
and 9.73 (CHO). 
(B) 3-Hydroxy-12-phenyltridecanoic acid Ethylester 
##STR10## 
When 78 g (0.317 mole) of 10-phenylundecanal was treated with lithioethyl 
acetate according to the procedure for the preparation of 3(R=CH.sub.3, 
m=12), 73.8 g (70% yield) of 3-hydroxy-12-phenyltridecanoic acid ethyl 
ester was obtained; .delta. (CDCl.sub.3) 1.28 (m, CH.sub.2), 2.1-2.6 (m, 
phenyl CH, CH.sub.2 CO), 3.98-4.33 (CHOH, OCH.sub.2) and 7.20-7.22 (m, 
phenyl). This compound could be used without further purification. 
(C) 4-Allyl-4,6-dihydroxy-15-phenylhexadec-1-ene 
##STR11## 
When 73 g (0.219 mole) of 3-hydroxy-13-phenyltridecanoic acid ethyl ester, 
102.8 g (0.85 mole) of allyl bromide and 24.32 g (1.0 mole) of Mg was 
reacted as in the procedure for the preparation of 
4-allyl-4,6-dihydroxyheptadec-1-ene, 74 g (90%) of product was obtained; 
.delta. (CDCl.sub.3), 1.28 (m, CH.sub.2), 2.15-2.60 (phenyl CH, CH.sub.2, 
CH.dbd.), 4.90-6.2 (m, CH.dbd.CH.sub.2) and 7.22 (m, phenyl). 
(D) 3-Carboxymethyl-3,5-dihydroxy-14-phenylpentadecanoic acid 1,5-Lactone 
##STR12## 
When 4.8 g (0.0129 mole) of 4-allyl-4,6-dihydroxy-15-phenylhexadec-1-ene 
was substituted for 4-allyl-4,6-dihydroxynonadec-1-ene in the procedure 
for the preparation of 5 (R=CH.sub.3, m=12) 200 mg of 5 
##STR13## 
was obtained: .delta. (CDCl.sub.3): 0.65-0.95 (m, CH.sub.3), 1.28 (m, 
CH.sub.2), 2.4-2.9 (m,2CH.sub.2 CO), 4.62 (m, CHOC), 7.2 (m, C.sub.6 
H.sub.5). 
Anal. Calcd for C.sub.23 H.sub.34 O.sub.5. 1/3H.sub.2 O C: 69.75; H: 
8.73. Found C: 69.81; H, 9.03. 
Example 3 
3-Carboxymethyl-14-Cyclohexyl-3,5-dihydroxy pentadecanoic acid-1.5-Lactone 
##STR14## 
(A) 3-Hydroxy-12-cyclohexyltridecanoic acid Ethylester. 3, 
##STR15## 
Sixty grams of 3-hydroxy-13-phenyltridecanoic acid ethyl ester in methanol 
solution was hydrogenated in the presence of 12 g of 5% Rhodium on carbon 
catalyst at 60 lbs. of hydrogen 1 in.sup.2 at 60.degree. for 25 hours. The 
methanol solution was distilled to dryness and the product was purified by 
chromatography on silica, utilizing ethylacetate-hexane (15:85) as an 
eluant, to yield 16.5 g of the hydroxyester; as a liquid; .delta. 
(CDCl.sub.3) maxima centered at about 0.85 (m, CH.sub.3), 1.28 and 1.64 
(m, CH.sub.2, C.sub.6 H.sub.11) and 2.3 (m, CH.sub.2 CO). 
(B) 4-Allyl-4,6-dihydroxy-15-cyclohexylhexadec-1-ene. 
##STR16## 
When 16.5 g (0.049 mole) of 3-hydroxy-12-cyclohexyltridecanoic acid 
ethylester, 20.6 g (0.17 mole) of allyl bromide and 24.3 g (0.20 mole) of 
magnesium was reacted as described for the preparation of 
4-allyl-4,6-dihydroxyheptadec-1-ene, 14.9 g of a yellow oil was isolated 
in 80% yield which was used without purification; .delta. (CDCl.sub.3) 
1.28(m,CH.sub.2) and 4.90-6.2(m,CH.dbd.CH.sub.2). 
(C) 3-Carboxymethyl-14-cyclohexyl-3,5-dihydroxypentadecanoic acid 
1,5-lactone 
##STR17## 
When 14.9 g (0.0394 mole) of 
4-allyl-4,6-dihydroxy-15-cyclohexylhexadec-1-ene was substituted for 
4-allyl-4,6-dihydroxynonadec-1-ene in the procedure for the preparation of 
5 (R=CH.sub.3, m=12) a yellow oil was isolated. The crude material was 
purified by chromatography on silica gel to yield 
3-carboxymethyl-14-cyclohexyl-3, 5-dihydroxypentadecanoic acid; .delta. 
(CDCl.sub.3) 0.6-1.8 (broad m, CH.sub.3, CH.sub.2, C.sub.6 H.sub.11), 
2.5-2.8(m,2CH.sub.2 CO) and 4.4-4.95(m,CH--O). 
Anal. Calcd for C.sub.23 H.sub.40 O.sub.5.1/3H.sub.2 O C: 68.71 H: 10.11 ; 
Found: C: 68.82 H: 10.10. 
The following data illustrate the HMG CoA reductase activity (i.e. 
antihypercholesterolemia or antihyperlipidemia) of the compounds of the 
present invention as determined in the previously identified standard 
laboratory test: compound of Example 1=52% inhibition at 
5.0.times.10.sup.-5 M; compound of Example 2=57% inhibition at 
5.0.times.10.sup.-5 M; compound of Example 3=49% inhibition at 
7.5.times.10.sup.-5 M. 
While the invention has been described and illustrated with reference to 
certain preferred embodiments thereof, those skilled in the art will 
appreciate that various changes, modifications, and substitutions can be 
made therein without departing from the spirit of the invention. For 
example, effective dosages other than the preferred ranges set forth 
hereinabove may be applicable as a consequence of variations in the 
responsiveness of the mammal treated, severity of condition treated, 
dosage related adverse effects, if any, observed and analogous 
considerations. Likewise, the specific pharmacological responses observed 
may vary depending upon the particular active compounds selected or 
whether different active compounds are used in combination or in the 
presence of suitable pharmaceutical carriers, as well as the type of 
formulation and mode of administration employed, and such expected 
variations or differences in results are contemplated in accordance with 
the objects and practices of the present invention. It is intended, 
therefore, that the invention be limited only by the scope of the claims 
which follow.