Note: Descriptions are shown in the official language in which they were submitted.
AHP-9148/9148-1-C1
- ~3310~0
- 1 - ,,
NAPHTHALENEPROPIONIC ACID DERIVATIVES
AS AllTl-rNFLAMMATORY/ANTlALLERGlC AGI~NTS
' , ~
This invention relates to novel naphthalenepropionic acid deriva-
tives possessing lipoxygenase inhibitory and leukotriene antagonist activity,
which are useful as anti-inflammatory, antiallergic and cytoprotective agents.
It is known that arachidonic acid (AA) is metabolized in mammals
by two distinct pathways. The metabolism of arachidonic acid by cyclooxy-
genase enzymes results in the production of prostaglandins and thromboxanes.
The physiological activity of the prostaglandins has already been amply
elucidated in recent years. The other pathway of AA metabolism involves
lipoxygenase enzymes and results in the production of a number of oxidative
products called leukotrienes. The latter are designated by the LT nomencla-
ture system, and the most significant products of the lipoxygenase metabolic
¦ pathway are the leukotrienes B4, C4, D4 and E4. The substance denominated
slow-reacting substance of anaphylaxis (SRS-A) has been shown to consist of a
mixture of sulfidopeptide leukotrienes, C4, D4 and E4 [see Bach et al., J.
, Immun., 215, 115-118 (1980); Biochem. Biophys. Res. Commun., 93, 1121-1126
(1980)].
The significance of these leukotrienes is that a great deal of
evidence has been accumulated showing that leukotrienes participate in
. inflammatory reactions, exhibit chemotactic activities, stimulate lysosomal
enzyme release and act as important factors in the immediate hypersensitivity
reaction. It has been shown that LTC4 and LTD4 are potent broncho-
constrictors of the human bronchi [see Dahlen et al., Nature 288, 484-486
(1980) and Piper, Int. Arch. Appl. Immunol., q6, suppl. 1, 43 (1985)] which
stimulate the release of mucus from airways in vitro [Marom et al., Am. Rev.
Resp. Dis., 126, 449 (1982)], are potent vasodilators in skin [see Bisgaard et al,
Prostaglandins, 23, 797 (1982)~, and produce a wheal and nare response tCamp
et al., Br. J. Pharmacol" 80, 497 (1983)]. The nonpeptide leukotriene, LTB4,
is a powerful chemotactic factor for leukocytes [see A.W. Ford-Hutchinson, J.
Roy. Soc. Med., 74, 831-833 (1981)1, which stimulates cell accumulation and
affects vascular smooth muscle [see Bray, Br. Med. Bull., 39, 249 11983)]. The
activity of leukotrienes as mediators of inflammation and hypersensitivity is
~ : : .. :-
,: .:'~ ~
~s ~ ` 5;~
~331~0~
-2-
extensively reviewed in Bailey and Casey, Ann. Reports Med. Chem., 17, 203-
217 (1982) and in Bray, Agents snd Actions, 19, 87 (1986).
There is slso evidence thst products of the cyclooxygenase/lipoxy-
gensse pathways play key roles in both the pathogenesis of gsstric mucossl
S dQmage due to extrscellular (gastric and intestinal contents, microorganisms, -
and the like) or intracellulsr (ischemia, viruses, etc.) agents, ss well ss in
cytoprotection agsinst such dsmage. Thus, on the one hand prostaglandins
exert a cytoprotective effect on the gsstric mucosa ~see Robert, Gastroen-
terology, 77, 761-767 (1979)1 snd this action of the prostsglandins, especislly
of the E series, is considered to be of importsnce in the treatment of
gsstrointestinal ulcerstion [see Isselbacher, Drugs, 33 ~;uppl.), 38-46 (1987)].On the other hand, ex vivo experiments have shown that gsstric mucosal tissue ;-
from ethanol-pretrested rats is capable of LTC4 generation and that this ~ ~ -
LTC4 production is guantitstively relHted to the severity of the ethanol -
dsmsge [see Lange et al., Nsunyn Schmiedeber~s Arch. Pharmscol. Suppl.,
330, R27, (1985) 1. It hss also been demonstrsted thQt LTC4 csn induce V8SO~
constriction in both venous snd srteriolsr vessels in the rst submucoss tsee -~
Whittle, IUPHAR Ninth Int. Con~. of Pharm., S30-2, London, England (1984)].
This is significant since ethsnol-induced lesion formstion in gastric mucosa ; -
20 msy be multifactorisl with, for example, stssis of gsstric blood nOw contri~
buting significsntly to the development of the hemorrhagic necrotic aspects of
- the tissue injury lsee Guth et al., Gastroenterology, 87, 1083-90 (1984)].
` ~ Moreover, in the anesthetized cst, exogenous LTD4 evokes both incressed
~;~ pepsin secretion snd decressed transgastric potential [Pendlcton et al., Eur. J.
25 Phsrmscol., 125, 297-99 (1986)1. A particularly significant recent finding inthis regsrd is thst 5-lipoxygensse inhibitors snd some leukotriene sntagonists
i~ protect the gastric mucoss against lesions induced by the orsl or psrentersl -`
sdministrsion of most nonsteroidal antiinflammatory drugs [see Rainsford,
!,, ~ Agents and Actions, 21, 316-19 (1987)]. Accordingly, a significant body of
30 evldence implicates the involvement of lipoxygenase products in the develop~
` ~ ment of patholoficsl features sssociated with gsstric mucosal lesions, such ss
for example those induced by ethanol exposure and administration of non~
;- ~ steroidal Qnti-inflsmmstory drugs. Thus, compounds which inhibit the
biological effects of leukotrienes and/or which control the biosynthesis of
these substances, ss by inhibiting ~lipo~cygensse, are considered to be of vslue~s cytoprotective agents. ~ ~ -
1331~D
:
-- 3 --
Accordingly, the biological activitg of the leukotrienes and SRS's,
and of lipoxygenase as the enzyme leading to the metabolism of AA to
leukotrienes, indicates that a rational approach to drug therapy to prevent,
remove or ameliorate the symptoms of allergies, anaphylaxis, asthma and
5 inflammation and for gastric cytoprotection must focus on either blocking the
release of mediators of these conditions or antagonizing their effects. Thus
compounds, which inhibit the biological effects of the leukotrienes and SRS's
and/or which control the biosynthesis of these substances, as by inhibiting
lipoxygenase, are considered to be of value in treating such conditions as
10 allergic bronchial asthma, allergic rhinitis, as well as in other immediate
hypersensitivity reactions and in providing gastric cytoprotection.
It has now been found that certain novel naphthalenepropionic acid
derivatives inhibit lipoxygenase and antagonize products of the lipoxygenase
pathway, and so are useful as anti-inflammatory, anti-allergic and cytopro-
15 tective agents. The present invention provides novel compounds having thefollowing formula:
Cl H3
CHCOOH
~
~. ~"''','''.
A--CH2O~ 11 /J
wherein
A is alkgl of 3-19 carbon atoms, diloweralkylvinyl, dihalovinyl,
20diphenylvinyl, lower alkynyl,
l~X
S 11 or R2J~Z
X is N or CR;
R R R R R
Z is-C=/~-, -C=N-,-N=C-, N-,~ or -O-;
R is hydrogen or lower alkyl;
25Rl is hydrogen, lower alkyl or phenyl;
R2 is hydrogen or loweralkyl; or
~ ' '
,. .;
. .~. ~`,
~ ' ~ ' ' '; ' ' ' ` ' ' ' ' ~'~ ' ' ` ~ ' ` .' ' ' "!~. ' . ` - . ~,
3 3 ~ ~ 0 ~
- 4- ~ ;
~ ,:
Rl and R2 taken together form a benzene ring,
and the pharmaceutically acceptable salts thereof.
The terms "lower alkyr' and "lower alkynyr' refer to moieties
having 1 to 6 carbon atoms in the carbon chain.
S The compounds of the invention can be readily prepared according
to the following reaction sequence~
Cl H3 ~ -
CHCOOH
~ NaOCH3
A-CH2hal + ~\ 11 ~J -
CH3
I!~HC-OCH2--A
~ 1 NaOH >
A--CH2~
CH3 `
CNCOOH
A--CH20--_ ¦ 1 :- ,., - :.,
l ~
,`': '~'. '~.'`' ': '`'
l ~
~ 10 In the above reaction sequence, A is as defined hereinbefore and hal denotes
- c~oro, bromo or iodo. This preparative sequence reacts two equivalents of
the starting material ACH2hal with a metal derivative of the hydroxynaphtha- ;~
- lene propionic acid to form an intermediate ester ether which is hydroly~ed to ~ ;
yield the desired final products. The metal derivative of the hydroxynaphtha- ~ - -n
IS lene propionic acid may be prepared by treating the acid with an alkali metalalkoxide, such as sodium methoXide. In an alternative sequence, it is poæible
to use only one equivalent of starting material ACH 2hal with the metal
",,.-
.' :',,.'' :""
' '
.' '.. '' " ',~
A ,
, , ~ ~ :: ' ! ` ; ~. ' ~ . . . `'
L ~ . . ' ~ ; ~ r - ~ ~
'`- 1 33l00a
- s -
derivative to obtain the desired final product directly, without proceeding
through the ethyl ester intermediate.
In another reaction sequence, the desired final products can be
prepared by the alkylation of alkyl esters of the hydroxynaphthalene propionic
S acid:
fH3
CHCo2R3
~ K2C03/Acetone ', -
HO--~ A-CH2hal
f H3
CHCo2R3
A--CH20~1 hydrolysis >
',.
CIH3 :
CHC02H
A~112~
;`. . '
where hal is as defined hereinbefore and R3 is lower alkyl. Hydrolysis is
10 carried out using a dilute hydroxide, such as for example sodium hydroxide.
The starting materials used in the reaction sequences are available
commercially or can be prepared by known methods conventional in the art.
Thus, for example, the benzo-fused heterocyclic compounds such as l-methyl-
2-chloromethylbenzimidazole, 2-chloromethylbenzthiazole and 2-chloro-
15 methylbenzoxazole can be prepared by the following reaction scheme:
1 33100~
.,
- 6-
~ CICN2C~CH3 ~ CN2CI
wherein X is O, S or NCH3. The reaction is preferably carried out at a
controlled low temperature in an organic solvent, such as methylene chloride.
Compounds of the invenffon which contain a basic nitrogen are
capable of forming pharmaceutically acceptable s~lts, including the salts of
pharmaceutically acceptable organic and inorganic acids such as hydrochloric,
hydrobromic, sulfuric, nitric, phosphoric, methanesulfonic, benzenesulfonic,
acetic, citric, fumaric, maleic, succinic and the like. Also, the carboxylic acid
funcffon of the compounds of the invention is capable of forming salts with
10 pharmaceutically acceptable cations, such as alkali metal and alkaline earth
metal salts and ammonium and amine salts.
The compounds of the invention, by virtue of their ability to inhibit
the activity of lipoxygenase enzyme and to antagonize mediators arising from
this enzymaffc pathway, are useful in the treatment of inflammatory
15 conditions. Accordingly, the compounds are indicated in the treatment of suchdiseases as rheumatoid arthriffs, osteoarthritis, tendinitis, bursitis and similar
conditions involving inflammation. Moreover, by virtue of their ability to
inhibit the activity of lipoxygenase enzyme and by their ability to antagonize
the effect of LTC4, LTD4 and LTE4 which are the constituents of SRS-A, they
20 are useful for the inhibiffon of symptoms induced by these leukotrienes.
` Accordingly, the compounds are indicated in the prevention and treatment of
those disease states in which LTC4, LTD4 and LTE4 are causative factors, for
example allergic rhinitis, allergic bronchial asthma and other leukotriene
mediated nas~bronchial obstructive air-passageway conditions, as well as in
` 25 ot~er immediate hypersensitivity reactions, such as allergic conjunctivitis
The compounds are especially valuable in the prevention and treatment of
allergic bronchial asthma.
The compounds of the invention are cytoprotective agents and are
considered especially useful when edministered with convention~l non-
30 steroidal anti-inflammatory drugs, whose major side effect is gastrointestinal
irritation. The cytoprotective effect of the compounds of the invention
, ' ::'- '' '
; ~ ~331~0~ ~ -
-- 7 --
significantly reduces the gastroirritant impact of conventional anti-inflamma-
tory drugs. This effect is based not only on the ability of the compounds of
the invention to inhibit the biological effects of leukotrienes andlor control
the biosynthesis of these substances, as by inhibiting lipoxygenase, but also by ~ -
a shunting effect, whereby the control of the lipoxygenase pathway "shuntsn
the oxidation of arachidonic acid into the cyclooxygenase pathway, giving rise
to an increase in the formation of cytoprotective prostaglandins. These
biological effects make the compounds of the invention especially useful in
treating such conditions as erosive esophagitis, inflammatory bowel disease
and induced hemorrhagic lesions such as those induced by alcohol or non-
- steroidal anti-inflammatory drugs (NSAlD's), hepatic ischemia, noxious agent
induced damage ~x necrosis of hepatic, pancreatic, renal or myocardial tissue;
liver parenchyrnal damage caused by hepatotoxic agents such as carbon
tetrachloride snd ~galactosamine; ischemic renal failure; disease-induced
hepatic damage; bile salt-induced pancreatic or gastric damage; trauma or
stress-induced cell damage; and glycerol-induced renal failure.
When the compounds of the invention are employed in the treat-
ment of allergic airway disorders, as antiinflammatory agents and/or as - -
cytoprotective agents, they can be formulated into oral dosage forms such as
tablets, capsules and the like. The compounds can be administered alone or by
combining them with conventional carriers, such as magnesium carbonate,
magnesium stearate, talc, sugar, lactose, pectin, dextrin, starch, gelatin,
- tragacanth, methylcell~ose, sodium carboxymethylcellulose, low melting wax,
- cocoa butter and the like. Diluents, flavoring agents, solubilizers, lubricants, ~ - -
suspending agents, binders, tablet-disintegrating agents and the like may be ~ ~ -
employed. The compounds may be encapsulated with or without other
` carriers. In all cases, the proportion of active ingredients in said compositions
both solid and !iquid will be at least to impart the desired activity thereto onoral sdministration. The compounds may also be injected parenterally, in
which case they are used in the form of a sterile solution containing other
solutes, for example, enough saline or glucose to mHke the solution isotonic.
For administration by inhalation or insufflation, the compounds may be ~ -
formulated into an aqueous or partially aqueous solution, which can then be ;
utilized in the form of an aerosol. -
The dosage requirements vary with the particular compositions
employed, the route of administration, the severity of the symptoms presented
.'
~33~
-- 8 --
and the particular subject being treated. Treatment will gener~lly be initiated
with small dosages less than the optimum dose of the compound. Thereafter
the dosage is increased until the optimum effect under the circumstances is
reached. In general, the compounds of the invention are most desirably
5 administered at a concentration that will generally afford effective results
without causing any harmful or deleterious side effects, and can be
administered either as a single unit dose, or if desired, the dosage may be
divided into convenient subunits administered at suitable times throughout the
day.
10The lipoxygenase inhibitory and leukotriene antagonist effects as
well as the antiinflsmmatory and cytoprotective effects of the compounds of -
the invention may be demonstrated by standard pharmacological procedures,
which are descrlbed more fully in the examples given hereinafter.
These procedures illustrate the ability of the compounds of the
15 invention to inhibit the polymorphonuclear leukocyte synthesis of the lipoxy-genase product 5-HETE; the in vivo ability of the compounds to inhibit
bronchospasm induced by exogenously administered mediators of bronchocon-
striction; measure the in vivo activity of the compounds as antiinflammatory
agents in the rat carrageenan paw edema assay; measure the potential of the ;
20 compounds to induce acute gastroirritation in rats; measure the ability of the
compounds to prevent acute gastroirritation in rats induced by non-steroidsl
anti-inflammatory drugs.
The following examples show the preparation and pharmacological ~ -;
testing of compounds within the invention.
:' ~ - '. . , ' .
~: , . .,: .
~ r .... .' .
'."' '~ . ' '~'. ."'
.' .' ' ' ''' '` ~.
I ~ ... .. ...
~ . ' "~' '; ,.'''';
' ~
1331~00
g
l~sample I
-Methy~6 (2~nnolinylmeth~y)-2~aphthaleneaceffc acid
To a solution of 6-hydroxy- a-methyl-2-naphthaleneacetic acid
¦10.8 g, 50 mmol) in methanol (100 ml) is added sodium methoxide (100 mmol).
5 After 10 minutes the solvent is removed and replaced with dimethylformamide
(250 ml). 2-(Chloromethyl)quinoline (17.8 g, 100 mmol) is then added and the
reaction mixture is stirred for 8 days at room temperature. The reaction
mixture is partitioned between water and methylene chloride, the organic
layer is washed with water and evaporated to yield 27 g of an oil. Recrystalli-
10 zation of this oil twice from acetonitrile gives 10.6 g of white crystals ofintermediate ether ester (42% yield, m.p. 106-108C).
The ether ester from above is hydrolyzed as follows: a solution of
the ether ester (14.4 g, 28.9 mmol) in a mixture of 110 ml of lN NaOH and
110 ml tetrahydrofuran is refluxed for 1 hour. The organic solvent is then
15 removed and 2-(hydroxymethyl)quinoline is filtered off. The aqueous solution
is acidified to pH 6 and the precipitate is filtered and recrystallized from
ethanol to afford 2.2 g of white crystals (21% yield, m.p. 186-187C).
Analysis for: C23HlgNO3
Calculated C, 77.30; H, 5.36; N, 3.92.
20 Found C, 77.69; H, 5.36; N, 3.93.
E ample 2
a-Meth~6~ylmetho y)-~aphthl~leneaoeffc acid
` To a solut.on cf 6-hydroxy-o~-methyl-2-naphthaleneacetic acid
` (6.48 g, 30 mmol) in methanol (100 ml) is added sodium methoxide (60 mm).
25 After 10 minutes the solvent is removed and replaced with dimethylformamide
(100 ml). Benzyl chloride (7.6 g, 60 mmol) is then added and the reaction
mixture is stirred overnight at room temperature. The reaction is then heated
at 150C for 1 hour. The solvent is then removed and the residue is
partitioned between water and methylene chloride. The organic layer is
30 washed with water, dried over magnesium sulfate and evaporated to 11.5 g of
crude product. This crude product is extracted with 100 ml hot hexane which
affords 3.5 g of white crystals. A final recrystallization from methanol
affords a.s g of white crystals of the intermediate ethyl ester (2196 yield, m.p.
76-78C~.
'`'.~ .
.,
-
1 3 ~
- 10-
The ether ester (2.2 g, 5.56 mmol~ is hydrolyzed usine the method
of Example 1. RecrystaUization from ethanol affords 1.1 g of white crystals
(65% yield, m.p. 149-151C~.
AnalySiS for: C20H183
Calculated C, 78.40, H, 5.92.
Found: C, 78.31; H, 6.02.
l~samele 3
ai-lllethyl 6 Ul-methy}lH~ imid~o}~yl~ -
methosy ]-~naphthlllenell¢eti¢ a~id
The procedure of Example 2 is followed on a 10 mmol scale,
substituting 2-chlormethyl-N-methyl-benzimidazole for benzyl chloride. -;
Normal workup affords 2.5 g of white crystals of ether ester (50% yield, m.p. ~ -~
228-230 C~. This material is analyticaUy pure and needs no recrystaUization. ~d ~ -
The ether ester (2.5 g, 4.9 mmol) is hydrolyzed according to the -~
- lS method of Example 1. RecrystaUization from ethanol affords 0.75 g of white - ~ :
crystals (43% yield, m.p. 216-219C).
Analysis for: C22H2oN2o3 3/4 N2O
Calculated: C, 70.66; H, 5.79; N, 7.49. : - ~
Found C, 70.89; H, 5.72; N, 7.28. -
20 l~sample 4 ~
-MeUly}6-(~pyridinyhnetho~y)-2 naphthaleneaceticacid ~ -;To a solution of 6-hydroxy- ati-methyl-2-naphthaleneacetic acid
(10.8 g, S0 mmol) in methanol (100 ml) is added sodium methoxide (100 mmoV.
After 10 minutes the solvent is removed and replaced with hexamethylphos- --~
25 phoric triamide (250 ml). 2-(Chloromethyl)pyridine (100 mmol) is then added ` ~ ~;
and the reaction is stirred for 5 days. The reaction is worked up by -
partitioning between water and methylene chloride, and the organic extract is
evaporated to a crude oil which is chromatographed on silica gel (eluant: - --
methylene chloride - ethyl acetate) to afford 6.0 g of ether ester as an oil.
The ether ester is hydrolyzed using the method of Example 1. -
Reorystallization from ethanol affords S.S g of- white crystals (36~6 yield, m.p.
185-187C).
Analysis for: ClgH17NO3
Calculated C, 74.24; H, 5.5q; N, 4.56.
35 Found C, 74.49; H9 5.63; N, 4.49.
, ...
,.
~3310~
E amPle 5
a-Methy~6~2 ben~othiazolylmethoxy~2-naphtl~leneaceffc acid
The title compound is prepared according to the method of
Example 1, using 2-(chloromethyl)benzothiazole. White crystals are obtained
s having a melting point of 169-171C.
Analysis for: C21H17NO3S 1/4 H2O
Calculated C, 68.55; H, 4.79; N, 3.80.
FoundC, 68.44; H, 4.89; N, 4.21.
Example 6
10 a-Yetl~6 (2-naphthlllenylmethoxy~2-naphtluleneaceffca~id
The title compound is prepared according to the method of
Example I using 2-(chloromethyl)naphthalene. White crystals are obtained
having a melting point of 216-218C.
AnalySiS for: C24H203
Calculated: C, 80.88; H, 5.66.
FoundC, 80.68; H, 5.89.
E ample 7
S~ a-ll~etl~6-(2-quinolinylmethosy~2-naphthsleneacetic acid
~o a solution of ~(~-methyl-6-hydroxy-2-naphthalene acetic
20 acid*(21.6 g, 100 mmol) in methanol (250 ml) is added sodium methoxide
(200 mmol). The solvent is removed in vacuo and replaced with dimethylform-
amide (300 ml). To this solution is added 2-(chloromethyl)quinoline (17.7 g,
100 mmol). After 90 minutes, the solvent is removed in vacuo at 50C and the
residue is partitioned between ethyl acetate and p~l = 4 buffer. The insolubles `
and the ethyl acetate are heated to 60C at which point H homogeneous
solution is obtained. Cooling the solution to room temperature affords 8.9 g of
white crystals (2696). A second recrystallization of 4.0 g of this material fromMethanol (200 ml~ affords 1.78 g of white crystals, m.p. 192-194C.
An~ysis for: C23HlgNO3
Calculated: C, 77.30; Hl, 5.36; N, 3.92. ~ ~ ~
Found: C, 76.96; H, 5.44; N, 3.89. ~,
(a)D = ~Sl~1 (Pyr c = 1.115) ~ -
* Prepared according to the procedure described in J. Med. Chem., 17, 377
~1974).
. ,~
. ::.:
~331~
- 12-
E ample 8
R-(-~a-ldethy~6 (2 quinolinylmetho~2-naphtheleneacetic acid
The title compound can be prepared according to the method of
Example 7, using R-(-}a-methyl-6-hydroxy-2-naphthalene acetic acid. White ~ -~
crystals are obtained having a melting point of 190-193C.
Analysis for: C23HlgNO3
Calculated: C, 77.30; H, 5.36; N, 3.92. -
Found C, 76.87; H, 5.54; N, 3.77.
()D = -52.96 (Pyr c = 1.08)
E ample 9
a-~etl~6{(2-pheny~thiazolyl~methosy ]-2-naphth~leneacetic acid
The title compound is prepared according to the method of
Example 7 using 2-phenyl-4-(chloromethyl)thiazole and racemic ~hydroxy--
methyl-2-naphthalene acetic acid. White crystals are obtained having a - ~ -
meltingpointof 163-164C. -
Analysis for: C23HlgNO3S
Calculated: C, 70.93; H, 4.92; N, 3.60.
Found:C, 70.75; H, 4.77; N, 3.32.
Esample 10
a-Methyi 6-(hesylosy)-2-naphthaleneacetic acid
The title compound is prepared according to the method of
Example 7 using l-iodohexane and racemic 6-hydroxy-a-methyl-2-naphthalene ~ ~
acetic acid. White crystals are obtained having a melting point of 88-91C. -
Analysis for: Cl9H243 ~- -
Calculated C, 75.97; H, 8.05.
FoundC, 76.20; H, 8.25.
, -, -
Esample 11
- a-Methy~6 (butylo y~phth~leneacetic acid -
The title compound is prepared according to the method of
30 Example 7 using l-iodobutane and racemic 6-hydroxy--methyl-2-naphthalene
acetic acid. White crystals are obtained having a melting point of 125-127C.
: - ' ;
;' :
.. ..
. ",. ,
. .
- 133~00~
- 13-
Analysis for: C17H 203
Calculated: C, 74.97; H, 7.40.
Found. C, 75.04; H, 7.02.
E amlile 12
a-MethyW (pentylo~y~2~aphtlul1eneacetic acid
The title compound is prepared according to the method of
Example 7 using l-iodopentane and racemic 6-hydroxy-a-methyl-2-naph-
thalene acetic acid. White crystals are obtained having a melting point of 97-
99C.
10 Analysis for: ClgH22O3
Calculated: C, 75.50; H, 7.74.
Found:C, 75.54; H, 7.68.
Example 13
a-l~ethyl~(dodecylo~2 naphthaleneacetic acid
The title compound is prepared according to the method of
' Example 7 using l-iodododecane and racemic 6-hydroxy- a-methyl-2-naph-
th~lene acetic acid. White crystals are obtained having a melting point of 96-
- 9~ C .
AnalysiS for- C2sH363
20 Calculated C, 78.08; H, 9.44.
FoundC, 78.10; H, 9.77.
l ..
E~ample 14
-Methy~6 (3-meU~y~2 butenoxy~2~aphthsleDeaceffcacid
The title compound is prepared according to the method of - -
25 Example 7 using 1-bromo- 3-methyl-2-butene and racemic 6-hydroxy-a- ;~
methyl-2-naphthalene acetic acid. White crystals are obtained having a
melting point of 97-100C.
AnalYsisfOr: - C18H203 ~ f
` Calculated: C, 76.03; IX, 7.09. ~ ;
30 Found C, 76.38; H, 7.10.
~"
~3310~0
- 14-
Example 15 - e~
a ethyl~(3,3~icliloro~1ylo y~uphth~l1eneacetic acid
The title compound is prepared according to the method of ' ~ ~
Example 7 using 3,3-dichloroallyl chloride and racemic ~hydrox~ a-methyl-2- ~ -
naphthalene acetic acid. White crystals are obtained having a melting point of ; ~ : ;
107-109C.
AnalysiS for: C16H14O3C12
Calculated: C, 59.10; H, 4.34.
Found C, 59.43; H, 4.50. - -
: . .: , .
E ample 16
The compounds 5- and 12-hydroxyeicosatetraenoic acid (5-HETE
and 12-HETE) srld 5,12-dihydroxyeicosatetraenoic acid (5,12-diHETE) are early
arachidonic acid oxidation products in the lipoxygenase cascade, which have
been shown to mediate several aspects of inflammatory and allergic response.
- 15 The assay of this Example measures the ability of the compounds of the
invention to inhibit the synthesis of 5-HETE by rat glycogen elicited poly-
morphonuclear leukocytes.
The ~ssay is carried out as follows:
Peritoneal PMN are obtained from female Wistar rats (150-250 g)
~` 20 that received an i.p. injection of 696 glycogen (10 ml). After 24 hours, rats
are killed by C2 asphyxiation and peritoneal cells are harvested by peritoneal
- lavage using Ca+~and Mg+~ free Hanks' balanced salt solution (HBSS). The
peritoneal exudate is centrifuged at 400 g for 10 minutes. After centrifuga-
tion, the lavaged fluid is removed and the cell pellet is tesuspended in HBSS
25 containing CA~+ and Mg+~ and 10 mM L-cysteine at a concentration of 2 x 107
- cells/ml. To I ml portions of cell suspension, test drugs or vehicle are added
and incubated at 37C for 10 minutes. Following this preincubation, the
`~ calcium ionophore (10 llM), A23187, is added together with 0.5 uCi [14~arachidonic acid and further incubated for 10 minutes. The reaction is stopped
30 by the addition of ice cold water (3 ml) and acidifying to pH 3.5. Lipoxygenase
products are then extracted twice into diethyl ether. The pooled ether
extracts are evaporated to dryness under nitrogen and the residue is redis-
solved in a small volume of methanol and spotted on aluminum backed pre-
coated thin layer chromatographic plates. The samples are then cochromato-
35 graphed with authentic reference 5-HETE in the solvent system -
~ 33~ 0~
- 15-
hexane: ether: acetic acid (50:50:3). After chromatography, the areas
associated with 5-HETE standard are identified by autoradiography, cut out
and quantitated by liquid scintillation.
The compounds of the invention and the nonsteroid~l anti-inflam-
5 matory drug naproxen, when tested in this assay at the level of lOllM, gave
the following resuits in inhibiting the synthesis of the arachidonic acid
lipoxygenase oxidation product S-HETE.
Table 1
Compound of % Inhibition of 5-LO
10Example No.~as S-HETE~
naproxen -94 *
100
2 84
3 24
lS 4 82
~ 6 10O
': ~ 100 ~ ,:.
8 33
20 9 86
81
1l 28 -~
12 48
14 81 ~ -~
25 * The negative value denotes a potentiation of 5-HETE synthesis.
*~ Tested at a level of 0.1 llM.
These resuits show that the compounds of the invention exhibit
very significant activity in inhibiting the enzyme, ~lipoxygenase.
E~mple 1~
The procedure of Example 16 is also employed for the determina-
tion of the ability of the compounds of the invention to inhibit the synthesis of ~ ~ ~
the arachidonic acid cyclooxygenase oxidation products TxB2 and PGE2. ~ ~ :
: '' ': ~" ,:
- l 3 ~
- 16- ~
.
In this assay, the procedure of Example 16 is carried out as
described. However, in order to determine cyclooxygenase activity, the
samples are cochromatographed with authenffc reference TxB2 and PGE2 in
the solvent system ethyl acetate: formic acid (80:1) and the upper phase of
ethyl acetate: isooctane: acetic acid: water (110:50:20:100). After chroma-
tography, the areas associated with the TxB2 and PGE2 standards are
identified by autoradiography, cut out and quantitated by liquid scintillation - -
techniques.
The results are calculated as in Example 16.
When tested in this assay the compounds of the invention and the
nonsteroidal anti-inflammatory drug naproxen, a wel} established inhibitor of
cyclooxygenase, at a level of 10 llM, gave the following results in inhibiffng
the synthesis o the arachidonic acid cyclooxygenase oxidation products TxB2 ;
and PGE2-
Table 2
Compound of% Inhibition of CO % Inhibition of CO : ;
Example No.tas TxB~) (as PGE2)
naproxen 85
a8 -37* ~ ~
20 2 6 -a6* ~ -
3 15 -29 * - - --
4 12 18
-35 *
; -
6 -lS * - -
25 7 ~lq *
8 -7 *, * * ~ -
9 -11 *
~5 ~
11 .-: '
12 -13 *
14 -15 *
* The negative values denote H potentiHtion of PGE2 synthesis. ~~-
** Tested at level of 0.1 IIM.
,'~
':
' ' ' :', , '
i33100~ -
-- 17 --
These results show that the compounds of the invention, in
contradistinction to naproxen, are virtually devoid of cyclooxygenase inhibi-
tory activity, having activity substantially only on the lipoxygenase pathway ofarachidonic acid oxidation.
Example 18
The assay of this Example measures the in vivo ability of the
compounds of the invention to inhibit the bronchospasm induced in guinea pigs
by the exogenously administered leukotrienes C4 and/or D4.
This assay is carried out as followæ
Male Hartley strain guinea pigs ~350-600g) are anesthetized with
pentobarbital sodium (50 mg/kg, i.p.). The jugular vein is cannulated for
injection of drugs and the carotid artery for monitoring blood pressure. The ~ ~;
trachea is cannulated for artificial ventilation by a miniature Starling pump
and for indirect measurement of respiratory volume changes. The animals are
then pretreated with succinylcholine (2 mglkg i.v.) and indomethacin
~10 mg/kg i.v in trizma 8.3 buffer, 9 minutes prior to leukotriene challenge). ~; -
Submaximal bronchoconstrictor responses are established in control animals by
varying the dose-levels of leukotriene. Intravenous dose-levels for LTC4 range -
from 0.4 to 0.6 ~g/kg and for LTD4 the range is from 0.3 to 0.5 lJglkg. The ~- -
aerosol bronchoprovocation dose for LTC4 is generated from 1.6 llM solution
and for LTD4 from a 2.0 llM solution.
Test drugs (dissolved in a solvent such as propylene ~Iycol, poly~
ethylene glycol 400 or saline) are administered either intraduodenally, by
aerosol or intragastrically at 2 or 10 minutes before induction of bronchospasm - ; --
25 by administration of either LTC4 or LTD4 at the predetermined dose-levels.
Aerosols of soluble drugs or leukotrienes are produced in-line for 10 seconds
only by actuation of an ultrasonic nebulizer (Monaghan). Aerosdlized drug
` dosage is expressed in terms of solution concentration and by a fixed aerosol - - ~ -
s~ exposure time (approximately 10 seconds). Control animals receive solvent -
30 (2 mVkg i.d. or appropriate aerosol) in place of drug.
Respiratory volume changes are determined by a calibrated piston
` whose travel is recorded, via a linear transducer, on a Beckman Dynograph -
recorder. Maximal bronchoconstrictor volume is determined by clamping off `~
:~
:: :
-- 18-
the trachea at the end of the experiment. Overflow volumes at 1, 3 and S
minutes are obtained from the recorded charts.
Area under the volume overflow curve (AUC) is estimated, using
the overflow values at 1, 3 and 5 minutes, and expressed as a percentage of
5 the maximal overflow AUC (equation 1).
3 (1 min) + 4 (3 min) + 2 (5 min)
% maxAUC = X 100 (1
10 (max)
Drug effects are reported as percent inhibition of % max AUC values obtained
from appropriate control animals (eguation 2):
% max AUC control- % max AUC treated
% inhibition = X 100 (2 - ~
% max AUC control ~ -
Student's t-test for unpaired data is used to determine statistical
sigi~ificance (p < 0.05). ICso values can also be determined by inverse
prediction from linear regression lines through points between 10 and 90%
inhibition.
; The results for compounds of the invention are as follows: -
Table 3
Compound administered at 10 minutes before induction of bronchospasm
using LTD4
~ Compound of {)ose*
Example Numbermg/kg% Inhibition
20 1 25 75
2 25 -76 ~ -
3 25 80 ~ ~;
~' - 4 25 52 ~ -
* = intraduodenally administered
: '',''
The results show that compounds of the invention have significant
in vivo activity against LTD4 induced bronchoconstriction.
. .
~ ,
. ' .
: ' . . '.
.~ `.`' ' . 1 . :
.
.
` 1~31~
:
-- 19--
~ ample 19
The compounds of the invention are further tested in the rat
carrageenan paw edema assay to determine their ability to inhibit the scute
inflammatory response.
This assay is carried out as follows:
140-180 gm male Sprague-Dawley rats, in groups of 6 animals are
injected subcutaneously in the right paw with 0.1 ml of 1% carrageenan at
zero ffme. Mercury plethysmographic readings (ml) of the paw are made at
zero time and 3 hours later. Test compounds are suspended or dissolved in
0.5% methylcellulose and given perorally 1 hour prior to carrageenan admini- ;
stration.
The increase in paw volume (edema in ml.) produced by the
- ~ carrageenan is measured. Paw edema is calculated (3 hour volume minus zero ~ ;
- time volume), and percent inhibition of edema is determined. Unpaired ~ -
Student's t-test is used to determine statistical significance. ~ - - -
The activity of standard drugs in this assay is as follows: - - --
Drug Oral EDso (9596 C.L mg/kg
Indomethacin 3.7 (0.6, 23.8)
~;~ Rspirin 145.4 (33.1, 645.6)
` ~ 20 Phenylbutazone 26.2 (2.3, 291.0) ` ~ - ;
~; When tested in this assay, the compounds of the invention gave the ~ ~ -
r ~ ; following results:
Tabh 4
Compound of% Inhibition at
` ~ 25 Example No.50 mglkg(peroral)
1 42
~ 2 24
4 30
' ~
The results show that the compounds tested have activity in the rat ~ ;
carrageenan paw edema assay, evidencing an effect on the acute inflammatory
response.
`..................................................................... ~,.. -~'.
-~ ~33~.~3~ ;
- 20- "
EDample 20
The ~ompounds of the invention are tested in the rat acute
gastroirritation asgay to examine their potential for causing gastric irritationwhen administered at doses exceeding the effective dose. The nonsteroid~l
S anti-inflammatory drug naproxen is tested as a standard of a compound known _
to p~ssess gastroirritant side effects.
This assay is carried out as follows: ,
Male Sprague Dawley rats (190-220 g) are fasted for 18 hours prior
to drug administration. Rats are divided into groups of 8 and coded (i.e7 ,
observer of gastric lesions is not aware of drug treatment). Drugs, were~
dissolved or sugpended in 0.5% Tween~80 and administered by gastric intuba~-
Ç tion in a volume of I mV100 g body weight, control rats receiving only Tw~
80. Four hours after drug administration, rats are evaluated by recording the
incidence and severity of gastroirritation using the following scoring system:-
- 15 0) No irritation or lesion$ 1) irritation (rednessk 2) < 5 ledons (ulcers) and
3) > 5 legions. Dunnett's test (a = 0.05) was used to cslculste the mean,~
of ea~h test grou,o and the statistical signiffcance. ,,
The results of this- ~y are presented ln Tsble 5.
Table 5
~- 20CompoundofDose% of rats with - - -~
E~ample No. ma/k~ Gl lesi~
, ~ n~lproxen 25 100 '''
~- 1 300
~', 10 400 0
2511 400 50
12 400 25 ',~
' The results sho~v the compoun~ of the invention to have litUe potential,for
acut~ gastroirritation ~hen compsred to naproxen.
.
* trade-mark
'~
. -
: , '
'A
. ,~
~3310~0
- 21-
l~ ample 21
The assay of Example 20 is also used to measure the cytoprotective :~ .
activity of the compounds against acute gastroirritation induced by a
nonsteroidal anti-inflammatory drug (NSAlD). ~ -
5The assay is carried out as presented in Example 20 with the
following modifications: one hour before administration of the NSAID, the -
rats perorally receive either vehicle (1 mV100 g body weight of 0.5% Tween :
. 80) or drug (100 mg/kg), at 0 hour the rats perorally receive 25 mg/kg of the
NSAID naproxen or vehicle (I ml/100 g body weight of 0.5% Tween 80k at 4
10hours post~osing the rats are sacrificed and gastroirritation assessed as ~ ~ :
described in Example 20. . ~
The results are presented in Table 6. ~- :
Table 6
CompoundofDose% of Rats
15 Example No.m~/kg~ith Lesions Comments
: vehicle 12.5
naproxen 25 75.0
~`-; 7 100 87.5 no protection
100 37.5 protection
11 100 50.0moderate protection ~ ;
r . , , . .'
' ' ~
~ ' ',~'~` ~; ,"'`''~
