U.S. patent application number 09/804742 was filed with the patent office on 2001-08-16 for method for preventing and treating alcoholism.
Invention is credited to Bueno, Lionel, Chovet, Maria, Diop, Laurent, Guglietta, Antonio, Little, Hilary J., Rafferty, Michael Francis, Ren, Jiayuan, Taylor,, Charles Price JR., Watson, W. P..
Application Number | 20010014698 09/804742 |
Document ID | / |
Family ID | 27369092 |
Filed Date | 2001-08-16 |
United States Patent
Application |
20010014698 |
Kind Code |
A1 |
Bueno, Lionel ; et
al. |
August 16, 2001 |
Method for preventing and treating alcoholism
Abstract
GABA analogs are useful to prevent and treat gastrointestinal
damage and ethanol withdrawal syndrome. Preferred treatments employ
gabapentin or pregabalin.
Inventors: |
Bueno, Lionel; (Aussonne,
FR) ; Chovet, Maria; (Montrouge, FR) ; Diop,
Laurent; (Saclay, FR) ; Guglietta, Antonio;
(Ann Arbor, MI) ; Little, Hilary J.; (County
Durham, GB) ; Rafferty, Michael Francis; (Ann Arbor,
MI) ; Ren, Jiayuan; (Oklahoma City, OK) ;
Taylor,, Charles Price JR.; (Chelsea, MI) ; Watson,
W. P.; (County Durham, GB) |
Correspondence
Address: |
Warner-Lambert Company
2800 Plymouth Road
Ann Arbor
MI
48105
US
|
Family ID: |
27369092 |
Appl. No.: |
09/804742 |
Filed: |
March 13, 2001 |
Related U.S. Patent Documents
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Application
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09804742 |
Mar 13, 2001 |
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09567191 |
May 9, 2000 |
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6242488 |
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09567191 |
May 9, 2000 |
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09284710 |
Apr 19, 1999 |
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6127418 |
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09284710 |
Apr 19, 1999 |
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PCT/US98/17082 |
Aug 18, 1998 |
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60082936 |
Apr 24, 1998 |
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60074794 |
Feb 16, 1998 |
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60056753 |
Aug 20, 1997 |
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Current U.S.
Class: |
514/561 |
Current CPC
Class: |
A61P 1/00 20180101; A61P
25/32 20180101; A61K 31/197 20130101; A61K 31/195 20130101; A61P
1/04 20180101; A61K 31/40 20130101; A61K 31/42 20130101; A61K 31/54
20130101; A61P 29/00 20180101; A61K 31/405 20130101; A61K 45/06
20130101; A61K 31/60 20130101; A61K 31/60 20130101; A61K 31/195
20130101; A61K 31/54 20130101; A61K 31/195 20130101; A61K 31/42
20130101; A61K 31/195 20130101; A61K 31/405 20130101; A61K 31/195
20130101; A61K 31/40 20130101; A61K 31/195 20130101; A61K 31/195
20130101; A61K 31/195 20130101; A61K 31/195 20130101; A61K 31/19
20130101; A61K 31/195 20130101; A61K 2300/00 20130101; A61K 31/40
20130101; A61K 2300/00 20130101; A61K 31/405 20130101; A61K 2300/00
20130101; A61K 31/42 20130101; A61K 2300/00 20130101; A61K 31/54
20130101; A61K 2300/00 20130101; A61K 31/60 20130101; A61K 2300/00
20130101 |
Class at
Publication: |
514/561 |
International
Class: |
A61K 031/195; A61K
031/192 |
Claims
What is claimed is:
1. A method for preventing and treating gastrointestinal damage and
disorders comprising administering to a subject in need of
treatment an effective amount of a GABA analog.
2. A method according to claim 1 employing a compound of Formula I
3wherein R.sub.1 is hydrogen or lower alkyl and n is an integer of
from 4 to 6, and the pharmaceutically acceptable salts thereof.
3. The method according to claim 2 employing gabapentin.
4. The method according to claim 1 employing a compound selected
from (1 -aminomethyl-3-methylcyclohexyl)acetic acid,
(1-aminomethyl-3-methylcyclo- pentyl)acetic acid, and
(1-aminomethyl-3,4-dimethylcyclopentyl)acetic acid.
5. A method according to claim 1 employing a compound of Formula II
4wherein R.sub.1 is a straight or branched alkyl of from 1 to 6
carbon atoms, phenyl, or cycloalkyl of from 3 to 6 carbon atoms;
R.sub.2 is hydrogen or methyl; and R.sub.3 is hydrogen, methyl, or
carboxyl, and the pharmaceutically acceptable salts thereof.
6. The method according to claim 5 employing pregabalin.
7. The method according to claim 5 employing
R-(3)-(aminomethyl)-5-methyl-- hexanoic acid.
8. The method according to claim 5 employing
3-(1-aminoethyl)-5-methylhexa- noic acid.
9. The method according to claim 1 wherein the gastrointestinal
damage is in a subject who is receiving or will receive NSIAD
therapy or alcohol.
10. The method according to claim 1 wherein the gastrointestinal
damage is characterized as inflammatory bowel disorder or irritable
bowel syndrome.
11. The method according to claim 1 wherein the condition treated
is selected from Crohn's disease, ileitis, ischemic bowel disease,
dyspepsia, and ulcerative colitis.
12. A method for treating ethanol withdrawal syndrome in a mammal
in need of treatment comprising administering an effective amount
of a GABA analog.
13. A method according to claim 12 employing a compound of Formula
I 5wherein R.sub.1 is hydrogen or lower alkyl and n is an integer
of from 4 to 6, and the pharmaceutically acceptable salts
thereof.
14. The method according to claim 13 employing gabapentin.
15. A method according to claim 12 employing a compound of Formula
II 6wherein R.sub.1 is a straight or branched alkyl of from 1 to 6
carbon atoms, phenyl, or cycloalkyl of from 3 to 6 carbon atoms;
R.sub.2 is hydrogen or methyl; and R.sub.3 is hydrogen, methyl, or
carboxyl, and the pharmaceutically acceptable salts thereof.
16. The method according to claim 15 employing pregabalin.
17. A pharmaceutical composition comprising a GABA analog and a
non-steroidal anti-inflammatory drug together with a
pharmaceutically acceptable excipient, carrier, or diluent
therefor.
18. A composition of claim 17 wherein the GABA-analog is a compound
of Formula I 7wherein R.sub.1 is hydrogen or lower alkyl and n is
an integer of from 4 to 6, and the pharmaceutically acceptable
salts thereof.
19. The composition of claim 18 wherein the GABA analog is
gabapentin.
20. A composition according to claim 17 wherein the GABA analog is
a compound of Formula II.
21. The composition of claim 20 wherein the GABA analog is
pregabalin.
22. A composition according to claim 17 wherein the non-steroidal
anti-inflammatory drug is selected from sulindac, naproxen,
indomethacin, mefenamic acid, diclofenac, fenoprofen, diflunisal,
etodolac, ibuprofen, piroxicam, acetylsalicylic acid, oxaprozin,
and bromfenac, or pharmaceutical salts thereof.
23. A composition according to claim 22 wherein the non-steroidal
anti-inflammatory drug is selected from naproxen sodium, ibuprofen,
or indomethacin.
24. A composition of claim 23 comprised of naproxen sodium and
pregabalin.
25. A composition of claim 23 comprised of naproxen sodium and
gabapentin.
26. A composition of claim 23 comprising ibuprofen and
pregabalin.
27. A composition of claim 23 comprising ibuprofen and gabapentin.
Description
FIELD OF THE INVENTION
[0001] This invention relates to a method for preventing visceral
and gastrointestinal damage such as gastric ulcers by administering
a gamma-aminobutyric acid (GABA) analog, and for treating
gastrointestinal diseases such as inflammatory bowel disorders
(IBD), functional bowel disorders (FBD), including dyspepsia and
other visceral pain.
BACKGROUND OF THE INVENTION
[0002] Nonsteroidal anti-inflammatory drugs (NSAIDs) are the most
frequently prescribed drugs for the treatment of pain associated
with osteoarthritis and many other musculoskeletal and inflammatory
disorders. In the United States, about 100 million prescriptions
are written each year to provide effective relief of pain and
treatment of inflammatory diseases. Commonly used NSAIDs include
sulindac, naproxen, indomethacin, mefenamic acid, diclofenac,
fenoprofen, and diflunisal.
[0003] However, considerable evidence indicates that NSAIDs have
frequent, serious, and costly gastrointestinal tract toxic side
effects. These include mild dyspepsia, gastritis, peptic ulcer
disease, as well as more serious gastrointestinal complications
such as bleeding and perforation, leading sometimes to significant
morbidity and, to a lesser extent, mortality. Serious GI
complications due to NSAID use represent the greatest threat to
life in patients with connective tissue diseases, second only to
the primary disease and its complications. Similar gastrointestinal
damage is caused by ingestion of alcohol. Indeed, a condition known
as ethanol withdrawal syndrome is commonly encountered when
prolonged ethanol consumption is terminated. In addition to
gastrointestinal damage, this syndrome often results in tremors,
anxiety, convulsions, hallucinations, and confusion.
[0004] Other commonly encountered gastrointestinal disorders
include inflammatory bowel disorders (IBD) and functional bowel
disorders (FBD), including dyspepsia. These GI disorders include a
wide range of disease states that are currently only moderately
controlled, including Crohn's disease, ileitis, ischemic bowel
disease, and ulcerative colitis, as well as IBD, the irritable
bowel syndrome, dyspepsia, and gastro-esophageal reflux for FBD,
and other forms of visceral pain.
[0005] Gamma-aminobutyric acid has been shown to activate gastric
afferent nerves which, in turn, have been shown to participate in
gastric defense mechanisms. We have now discovered that GABA
analogs dramatically reduce the gastrointestinal damage caused by
drugs and alcohol. The GABA analogs also treat the conditions
resulting from ethanol withdrawal syndrome, and GI disorders
characterized as IBD and IBS.
[0006] All that is required to prevent gastrointestinal damage and
to treat IBD, IBS, and alcoholism according to this invention is to
administer to a subject who is in need of treatment an effective
amount of a GABA analog.
[0007] Several GABA analogs are known. Gabapentin, a cyclic GABA
analog, is now commercially available and extensively used
clinically for treatment of epilepsy and neuropathic pain. Such
compounds are described in U.S. Pat. No. 4,024,175. Another series
of GABA analogs which are anti-seizure agents is described in U.S.
Pat. No. 5,563,175.
SUMMARY OF THE INVENTION
[0008] This invention provides a method for preventing and treating
gastrointestinal damage and disorders comprising administering to a
subject in need of treatment an effective amount of a GABA analog.
A preferred embodiment utilizes a cyclic amino acid compound of
Formula I 1
[0009] wherein R.sub.1 is hydrogen or lower alkyl and n is an
integer of from 4 to 6, and the pharmaceutically acceptable salts
thereof. An especially preferred embodiment utilizes a compound of
Formula I where R.sub.1 is hydrogen and n is 5, which compound is
1-(aminomethyl)-cyclohe- xane acetic acid, known generically as
gabapentin. Other preferred GABA analogs have Formula I wherein the
cyclic ring is substituted, for example with alkyl such as methyl
or ethyl. Typical compounds include
(1-aminomethyl-3-methylcyclohexyl)acetic acid,
(1-aminomethyl-3-methylcyc- lopentyl)acetic acid, and
(1-aminomethyl-3,4-dimethylcyclopentyl)acetic acid.
[0010] In another embodiment, the method of the invention utilizes
a GABA analog of Formula II 2
[0011] or a pharmaceutically acceptable salt thereof, wherein
[0012] R.sub.1 is a straight or branched alkyl of from 1 to 6
carbon atoms, phenyl, or cycloalkyl of from 3 to 6 carbon
atoms;
[0013] R.sub.2 is hydrogen or methyl; and
[0014] R.sub.3 is hydrogen, methyl, or carboxyl.
[0015] Diastereomers and enantiomers of compounds of Formula II can
be utilized in the invention.
[0016] An especially preferred method of the invention employs a
compound of Formula II where R.sub.2 and R.sub.3 are both hydrogen,
and R.sub.1 is --(CH.sub.2).sub.0-2-i C.sub.4H.sub.9 as an (R),
(S), or (R,S) isomer.
[0017] A more preferred embodiment of the invention utilizes
3-aminomethyl-5-methyl-hexanoic acid, and especially
(S)-3-(aminomethyl)-5-methylhexanoic acid, now known generically as
pregabalin, as well as CI-1008. Another preferred compound is
3-(1-aminoethyl)-5-methylhexanoic acid.
[0018] The invention additionally provides a composition comprised
of an anti-inflammatory amount of an NSAID and a cytoprotective
amount of a GABA analog.
DETAILED DESCRIPTION OF THE INVENTION
[0019] As noted above, the method of this invention utilizes any
GABA analog. A GABA analog is any compound derived from or based
upon gamma-aminobutyric acid. The compounds are readily available,
either commercially, or by synthetic methodology well-known to
those skilled in the art of organic chemistry. The preferred GABA
analogs to be utilized in the method of this invention are cyclic
amino acids of Formula I. These are described in U.S. Pat. No.
4,024,175, which is incorporated herein by reference. Another
preferred method utilizes the GABA analogs of Formula II, and these
are described in U.S. Pat. No. 5,563,175 which is incorporated
herein by reference.
[0020] All that is required to practice the method of preventing
and treating gastrointestinal damage and disorders of this
invention is to administer a GABA analog in an amount that is
effective to prevent or treat the damage condition, i.e., to combat
the effects of a NSAID or alcohol, or to control IBD and IBS. The
invention includes a method for treating ethanol withdrawal
syndrome and general alcoholism. The effective amount of GABA
analog to be utilized will generally be from about 1 to about 300
mg per kg of subject body weight. Typical doses will be from about
10 to about 5000 mg per day for an adult subject of normal
weight.
[0021] Typical "gastrointestinal damage" conditions caused by NSAID
use include dyspepsia, gastritis, peptic ulcer, as well as lower
gastrointestinal bleeding and perforation. Further effects of
ethanol withdrawal syndrome include tremor, anxiety, and
convulsions. Typical IBD conditions include ileitis, ulcerative
colitis, and Crohn's disease.
[0022] Pharmaceutical compositions of the compound of the present
invention or its salts are produced by formulating the active
compound in dosage unit form with a pharmaceutical carrier. Some
examples of dosage unit forms are tablets, capsules, pills,
powders, aqueous and nonaqueous oral solutions and suspensions, and
parenteral solutions packaged in containers containing either one
or some larger number of dosage units and capable of being
subdivided into individual doses. Some examples of suitable
pharmaceutical carriers, including pharmaceutical diluents, are
gelatin capsules; sugars such as lactose and sucrose; starches such
as corn starch and potato starch, cellulose derivatives such as
sodium carboxymethyl cellulose, ethyl cellulose, methyl cellulose,
and cellulose acetate phthalate; gelatin; talc; stearic acid;
magnesium stearate; vegetable oils such as peanut oil, cottonseed
oil, sesame oil, olive oil, corn oil, and oil of theobroma;
propylene glycol, glycerin; sorbitol; polyethylene glycol; water;
agar; alginic acid; isotonic saline, and phosphate buffer
solutions; as well as other compatible substances normally used in
pharmaceutical formulations. The compositions of the invention can
also contain other components such as coloring agents, flavoring
agents, and/or preservatives. These materials, if present, are
usually used in relatively small amounts. The compositions can, if
desired, also contain other therapeutic agents. For use in
combating the gastrointestinal effects of NSAIDs, the GABA analogs
can be administered alone in unit dosage form, or in combination
with the NSAID being utilized for the particular patient.
[0023] The percentage of the active ingredient in the foregoing
compositions can be varied within wide limits, but for practical
purposes it is preferably present in a concentration of at least
10% in a solid composition and at least 2% in a primary liquid
composition. The most satisfactory compositions are those in which
a much higher proportion of the active ingredient is present, for
example, from 10% to 90% by weight.
[0024] Routes of administration of the subject compound or its
salts are oral or parenteral. For example, a useful intravenous
dose is between 5 and 50 mg and a useful oral dosage is between 20
and 800 mg. The dosage is within the dosing range used in treatment
of gastrointestinal diseases such as ulcers and IBS, or as would be
dictated by the needs of the patient as described by the
physician.
[0025] A unit dosage form of the GABA analog to be used in this
invention may also comprise other compounds useful in the therapy
of gastrointestinal diseases.
[0026] The advantages of using the compounds of Formula I and II,
especially gabapentin and pregabalin, in the instant invention
include the relatively nontoxic nature of the compounds, the ease
of preparation, the fact that the compounds are well-tolerated, and
the ease of IV and oral administration of the drugs. Further, the
drugs are not metabolized in the body to any great extent.
[0027] The subjects as used herein are mammals, including
humans.
[0028] The invention also provides a composition comprising an
NSAID (non-steroidal anti-inflammatory drug) together with a GABA
analog. The NSAID will be present in an anti-inflammatory amount,
preferably somewhat less than normally used, and the GABA analog
will be present in a cytoprotective amount, namely an amount which
will be effective in preventing or reducing the gastrointestinal
damage otherwise caused by the NSAID. In general, the NSAID will be
present for doses of about 10 to about 500 mg, and the GABA analog
will be present at about 1 to about 1500 mg. Any NSAID can be
combined with any GABA analog according to this invention.
Preferred GABA analogs to be employed are the compounds of Formulas
I and II, especially gabapentin and pregabalin. Preferred NSAIDs to
be employed in the compositions include sulindac, naproxen,
indomethacin, mefenamic acid, diclofenac, fenoprofen, diflunisal,
etodolac, ibuprofen, piroxicam, acetylsalicylic acid, oxaprozin,
and bromfenac. Most of the NSAIDs to be used are commercially
available, generally as salts such as calcium, sodium, or
potassium, for example, fensprofen calcium and bromfenac sodium.
Especially preferred combinations include pregabalin or gabapentin,
together with naproxen sodium or ibuprofen. The compositions may
contain common pharmaceutical excipients such as those described
above.
[0029] The ability of GABA analogs to treat gastrointestinal
diseases according to this invention has been established in
several animal models of induced gastric lesions and
alcoholism.
BRIEF DESCRIPTION OF FIGURES
[0030] FIG. 1 shows the effect of gabapentin on gastric lesions
caused by indomethacin.
[0031] FIG. 2 shows the effect of gabapentin on handling responses
following withdrawal of chronic ethanol treatment.
[0032] FIG. 3 shows the effect of gabapentin on memory and
drowsiness in animals receiving chronic ethanol treatment.
[0033] FIG. 4 shows the effects of gabapentin, CI-1008
(pregabalin), and morphine on colonic allodynia.
[0034] FIG. 5 shows the effects of gabapentin and CI-1008 on
colonic pain threshold in rats.
EXAMPLE 1
[0035] Gabapentin was evaluated in animals to determine its ability
to prevent gastric lesions otherwise caused by indomethacin.
[0036] Male Sprague-Dawley rats weighing 240 to 250 g were fasted
for 24 hours and allowed free access to water prior to experiment.
All test drugs were given intragastrically. Rats were pretreated
with different doses of gabapentin at doses of 40 and 60 mg. Thirty
minutes later indomethacin (25 mg/kg) was administered. Another
group of rats received 10 mg of gabapentin twice, 3 hours apart,
followed by indomethacin administration. Three hours after
indomethacin treatment, the rats were killed and gastric lesions
were assessed. The severity of the lesions were determined by the
measurement of the square area (mm.sup.2) of visible lesions.
[0037] Results
[0038] 1. Indomethacin caused severe gastric hemorrhagic injury;
the areas of injury were measured at 42.6.+-.5.2 mm.sup.2
(mean,.+-.standard error of mean).
[0039] 2. Gabapentin pretreatment significantly reduced
indomethacin-induced gastric injury. The gastric lesion with
different doses of gabapentin pretreatment after indomethacin
treatment were measured: 22.3.+-.2.8 mm.sup.2 with 40 mg,
16.5.+-.2.2 mm.sup.2 with 60 mg/kg, and 4.2.+-.0.39 mm.sup.2 with
10 mg twice.
[0040] 3. Gabapentin pretreatment also dramatically reduced gastric
bleeding.
[0041] The foregoing data are presented in FIG. 1, where the first
bar is control (animals treated with indomethacin alone); Bar 2 is
for animals treated with one dose of 40 mg of gabapentin; Bar 3 is
for animals dosed one time with 60 mg of gabapentin; and Bar 4 is
for animals dosed two times with 10 mg of gabapentin.
EXAMPLE 2
[0042] To determine the effects of gabapentin on ethanol-induced
gastric lesions, rats were pretreated with 1 mL of 70% aq. ethanol
(v/v), followed by gabapentin at a dose of 40 and 60 mg,
respectively. Another group of rats were pretreated by 20 mg of
gabapentin given twice, 3 hours apart, followed by ethanol
administration. Thirty minutes later, all rats were killed and
gastric lesions were assessed.
[0043] Results
[0044] 1. Seventy percent ethanol induced significant gastric
injury. The area of injury measured 41.9.+-.3.7 mm.sup.2.
[0045] 2. Gabapentin pretreatment reduced ethanol-induced gastric
injury. With 2 doses of 20 mg gabapentin pretreatment, the area of
injury measured 2.1.+-.0.3 mm.sup.2. With 40 and 60 mg of
gabapentin pretreatment, the area of injury measured 24.4.+-.3.5
mm.sup.2 and 18.7.+-.2.2 mm.sup.2, respectively.
EXAMPLE 3
[0046] The following test in rats was carried out to further
establish the ability of gabapentin and Pregabalin to reduce the
gastrointestinal damage caused by NSAIDS.
[0047] Animals:
[0048] Male CD-Sprague-Dawley rats (132-202 g) were received and
housed in a room with controlled temperature, humidity, and 12-hour
light/dark cycle. After a period of acclimatization of 4 to 5 days,
and after a 24-hour food fasting period, animals were used for the
study described below.
[0049] Administration of GABA Analogs and Indomethacin:
[0050] Gabapentin or pregabalin (CI-1008) were dissolved in water
and administered orally at the following doses: 1, 10, 100, and 200
mg/kg in a volume of 1 mL. Control animals were dosed with an equal
volume of vehicle (1 mL of water). Sixty minutes later, all the
animals received 1 mL of a solution of indomethacin dissolved in 5%
aqueous NaHCO.sub.3 (80 mg/kg). Control animals received 1 mL of 5%
aqueous NaHCO.sub.3 orally. Experimental groups were as
follows:
1 Group Pretreatment Damaging Agent Group 1 Water None
(NaHCO.sub.3) Group 2 Water Indomethacin 80 mg/kg Group 3
Gabapentin 1 mg/kg Indomethacin 80 mg/kg Group 4 Gabapentin 10
mg/kg Indomethacin 80 mg/kg Group 5 Gabapentin 100 mg/kg
Indomethacin 80 mg/kg Group 6 Gabapentin 200 mg/kg Indomethacin 80
mg/kg Group 7 Pregabalin 1 mg/kg Indomethacin 80 mg/kg Group 8
Pregabalin 10 mg/kg Indomethacin 80 mg/kg Group 9 Pregabalin 100
mg/kg Indomethacin 80 mg/kg Group 10 Pregabalin 200 mg/kg
Indomethacin 80 mg/kg
[0051] Evaluation of the Effect:
[0052] Gastric damage caused by indomethacin correlates with
inhibition of the cyclooxygenase product prostaglandin E2
(PGE.sub.2). Animals were sacrificed by decapitation 4 hours
post-indomethacin administration. The stomach was removed and
opened along the greater curvature and its image digitized and
stored on an optical disk using a 486-based PC computer equipped
with CUE3 system imaging analysis software (Olympus Corp.,
Marietta, Ga., USA). Two 6-mm biopsies were taken from a constant
region of the gastric mucosa located in each side of the glandular
portion of the stomach, and their PGE.sub.2 content was measured
using a commercially available ELISA kit (Assay Designs Inc., Ann
Arbor, Mich., USA). The presence of gastric damage was determined
using the retrieved electronic image, and the extent of damage was
measured using the CUE3 imaging software. Data are expressed as a
percent of gastric area damaged and the PGE.sub.2 content (pg/mL).
The data are present below in Table 1.
2TABLE 1 % Area PGE.sub.2 Synthesis Compound Drug Dose N Damaged
(pg/mL) Control NA 10 0.00 .+-. 0.00* 9525.27 .+-. 156.00*
Indomethacin + Vehicle NA 9 5.56 .+-. 0.48 1908.03 .+-. 72.31
Indomethacin + Gabapentin 1 mg/kg 5 2.99 .+-. 0.46 1783.66 .+-.
73.47 Indomethacin + Gabapentin 10 mg/kg 9 3.96 .+-. 0.35 3065.78
.+-. 137.19 Indomethacin + Gabapentin 100 mg/kg 10 1.87 .+-. 0.1
2997.90 .+-. 226.80 Indomethacin + Gabapentin 200 mg/kg 5 1.43 .+-.
0.40 2615.74 .+-. 165.36 Indomethacin + Cl-1008 1 mg/kg 4 8.07 .+-.
2.19 1209.95 .+-. 105.50 Indomethacin + Cl-1008 10 mg/kg 10 4.07
.+-. 0.42 2666.16 .+-. 307.45 Indomethacin + Cl-1008 100 mg/kg 10
1.99 .+-. 0.25* 3994.45 .+-. 318.95 Indomethacin + Cl-1008 200
mg/kg 5 0.34 .+-. 0.06* 3288.92 .+-. 407.43 Values are average .+-.
standard error. *p < 0.05 based on Kruskal-Wallis one-way
analysis of variance followed by Dunn's test and compared to
indomethacin group.
[0053] Both gabapentin and CI-1008 caused a reduction of the amount
of gastric damage induced by indomethacin which, in the case of
CI-1008, reached statistical significance at doses of 100 and 200
mg/kg.
[0054] As expected, the gastric damage caused by indomethacin was
associated with a significant inhibition of the cyclooxygenase
product PGE.sub.2. Neither gabapentin nor CI-1008, at any dose
tested, were able to significantly modify this effect. This data
suggests that the significant reduction of the indomethacin-induced
gastric damage caused by CI-1008 is not related to an effect of
this GABA analog on the cyclooxygenase enzyme.
[0055] The foregoing data establish that GABA analogs such as
gabapentin and pregabalin are effective in preventing
gastrointestinal damage such as gastric lesions, peptic ulcers, and
even lower gastrointestinal bleeding, otherwise caused by
consumption of alcohol or NSAIDs. The GABA analogs also treat the
effects of alcohol withdrawal, which is a syndrome characterized by
tremor, hallucinations, and confusion, and general gastrointestinal
disorders such as IBD and IBS.
[0056] The following tests establish that GABA analogs are useful
to treat ethanol withdrawal syndrome.
EXAMPLE 4
[0057] Male albino mice of the outbred TO strain (Bantin and
Kingman, UK) were used in all studies. The weight ranged from 25 to
35 g, with no more than a 5 g range in any single experiment. The
mice were housed, eight per cage, at 21.degree. C..+-.1.degree. C.,
with 55.+-.10% relative humidity, and a 12-hour light/dark cycle
with the light phase between 09:00 to 21:00. All mice received ad
libitum access to tap water and standard laboratory chow (RM-1,
Special Diet Services, UK) until their use in experiments or until
their diet was replaced with a liquid diet.
[0058] Induction of Physical Dependence
[0059] Ethanol was administered in a liquid diet schedule. All mice
received control diet for an initial 2-day period. Ethanol treated
mice then received a diet containing 3.5% (v/v) ethanol/water for 2
days, followed by a diet containing 7% ethanol for a further 5
days. The average intake was 22 to 30 g/kg/day. Control groups were
pair-fed a control diet, balanced isocalorifically to match the
ethanol containing diet. There were no differences in the weights
of the ethanol-treated and control mice at the end of the treatment
periods. When mice were withdrawn from the ethanol (between 07:00
AM and 09:00 AM), they were provided with tap water until their use
in experiments.
[0060] Drug Treatment
[0061] Gabapentin was dissolved in saline, the solution being made
freshly each testing day. Intraperitoneal (i.p.) injections of
either gabapentin, 10 mL/kg, or saline, were given immediately on
withdrawal from the ethanol treatment in the studies on the
handling responses, and 2 hours prior to measurement of audiogenic
seizures. In the experiments using a standard elevated plus maze,
gabapentin or saline was injected i.p. at 8 hours after the removal
of the ethanol diet, and the mice were placed on the plus maze 60
minutes after the injections. In the studies on motor co-ordination
(ataxia) and on locomotor activity, gabapentin or saline was
injected into ethanol-naive chow fed mice immediately before
testing. Measurements were then made for 60 minutes for the ataxia
study, and for 30 minutes in the case of the locomotor
activity.
[0062] Measurement of Handling-Induced Behavior
[0063] Following withdrawal from the ethanol treatment at 09:00 AM,
ratings of handling-induced behavior were assessed by the same
experimenter, on the same mice, every hour for a period of 12 hours
after withdrawal from ethanol. Numerical ratings have the
definitions shown in Table 2.
3TABLE 2 Behavioral Ratings During Gentle Handling 1 Mild tremor on
lifting and turning 2 Continuous severe tremor on lifting and
turning 3 Clonic forelimb extensor spasm on lifting 4 Clonic
forelimb extensor spasm on lifting, which continued after placing
mouse on cage top 5 Spontaneous evidence of myoclonic activity
followed by (4)
[0064] Each mouse was lifted gently by the tail and held for 3
seconds, 30 cm under an "Anglepoise lamp" with a 60-watt bulb. The
animal was gently rotated and its ensuing behavior rated on a scale
of 1 to 5 according to the criteria in Table 2. Groups of 15 mice
were used in each of the treatment groups, and the data were
calculated as medians with interquartile ranges. The data were also
expressed as the area under the curve at 4 and 12 hours from the
withdrawal of the ethanol treatment. The results are shown in FIG.
2.
[0065] Elicitation of Audiogenic Seizures
[0066] At 8 and 12 hours from ethanol withdrawal, the
susceptibility to sound-induced convulsions was measured in
separate groups of ten mice. Mice were tested individually in a
sound-proof perspex box 30.times.30.times.30 cm containing an
electric door-bell. The bell was rung for 2 minutes or until the
first signs of convulsions. The number of mice which responded by
wild-running and clonic convulsions was counted. The mice were
humanely killed as soon as a full convulsion was seen.
[0067] Anxiety-Related Behavior
[0068] Mice were withdrawn from the ethanol diet at 7:00 AM and
tested for anxiety-related behavior 8 hours later using a murine
elevated plus-maze. It was constructed of perspex with two opposing
open arms (30.times.5.times.0.25 cm) and two opposing closed arms
(30.times.5.times.15 cm) which extended from a central platform
(5.times.5 cm). The floor was of matt black perspex. The animals
were acclimatized to the experimental room I hour prior to
experimentation. Experiments were conducted under dim red light,
and each 5 minute session was video-taped for later analysis, by an
observer unaware of the prior treatment. During this analysis
(Observer 3.0, Noldus Information Technology, Wageningen,
Netherlands) measurements were made of the time spent on each arm
of the maze, the number of entries onto each arm and rearing
activity. The measurements were made in accordance with the
definitions in Table 3.
4TABLE 3 Measurements of Behavior on the Elevated Plus Maze Arm
entry = All four paws onto either a closed or an open arm. "Head
Dip" = An exploratory forward head/shoulder movement over the side
of an open arm and down towards the floor. "Protected head dips" =
Exploratory forward head/shoulder movement over the side of a
closed arm and down towards the floor. "Stretch-attend posture" =
An exploratory flat body posture where the mouse stretches forward
and then retracts to original position without moving forward.
[0069] Measurement of Ataxic Actions
[0070] Possible ataxic effects of gabapentin were studied in
control animals (i.e., not treated with ethanol) by the rotorod
method. Mice were placed on a rod rotating at 4.5 rpm, and the time
they remained on the rod was measured automatically. A cut-off time
of 180 seconds was used in all experiments. Before the acute drug
injections, all mice were tested on the rotorod to ensure that they
stayed on for 180 seconds (a very small number did not do so and
were excluded from the studies). Measurements were made for 60
minutes, at 10-minute intervals, after the acute administration of
the drug under test. Eight mice were used in each treatment
group.
[0071] Locomotor Activity
[0072] The effects of gabapentin in control animals were also
tested on locomotor activity to determine the selectivity of the
effects in the withdrawal studies. Mice were injected with
gabapentin solution or saline and placed immediately in activity
test cages crossed by infra-red beams. The number of infra-red beam
breaks was measured every 5 minutes for the next 30 minutes.
Rearing activity was measured by a similar set of infrared beams
situated 4 cm above the cage floor.
[0073] Statistical Analysis
[0074] The results of the handling response ratings were compared
by nonparametric two-way analysis of variance, designed for repeat
measures on the same animal. The results of the area under the
curve calculations were compared by the Mann-Whitney U-test. The
convulsion incidence was analyzed by Fisher's exact probability
test. The measurements from the elevated plus maze were subjected
to one-way analysis of variance, followed by a Bonuferroni multiple
comparison test, comparing all groups to the control group which
received saline injections, and also comparing both ethanol treated
groups which received gabapentin to the ethanol treated group which
received saline. The ataxia measurements were analyzed by the
Mann-Whitney U-test and the locomotor activity by Student's
t-test.
RESULTS
[0075] Handling Response
[0076] The ratings of behavior in response to gentle handling
showed the expected increase following withdrawal from the ethanol
treatment. Gabapentin (GP), at 100 mg/kg (FIG. 2a), significantly
reduced this increase in ratings when the results were compared
over the 12-hour testing period (p<0.001). The effect of this
dose of gabapentin showed a marked reduction in handling scores for
around 4 hours. This time period was therefore used in later
analysis to examine the area under the handling curve with each
dose of the drug. The effects of lower doses of gabapentin were not
significant over the 12-hour period of measurement, but when the
areas under the curve were calculated for the first 4 hours of the
study (FIG. 2b), significant effects of the 20- and 50-mg/kg doses
were seen (p<0.05), as well as the 100-mg/kg dose
(p<0.01).
[0077] Audiogenic Seizures
[0078] At the 8-hour time interval, 50 and 100 mg/kg gabapentin
decreased the convulsion incidence after the audiogenic stimulus,
with the 100-mg/kg dose reaching statistical significance
(p<0.05). There was no effect of the lower doses (Table 4). No
effect was seen of any of the doses tested at 12 hours from the end
of the ethanol treatment (data not shown).
5TABLE 4 The Effect of Gabapentin on Audiogenic Convulsions
Measured 8 Hours From Ethanol Withdrawal Chronic Percentage of
Group Showing Treatment Acute Injection Clonic Convulsions Control
Diet Saline 0 Ethanol Diet Saline 80* p < 0.05 c.f.
Control/Saline group Ethanol Diet Gabapentin 5 mg/kg 92 Ethanol
Diet Gabapentin 20 mg/kg 70 Ethanol Diet Gabapentin 50 mg/kg 40
Ethanol Diet Gabapentin 100 mg/kg 30 p < 0.01 c.f.
Ethanol/Saline group
[0079] Elevated Plus Maze
[0080] The most prominent effect of ethanol withdrawal in this test
was a decrease in the percentage time spent on the open arms of the
maze (FIG. 3a, F(4,50)=5.12, p<0.002). Gabapentin decreased this
effect at both 50 and 100 mg/kg. The p values were p<0.05 for
the 50-mg/kg dose and p<0.01 for 100 mg/kg, for comparison with
saline administration in both cases.
[0081] Mice undergoing ethanol withdrawal also showed a significant
increase in head dips from the closed arms (protected head dips).
This effect was significantly reduced by gabapentin at 100 mg/kg
(p<0.01 compared with the effects of saline), as illustrated in
FIG. 3b (F(4,50)=6.53, p<0.001). In control animals, the number
of protected head dips was significantly decreased by gabapentin at
100 mg/kg (p<0.05, compared with control values after saline
administration). Although the mean time on the open arms was
increased in control animals after this dose of gabapentin, this
was not significantly different from controls with saline.
EXAMPLE 5
[0082] LPS-Colonic Hypersensitivity Assay
[0083] The GABA analogs also have been evaluated for their ability
to control and treat gastrointestinal disorders characterized as
IBD and IBS. The assay utilized to evaluate the GABA analogs
measures the effects of compound on lipopolysaccharide-induced
delayed rectal allodynia in rats. Intraperitoneal (IP) injections
of the endotoxin lipopolysaccharide (LPS) are known to induce
long-lasting hyperalgesia in somatic pain models. The following
assay LPS-colonic hypersensitivity assay was designed to evaluate
the effect of IP injections of LPS on pain visceral threshold in an
experimental model of rectal distension.
[0084] Animal Preparation
[0085] Male Wistar rats weighing 250 to 350 g were surgically
prepared for electromyography, according to a standard technique.
Rats were anesthetized by i.p. injection of acepromazine and
ketamine (Imalgene 1000, Rhone-Mrieux, Lyon, France) at doses of
0.6 and 120 mg/kg, respectively. Two groups of four electrodes of
nichrome wire (60 cm length and 80 .mu.m diameter) were implanted
bilaterally in the abdominal external oblique musculature just
superior to the inguinal ligament. Electrodes were exteriorized on
the back of the neck and protected by a glass tube attached to the
skin. Animals were individually housed in polypropylene cages and
kept in a temperature-controlled room (21.degree. C.). They were
allowed free access to water and food (UAR pellets, Epinay,
France).
[0086] Electromyographic Recording
[0087] Electromyographic recording began 5 days after surgery. The
electrical activity of abdominal striated muscles was recorded with
an electroencephalograph machine (Mini VIII, Alvar, Paris, France)
using a short-time constant (0.03 sec) to remove low-frequency
signals (<3 Hz) and a paper speed of 3.6 cm/minute.
[0088] Balloon Distension Procedure
[0089] Rats were placed in plastic tunnels (6 cm diameter; 25 cm
length) where they could not move, escape or turn around, in order
to prevent damage to the balloon. They are accustomized to this
procedure for 3 or 4 days before rectal distension (RD) in order to
minimize stress reaction during experiments. The animals were
determined to be accustomed to the plastic tunnel using two
criteria: (i) a behavioral component: when the animals tried to
escape or turn around no more than one time per 5 minutes, (ii) the
abdominal basal activity: when abdominal striated muscles exhibited
less than five abdominal contractions per 5 minutes in the absence
of distension. The balloon used for distension was an arterial
embolectomy catheter (Fogarty, Edwards Laboratories, Inc., Santa
Ana, USA). Rectal distension (RD) was performed by insertion of the
balloon (2-mm diameter; 2-cm long) in the rectum, at 1 cm of the
anus, the catheter being fixed at the tail. It was inflated
progressively by steps of 0.4 mL, from 0 to 1.6 mL, each step of
inflation lasting 5 minutes. To detect possible leakage, the volume
of water introduced in the balloon was checked by complete removal
with a syringe at the end of the distension period.
[0090] Experimental Protocol
[0091] In a first series of experiments, a group of 8 rats were
submitted to gradual rectal distention. The animals were previously
(30 minutes) treated by gabapentin at doses of 30 and 100 mg/kg IP
or its vehicle (NaCl 9% aqueous).
[0092] In a second series of experiments, the same group of 8 rats
received IP lipopolysaccharide (E Coli, serotype 0111:B4) or its
vehicle, at a dose of 1 mg/kg IP, 1 hour after a control rectal
distension. Then, RD was performed 12 hours after LPS injection and
was preceded (30 minutes) by IP administration of gabapentin (30
mg/kg) or its vehicle (0.3 mL/rat).
[0093] Drugs
[0094] LPS was dissolved in saline (NaCl 9%). Intraperitoneal
injection of vehicle was given in a volume of 0.3 mL. LPS was
purchased from Sigma-Aldrich (St. Quentin Fallavier, France).
[0095] Statistical Analysis
[0096] Statistical analysis of the number of abdominal contractions
occurring during each 5-minute period during RD was performed by
one-way ANOVA followed by Student's paired t-test. Values were
expressed as the mean.+-.SEM, and differences were considered
significant for p<0.05.
[0097] The results are presented in Tables 5 and 6, and establish
that gabapentin is effective in reducing lower gastrointestinal
disorders such as IBS.
6TABLE 5 Effect of Gabapentin on Abdominal Response Induced by
Rectal Distension (Number of abdominal contractions/5 minutes; mean
.+-. SEM, n = 7-8, *p < 0.05, **pp < 0.01, significantly
different from vehicle; n % of reduction vs vehicle) Volume of
Vehicle Gabapentin Vehicle Gabapentin Distension (0.3 mL/rat) (30
mg/kg) (0.3 mL/rat) (100 mg/kg) 0.4 mL 4.4 .+-. 1.6 5.0 .+-. 2.1
3.9 .+-. 1.8 2.0 .+-. 1.4 0.8 mL 19.1 .+-. 2.8 10.6 .+-. 34** 19.6
.+-. 2.3 7.6 .+-. 3.4** (-45%) (-61.2%) 1.2 mL 23.4 .+-. 2.6 16.1
.+-. 2.3* 19.1 .+-. 2.3 16.7 .+-. 2.9 (-31.2%)
[0098]
7TABLE 6 Effect of Gabapentin on LPS-Lnduced Delayed (12 Hours)
Allodynia (Number of abdominal contractions/5 minutes; mean .+-.
SEM, n = 7-8, +p < 0.001, significantly different from
"LPS/vehicle" value; n % of reduction vs "LPS/vehicle") LPS (1
mg/kg) + LPS (1 mg/kg) + Volume of Vehicle Gabapentin Distension
(0.3 mL/rat) (3.0 mg/kg) 0.4 mL 9.7 .+-. 1.0 0.7 .+-. 0.5 +
(-92.8%) 0.8 mL 11.7 .+-. 1.2 11.9 .+-. 0.8 1.2 mL 23.5 .+-. 2.2
16.3 .+-. 3.2
[0099] The foregoing experiment was carried out with the
GABA-analog pregabalin. Pregabalin, at 30 mg/kg, reduced the number
of cramps at both distension volumes of 0.4 and 0.8 mL. When
injected 120 minutes before rectal distension, pregabalin, at both
10 and 30 mg/kg, had a similar effect at all distension volumes.
LPS enhanced the number of abdominal contractions at the volume of
0.4 mL (9.7.+-.1.0 vs. 3.7.+-.1.0) 12 hours after its
administration. This effect was suppressed when animals received
pregabalin (1.8.+-.0.9 vs. 9.7.+-.1.0) at 30 mg/kg 30 minutes prior
to rectal distension. These results establish that pregabalin is
effective in reducing basal rectal sensitivity and in blocking
LPS-induced rectal allodynia in rats.
EXAMPLE 6
[0100] TNBS-Induced Allodynia
[0101] GABA analogs were evaluated in rats suffering from chronic
visceral allodynia induced by trinitrobenzene sulfonic acid (TNBS).
Injections of TNBS into the colon of animals have been found to
induce chronic colitis. In humans, digestive disorders are often
associated with visceral pain. In these pathologies, the visceral
pain threshold is decreased, indicating a visceral
hypersensitivity. Consequently, the following study was designed to
evaluate the effect of injection of TNBS into the colon on visceral
pain threshold in an experimental model of colonic distension.
[0102] Male Sprague-Dawley rats weighing 340 to 400 g were used in
the study. The animals were housed three per cage in a regulated
environment (20.+-.1.degree. C., 50.+-.5% humidity, with light 8:00
AM to 8:00 PM). Under anethesia (ketamine 80 mg/kg i.p.;
acepromazin 12 mg/kg i.p.), TNBS (50 mg/kg), or saline (1.5 mL/kg)
was injected into the proximal colon (1 cm from the cecum). After
the surgery, animals were individually housed in the regulated
environment.
[0103] A balloon catheter (5-6 cm length) was inserted through the
anus into the colon and kept in position (tip of balloon 5 cm from
the anus) by taping the catheter to the base of the tail. The
balloon was progressively inflated by step of 5 mm Hg, from 0 to 75
mm Hg, each step of inflation lasting 30 seconds. Each cycle of
colonic distension was controlled by a standard barostat. The
threshold corresponds to the pressure which produced the first
abdominal contraction, at which time the cycle of distension was
discontinued. To determine the colonic threshold, four cycles of
distension were performed on the same animal.
[0104] In a first series of experiments, a group of eight rats
treated with saline were subjected to a colonic distension
session.
[0105] In a second series, a group of eight rats treated with TNBS
were subjected to a colonic distension session.
[0106] In a third series, a group of eight rats treated with TNBS
received a subcutaneous (sc) injection of gabapentin or CI-1008 30
minutes prior to initiation of the colonic distension cycle.
[0107] All test compounds were dissolved in saline except TNBS.
TNBS was dissolved in EtOH 30% (w/v). Subcutaneous injection of
vehicle was given in a volume of 2 mg/kg.
[0108] Statistical significance between each group was determined
using a one-way ANOVA followed by Student's unpaired t-test.
Differences were considered statistically significant at
p<0.05.
[0109] Pain threshold (pressure of distension inducing the first
abdominal contraction) after distal colonic distension was
determined at Day 7 in two groups of awake rats: control animals
and TNBS-treated animals. A significant decrease in the pain
threshold was observed in TNBS-treated animals. Inflammatory
parameters (colon weight, area of hyperemia and necrosis and
colonic myeloperoxidase content) were measured in the proximal
colon at Day 7 after TNBS treatment. All the parameters were
significantly increased except the area of necrosis.
[0110] Gabapentin (100, 300, and 500 mg/kg sc) and CI-1008 (30, 60,
100, and 200 mg/kg sc) were administered 30 minutes before colonic
distension and measurement of the inflammatory parameters.
Gabapentin inhibited in a dose-related manner the TNBS-induced
colonic allodynia. At 500 mg/kg sc, gabapentin completely blocked
the effect of TNBS on colonic pain. CI-1008 also showed a
dose-related inhibition of the decrease in pain threshold. At 100
mg/kg, CI-1008 completely suppressed the allodynia induced by TNBS.
Morphine (0.1 mg/kg sc) completely suppressed the TNBS-induced
decrease in pain threshold after colonic distension (FIG. 4). In
contrast, neither gabapentin nor CI-1008 inhibited the colonic
inflammatory effect of TNBS in these experimental conditions.
[0111] In normal conditions (control animals), morphine (0.3 mg/kg
sc) significantly increased the colonic pain threshold while, in
the same conditions, neither gabapentin (500 mg/kg sc) nor CI-1008
(200 mg/kg sc) modified the colonic pain threshold (FIG. 5). The
results are further shown in Tables 7 and 8.
8TABLE 7 Effect of CI-1008, Gabapentin, and Morphine on
TNBS-Induced Chronic Colonic Allodynia in Rats Colonic Threshold
Treatment (mm Hg) SEM n p Control 43.39 .+-. 1.98 8 Sham 33.44 .+-.
3.25 8 * TNBS 17.81 .+-. 1.27 8 *** CI-1008 30 mg/kg sc 21.72 .+-.
1.51 8 ? 60 mg/kg sc 25.47 .+-. 1.03 8 100 mg/kg sc 33.13 .+-. 1.83
8 200 mg/kg sc 40.47 .+-. 3.75 8 Gabapentin 100 mg/kg sc 22.03 .+-.
2.23 8 300 mg/kg sc 24.69 .+-. 1.27 8 ? 500 mg/kg sc 36.88 .+-.
1.46 8 Morphine 0.1 mg/kg sc 34.22 .+-. 1.72 8 0.3 mg/kg sc 46.09
.+-. 1.43 8 1 mg/kg sc 64.84 .+-. 1.88 8 * = p < 0.05, ** = p
< 0.01, and *** = p < 0.001 vs control. ? = p < 0.05, = p
< 0.01, and = p < 0.001 vs TNBS.
[0112]
9TABLE 8 Effect of CI-1008 and Gabapentin on Colonic Threshold in
Normal Rats Colonic Threshold Treatment (mm Hg) SEM n p Control
43.33 .+-. 1.23 6 CI-1008 200 mg/kg sc 46.41 .+-. 2.26 8 NS
Gabapentin 500 mg/kg sc 43.75 .+-. 1.44 6 NS NS = Not significant
vs control.
[0113] The foregoing data establish that GABA analogs such as
gabapentin and CI-1008 suppress TNBS-induced colonic allodynia, and
are therefore effective in abnormal colonic hypersensitivity
reflecting the chronic pain in IBS.
EXAMPLE 7
[0114] Formalin-Induced Inflammatory Colonic Pain
[0115] The GABA analogs were evaluated in another model to
determine their effect on inflammatory visceral pain, including
pancreatitis and intestinal cystitis.
[0116] Administration of formalin into the wall of the rat colon
causes acute inflammation and visceral pain. The aim of this study
was to evaluate the antinociceptive activity of gabapentin and
CI-1008 in visceral pain induced by colonic intrapariental
injection of formalin.
[0117] Adult female Sprague-Dawley rats weighing 240 to 260 g were
used in the study. The animals were housed three per cage in a
regulated environment (20.+-.1.degree. C., 50.+-.5% humidity, with
light 8:00 AM to 8:00 PM) prior to use in the test.
[0118] Each test animal was placed in a transparent plastic cage
(27.times.43.times.28 cm) with a layer of wood shavings on the
floor. Drinking water was available. Cages were placed in such a
way that visual interaction between animals was avoided. A mirror
was positioned behind each cage to improve the recording of
behaviors. Each animal was initially allowed 20 minutes to get used
to its surroundings. Anesthesia was then achieved by using
isofurane (starting 4%, then 1.5% in a mixture of 2:3 nitrous oxide
and 1:3 oxygen). The animal was suspended by its tail, the rectum
was gently emptied with a cotton-capped stalk, and a coloscope was
inserted through the anus. This designed endoscope has a lateral
slot that allows for puncture of the intestinal wall under visual
control, using a 51-mm-long needle (26 ga), at about 35 mm from the
anal margin. Injected solutions were either 50 .mu.L 5% aq.
formalin (v/v), or the same volume of isotonic saline. Animals were
allowed to recover from anesthesia as soon as the injection was
completed (about 1 minute), and the observation period was started
and continued for 2 hours. Thirty minutes after the end of the
observation test, an IV injection of Evans Blue (1%) was
administered, and 30 minutes later the animal was sacrificed. The
abdomen was opened. The injection site and the zone of diffusion of
Evans Blue were recorded by image analysis software. Data from rats
in which the spread of the dye was not restricted to the sigmoid
wall were discarded.
[0119] As listed in increasing order of pain intensity, these
behaviors were: (i) abdominal licking and nibbling (L), (ii) body
stretching, i.e., backward extension of the hind limbs (B), (iii)
contraction of the flanks, sometimes evolving to a stretching
attitude (C), and (iv) whole body contraction, the rat standing
with its back curved, occasionally further graded according to the
duration of the given episode: W.sub.1 for less than 30 seconds,
W.sub.2 between 30 seconds and 1 minute, and W.sub.3 for more than
1 minute. Behaviors were recorded for each animal throughout the
2-hour test on individual charts. A pain score (S) was then
calculated for each of the successive 15-minute periods, using the
following formula:
S=1L+2B+3C+4W.sub.1+5W.sub.2+6W.sub.3
[0120] whereby the pain score was proportional to (i) the number of
episodes of each selected behavior, and (ii) the coefficient, from
1 to 6, attributed to the given behaviors.
[0121] All compounds were dissolved in saline. Subcutaneous
injection of vehicle was given in a volume of 2.5 mg/kg. Formalin
was purchased from Prolabo.
[0122] Statistical significance between each group was determined
by using a one-way ANOVA followed by Student's unpaired t-test.
Differences were considered statistically significant at
p<0.05.
[0123] Hyperalgesia is induced by intramural injection of formalin
(5%, 50 .mu.L/rat) into the colonic wall in unfasted female
Sprague-Dawley rats. Gabapentin and CI-1008 were tested at 100,
300, 500 and 100, 200 mg/kg sc, respectively. Gabapentin and
CI-1008 significantly and dose-dependently decreased the pain score
induced by intracolonic formalin. The maximal inhibitory effect was
observed after 500 mg/kg of gabapentin and 200 mg/kg of CI-1008.
The results are presented in Table 9.
[0124] This study establishes that GABA analogs exhibit an
antinociceptive effect on intra-colonic formalin-induced pain, and
thus are effective in treating IBD and IBS, and visceral pain,
including pancreatitis and intestinal cystitis.
10TABLE 9 Effect of Subcutaneous Injection of Gabapentin and
CI-1008 on Inflammatory Colonic Pain Induced by Intramural
Injection of Formalin 5% Treatment % Antinociception SEM n p
CI-1008 100 mg/kg sc 18.55 .+-. 7.41 7 *** 200 mg/kg sc 70.81 .+-.
7.47 6 *** Gabapentin 0.3 mg/kg sc -7.73 .+-. 10.43 3 NS 100 mg/kg
sc 13.62 .+-. 12.65 9 NS 300 mg/kg sc 55.07 .+-. 9.98 6 *** 500
mg/kg sc 88.01 .+-. 16.96 6 *** *** = NS = Not significant vs
control.
[0125] The following examples further illustrate compositions
provided by the invention which contain a GABA analog in
combination with an NSAID.
EXAMPLE 8
[0126]
11 Tablet Formulation Naproxen sodium 200 mg Gabapentin 300 mg
Magnesium stearate 20 mg Microcrystalline cellulose 100 mg Povidone
100 mg Talc 50 mg
[0127] The ingredients are blended to uniformity and pressed into a
tablet. The tablets are administered from 1 to 3 times a day for
treatment of inflammatory conditions such as rheumatoid arthritis,
ankylosing spondylitis, osteoarthritis, bursitis, tendinitis, and
acute gouty arthritis.
EXAMPLE 9
[0128]
12 Capsule Formulation Fenoprofen calcium, USP 150 mg Pregabalin 50
mg Cellulose 100 mg Gelatin 50 mg Titanium dioxide 10 mg Cornstarch
50 mg
[0129] The ingredients are blended to uniformity and placed into a
gelatin capsule. The capsules are administered from 1 to 4 times a
day for treatment of rheumatoid arthritis and osteoarthritis. The
combinations provided by this invention comprise an NSAID (eg,
naproxen or meclofenamic acid) and a GABA analog (eg, pregabalin or
gabapentin). Such combinations have been shown to be synergistic in
their ability to treat pain. For example, gabapentin and naproxen
sodium were combined in synergistic amounts and evaluated in a
standard rat carrageenan footpad thermal hyperalgesia assay. This
assay utilizes an extract of seaweed (carrageenan) that, when
injected into the footpad of test animals, causes a sterile
inflammation, thereby lowering the pain threshold. Analgesic
agents, including GABA analogs such as gabapentin, raise the pain
threshold back to normal, thereby enabling the animal to tolerate
an external source of pain for a longer period of time relative to
untreated control animals. Several fixed combinations of gabapentin
and naproxen sodium, ranging in concentrations of about 50 parts by
weight of GABA analog to 1 part by weight of NSAID, to 1:1
combinations, were evaluated in the foregoing assay. The results
are shown in FIG. 6 (for fixed 1:1 combinations at various dosages)
and in FIG. 7 (for fixed 50:1 combinations at various dosages). The
data establish that the combinations of a GABA analog and an NSAID
are synergistic in their ability to relieve acute and chronic pain
and to induce analgesia.
* * * * *