U.S. patent application number 09/325369 was filed with the patent office on 2002-02-07 for gel formulations for topical drug delivery.
Invention is credited to BEAURLINE, JOSEPH M., RODDY, PATRICK J., TOMAI, MARK A..
Application Number | 20020015715 09/325369 |
Document ID | / |
Family ID | 25057713 |
Filed Date | 2002-02-07 |
United States Patent
Application |
20020015715 |
Kind Code |
A1 |
BEAURLINE, JOSEPH M. ; et
al. |
February 7, 2002 |
GEL FORMULATIONS FOR TOPICAL DRUG DELIVERY
Abstract
Pharmaceutical gel formulations for topical drug delivery
include drug, colloidal silicon dioxide, triacetin and, preferably,
propylene glycol. The gel formulations are well suited for topical
delivery of the drug
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol, which when applied topically induces cytokines, such
as interferon and tumor necrosis factor, locally in the skin or
mucous membranes of a mammal. The gel formulations are also
well-suited for topical delivery of drugs for treatment of diseases
involving skin and/or mucosal lesions because the gel formulations
do not need to include irritating components.
Inventors: |
BEAURLINE, JOSEPH M.; (NORTH
ST PAUL, MN) ; RODDY, PATRICK J.; (MAHTOMEDI, MN)
; TOMAI, MARK A.; (OAKDALE, MN) |
Correspondence
Address: |
OFFICE OF INTELLECTUAL PROPERTY COUNSEL
3M INNOVATIVE PROPERTIES COMPANY
PO BOX 33427
ST PAUL
MN
551333427
|
Family ID: |
25057713 |
Appl. No.: |
09/325369 |
Filed: |
June 4, 1999 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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09325369 |
Jun 4, 1999 |
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08759992 |
Dec 3, 1996 |
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5939090 |
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Current U.S.
Class: |
424/401 ;
424/400 |
Current CPC
Class: |
Y10S 514/934 20130101;
A61K 9/0014 20130101; A61K 47/02 20130101; A61P 31/12 20180101;
Y10S 514/944 20130101; A61P 31/22 20180101; A61P 17/02 20180101;
A61K 47/10 20130101; A61K 47/14 20130101; A61K 31/4745 20130101;
A61P 17/00 20180101; Y10S 514/889 20130101 |
Class at
Publication: |
424/401 ;
424/400 |
International
Class: |
A61K 009/00; A61K
006/00; A61K 007/00; A61F 013/00 |
Claims
What is claimed is:
1. A gel formulation for the topical administration of a drug,
which formulation comprises a therapeutically effective amount of
drug, colloidal silicon dioxide, propylene glycol, and
triacetin.
2. The gel formulation of claim 1, wherein the colloidal silicon
dioxide is present in an amount of about 7 to about 12 percent by
weight based on the total weight of the formulation.
3. The gel formulation of claim 1, wherein the propylene glycol is
present in an amount of about 1 to about 30 percent by weight based
on the total weight of the formulation.
4. The gel formulation of claim 1, wherein the propylene-glycol is
present in an amount of about 5 to about 25 percent by weight based
on the total weight of the formulation.
5. The gel formulation of claim 1, wherein the triacetin is present
in an amount of about 58 to about 92 percent by weight based on the
total weight of the formulation.
6. A gel formulation for the topical administration of
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo
[4,5-c]quinoline-1-ethanol, which formulation comprises: a
therapeutically effective amount of
4-amino-2-ethoxymethyl-.alpha.,.alpha-
.-dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol; colloidal silicon
dioxide; and triacetin.
7. The gel formulation of claim 6, further including propylene
glycol.
8. A gel formulation according to claim 6, wherein
4-amino-2-ethoxymethyl--
.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol is
present in an amount of about 0.001 percent to about 0.6 percent by
weight based on the total weight of the formulation.
9. A gel formulation according to claim 6, wherein
4-amino-2-ethoxymethyl--
.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol is
present in an amount of about 0.01 percent to about 0.5 percent by
weight based on the total weight of the formulation.
10. A gel formulation according to claim 6, wherein the
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol is dissolved in the gel formulation.
11. A gel formulation according to claim 6, wherein the colloidal
silicon dioxide is present in an amount of about 7 to about 12
percent by weight based on the total weight of the formulation.
12. A gel formulation according to claim 6, wherein the colloidal
silicon dioxide is present in an amount of about 8 to about 10
percent by weight based on the total weight of the formulation.
13. A gel formulation according to claim 6, wherein the triacetin
is present in an amount of about 88 to about 93 percent by weight
based on the total weight of the formulation.
14. A gel formulation according to claim 7, wherein the triacetin
is present in an amount of about 58 to about 92 percent by weight
based on the total weight of the formulation.
15. A gel formulation for the topical administration of
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo
[4,5-c]quinoline-1-ethanol, which formulation comprises: (a) a
therapeutically effective amount of
4-amino-2-ethoxymethyl-.alpha.,.alpha-
.-dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol, (b) colloidal
silicon dioxide present in an amount of about 7 percent to about 12
percent by weight based on the total weight of the formulation; (c)
propylene glycol present in an amount of about 1 percent to about
30 percent by weight based on the total weight of the formulation;
and (d) triacetin present in an amount of about 58 percent to about
92 percent by weight based on the total weight of the
formulation.
16. A gel formulation according to claim 15, wherein the propylene
glycol is present in an amount of about 5 percent to about 25
percent by weight based on the total weight of the formulation.
17. A gel formulation according to claim 15, wherein
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol is present in an amount of about 0.001 percent to about
0.6 percent by weight based on the total weight of the
formulation.
18. A gel formulation according to claim 15 comprising from about
0.05 percent to about 0.3 percent by weight
4-amino-2-ethoxymethyl-.alpha.,.al-
pha.-dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol, from about 8
percent to about 10 percent by weight colloidal silicon dioxide,
and from about 90 percent to about 92 percent by weight
triacetin.
19. A gel formulation according to claim 15 comprising from about
0.05 percent to about 0.3 percent by weight
4-amino-2-ethoxymethyl-.alpha.,.al-
pha.-dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol, from about 8
percent to about 10 percent by weight colloidal silicon dioxide,
from about 15 percent to about 25 percent by weight propylene
glycol, and from about 65 percent to about 77 percent by weight
triacetin.
20. A method of inducing cytokines locally in the skin or mucous
membranes of a mammal, comprising placing on the skin or mucous
membranes of a mammal an amount of a gel formulation including
4-amino-2-ethoxymethyl-.a-
lpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol
effective to induce cytokines.
21. A method of inducing interferon locally in the skin or mucous
membranes of a mammal, comprising placing on the skin or mucous
membranes of a mammal an amount of a gel formulation including
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol effective to induce interferon.
22. A method of inducing tumor necrosis factor locally in the skin
or mucous membranes of a mammal, comprising placing on the skin or
mucous membranes of a mammal an amount of a gel formulation
including
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol effective to induce tumor necrosis factor.
23. The method of claim 20, where the gel formulation is applied to
lesions on the skin and/or mucosa.
Description
BACKGROUND OF THE INVENTION
[0001] This invention relates to improved pharmaceutical gel
formulations for the topical delivery of drugs. In another aspect,
this invention relates to pharmaceutical topical gel formulations
containing
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol.
[0002] Pharmaceutical gel and cream formulations for topical
delivery of drugs are well known. However, many such formulations
are not suitable for certain applications due to problems with, for
example, insolubility and/or degradation of the drug in the
formulation, physical instability of the formulation (separation of
components, thickening, precipitation/agglomerization of active
ingredient, and the like), and due to irritation of the skin or
mucosa to which the formulation is applied. Also, depending on the
purpose of the formulation, it may be desirable if the formulation
avoids systemic delivery of the active ingredient, particularly
where side effects may result from such systemic delivery.
[0003] U.S. Pat. No. 5,238,944 discloses a topical formulation of
1-isobutyl-1H-imidazo[4,5-c]quinolin-4-amine which is effective for
the treatment of genital warts and other diseases. However,
although useful for its intended purpose, this formulation is a
cream, subject to potential separation problems, and includes
isostearic acid which makes it painful if applied to open lesions
such as occur in the case of herpes simplex virus infection.
[0004] The compound
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imi-
dazo[4,5-c]quinoline-1-ethanol disclosed in U.S. Pat. No. 5,389,640
is from the same chemical class of compounds as
1-isobutyl-1H-imidazo[4,5-c]- quinolin-4-amine, although having
some significantly different chemical, physical, and biological
properties. The compound has been shown to induce interferon and
tumor necrosis factor in mice and rats following oral
administration. The compound has also been shown to induce
interferon-a, tumor necrosis factor, interleukin-1.alpha.,
interleukin-1.beta., interleukin-6 and interleukin-8 in cultures of
human peripheral blood mononuclear cells. The compound has also
shown antiviral activity against herpes simplex virus-challenged
guinea pigs when administered subcutaneously, dermally or
intravaginally 24 hours before infection.
[0005] However, systemic administration of
4-amino-2-ethoxymethyl-.alpha.,-
.alpha.-dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol may also be
associated with certain side effects, including fever, malaise,
headache, nausea and vomiting. Non-systemic topical cytokine
induction would thus have the advantage of avoiding the side
effects associated with the systemic induction of these
ctyokines.
SUMMARY OF THE INVENTION
[0006] Accordingly, it is one object of the present to provide a
highly stable pharmaceutical gel formulation that is suitable for
topical application to the skin and/or mucosa.
[0007] A related object is to provide a gel formulation that is
suitable for application to skin and/or mucosal lesions.
[0008] Another object is to provide a gel formulation in which
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol is soluble and does not substantially degrade during
storage.
[0009] Yet another object is to provide a topical gel formulation
for the topical delivery of
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-im-
idazo[4,5-c]quinoline-1-ethanol which does not deliver undue
amounts of the active compound systemically.
[0010] These objects, as well as others that will become apparent
upon reference to the following description, are provided by
pharmaceutical gel formulations including a drug, colloidal silicon
dioxide, triacetin and, preferably, propylene glycol. The drug is
preferably
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol, which has been found to be sufficiently soluble and
chemically stable in gel formulations of the present invention.
Moreover, it has been found that gel formulations of the present
invention, unlike certain other gel formulations, provide excellent
topical delivery of the drug while substantially avoiding unwanted
systemic delivery (thereby avoiding side effects).
[0011] As noted, the gel formulations of the invention preferably
include propylene glycol. One reason is because it appears,
surprisingly, that inclusion of propylene glycol thickens the gel
formulations and that the integrity of the resulting gel is
maintained at body temperature. It should be noted, however, that
gel formulations without propylene glycol, but including
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[-
4,5-c]quinoline-1-ethanol, colloidal silicon dioxide, and triacetin
have been found to be suitable, although less preferred.
[0012] The present invention also provides a method of inducing
cytokines, such as interferon and tumor necrosis factor, locally in
the skin or mucous membranes of a mammal, comprising placing on the
skin or mucous membranes of a mammal an amount of a formulation as
described above effective to induce cytokines. The formulations of
the present invention are also well-suited for treatment of
diseases by application of the formulation to skin and/or mucosal
lesions because the gel formulations do not need to include
irritating components.
BRIEF DESCRIPTION OF THE DRAWINGS
[0013] The invention will be described below with reference to
tests conducted using apparatus shown in the accompanying drawings,
wherein:
[0014] FIG. 1 is a modified Franz diffusion cell drug release test
apparatus; and
[0015] FIG. 2 is an alternative modified Franz diffusion cell drug
release test apparatus.
DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS
[0016] All weight percentages recited herein are based on the total
weight of the formulation unless otherwise indicated.
[0017] The present invention provides gel formulations preferably
containing
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-
-c]quinoline-1-ethanol (sometimes referred to herein as "the
drug").
[0018] The compound
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imi-
dazo[4,5-c]quinoline-1-ethanol is a known immune response enhancer
with antiviral properties. It can be synthesized using the method
disclosed in U.S. Pat. No. 5,389,640, the disclosure of which is
incorporated herein by reference. The compound can be used to treat
viral infections such as Type I or Type II Herpes simplex viral
infections and genital warts. Furthermore, the fact that the
compound induces a variety of cytokines including interferon
suggests that it and topical formulations containing it may be
useful in the treatment of other diseases where interferon has been
shown to be effective. The
4-amino-2-ethoxymethyl-.alpha.,.alpha.-di-
methyl-1H-imidazo[4,5-c]quinoline-1-ethanol will preferably be
present in a formulation of the invention in a therapeutically
effective amount i.e., an amount effective to treat the targeted
disease state or to prevent the recurrence of such a disease.
Generally
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol will preferably be present in a formulation of the
invention in an amount of 0.001 to about 0.6 percent by weight,
more preferably about 0.01 to about 0.5 percent by weight based on
the total weight of the formulation.
[0019]
4-Amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]qu-
inoline-1-ethanol exhibits substantial solubility in formulations
of the invention. Accordingly in a preferred embodiment of the
invention the drug is substantially fully dissolved in the
formulation.
[0020] Formulations of the invention contain colloidal silicon
dioxide as a gelling agent. Colloidal silicon dioxide is
commercially available under several trade names: AEROSIL from
Degussa (Deutsche Gold- und Silber-Schneideanstalt vormals
Roessler, Frankfurt, Germany), CAB-O-SIL from Cabot Corporation
(Tuscola, Ill., USA), and Wacker HDK from Wacker-Chemie GmbH
(Munich, Germany). Several grades of colloidal silicon dioxide
having different surface areas are commercially available. A
preferred grade has a specific surface area of about 200 m.sup.2/g
and is available under the trade designation AEROSIL 200. Colloidal
silicon dioxide will generally be present in a formulation of the
invention in an amount of about 7 to about 12 percent, preferably
about 8 to about 11 percent by weight based on the total weight of
the formulation.
[0021] Formulations of the invention preferably contain propylene
glycol (1,2-propanediol). The addition of propylene glycol has been
found, surprisingly, to thicken the gel formulations and to provide
a gel that maintains its integrity at body temperature. It is
believed that the propylene glycol may act as a solvent for the
colloidal silicon dioxide and as a solubilizer for the drug.
Generally propylene glycol will preferably be present in a
formulation of the invention in an amount of about 1 to about 30
percent, and more preferably about 5 to about 25 percent by weight
based on the total weight of the formulation.
[0022] Formulations of the invention also contain triacetin
(1,2,3-propanetriol triacetate). The triacetin is believed to act
as a solvent for the colloidal silicon dioxide and as a solubilizer
for the drug. In those formulations of the invention that do not
contain propylene glycol, triacetin will generally be present in
amount of about 88 to about 93 percent by weight based on the total
weight of the formulation. In those formulations of the invention
that do contain propylene glycol, triacetin will generally be
present in an amount of about 58 to about 92 percent, preferably
about 63 to about 88 percent by weight based on the total weight of
the formulation. However, the actual percentages of the triacetin
and other ingredients will depend on whether other ingredients are
included in the formulation.
[0023] A formulation of the present invention can be prepared by
combining the
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quin-
oline-1-ethanol with the triacetin and propylene glycol, if
present, and then heating with mixing to a temperature of about
50-55.degree. C. When the
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quin-
oline-1-ethanol appears to be completely dissolved, the colloidal
silicon dioxide is added and mixed until wetted. The resulting
mixture is sheared on a high speed propeller mixer until a
homogeneous gel is formed.
[0024] Formulations of the invention have been found to induce
interferon and tumor necrosis factor locally in the skin of mice.
Local induction helps avoid the side effects associated with the
systemic induction of these cytokines. As previously mentioned,
these side effects include fever, malaise, headache, nausea and
vomiting. The ability of the formulations of the invention to
induce interferon and tumor necrosis factor in the skin suggests
that they will be useful for the topical treatment of diseases such
as Type I and Type II Herpes simplex infections, warts including
genital warts, basal cell carcinoma, cervical intraepithelial
neoplasia and actinic keratosis.
[0025] The examples set forth below are intended to illustrate the
invention.
[0026] Cytokine Induction Test Method
[0027] The cytokine induction data given in the examples below were
obtained using the following test method.
[0028] For each formulation being tested, two groups of hairless
SKH-1 female mice (four mice per group) are dosed. A 10 .mu.L
portion of the drug containing formulation is applied to the right
flank and rubbed in for 1 minute. A 10 .mu.L portion of placebo gel
(which contains the same weight percent of silicon dioxide and
propylene glycol as the test gel with the remainder being
triacetin) is applied to the left flank and rubbed in for 1 minute.
One hour after dosing the first group of mice is sacrificed. Two
hours after dosing the second group of mice is sacrificed. The skin
is washed and tissue samples (100 mg) are removed from the right
flank (drug treated) and the left flank (placebo treated).
Individual samples are placed in cryovials and snap frozen in
liquid nitrogen. The samples are then homogenized in 1 mL of RPMI
medium containing 10% fetal calf serum and centrifuged at 2000 rpm
for 10 minutes. The supernatants are collected and frozen until
assayed for tumor necrosis factor (TNF) and interferon (INF). TNF
is assayed using a commercially available ELISA kit (Genzyme,
Cambridge, Mass.) and results are expressed as pg/mL.+-.SEM.
Interferon is measured by bioassay using L929 mouse fibroblasts
challenged with encephalomyocarditis virus. The details of the
bioassay method have been described by G. L. Brennan and L. H.
Kronenberg in "Automated Bioassay of Interferons in Micro-test
Plates", Biotechniques, June/July, 78, 1983, incorporated herein by
reference. Briefly stated, the method is as follows: interferon
dilutions and L929 cells are incubated at 37.degree. C. for 12 to
24 hours. The incubated cells are infected with an inoculum of
encephalomyocarditis virus. The infected cells are incubated for an
additional period at 37.degree. C. before quantifying for viral
cytopathic effect. The viral cytopathic effect is quantified by
staining followed by spectrophotometric absorbance measurements.
Results are expressed as units/mL.+-.SEM based on the value
obtained for NIH mouse interferon reference standard.
[0029] Drug Release Test Method
[0030] The drug release data given in the examples below were
obtained using the following test method.
[0031] A modified Franz diffusion cell 10 of the type shown in FIG.
1 is used. The cell is made of glass and holds approximately 11 mL
of receptor fluid in the cell body. The cell body opening is 1.6 cm
in diameter (2.0 cm.sup.2 area). A section of synthetic membrane 11
(microporous polyethylene film, CoTran.TM. 9711 from 3M Company) is
mounted between the upper portion 13 and lower portion 15 of the
cell. The membrane is held in place by means of a Teflon.RTM.
O-ring 14. The upper and lower portions are held together by means
of a clamp (not illustrated).
[0032] The portion of the cell below the mounted membrane is
completely filled with receptor fluid (0.1M sodium acetate buffer,
pH 4.0) such that the receptor fluid is in contact with the
membrane. The receptor fluid is stirred by means of magnetic stir
bar 17 and a magnetic stirrer (not illustrated). The sampling port
19 is covered except when in use.
[0033] When a gel formulation is evaluated, the membrane is placed
across the opening of the lower portion of the diffusion cell. The
O-ring is positioned on top of the membrane. A 1.50 g portion of
formulation is placed on top of the membrane and spread evenly over
the portion of the membrane surface which lies inside the O-ring.
The diffusion cell is assembled and the lower portion is filled
with 11 mL of warm (32.+-.1.degree. C.) receptor fluid.
[0034] The sampling port is covered and the cell is placed in a
constant temperature (32.+-.1.degree. C.) and humidity (50%.+-.15%
relative humidity) chamber. The receptor fluid is stirred
throughout the experiment. The entire volume of receptor fluid is
withdrawn at intervals of 30, 60, 120, 240 and 360 minutes elapsed
time and immediately replaced with fresh fluid. The withdrawn fluid
is analyzed for
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol using a uv spectrophotometer equipped with a flow cell
(1.0 cm for gels containing 0.05% or 0.01% drug; 0.2 cm for gels
containing 0.25% drug) and measuring the absorbance at 247 nm.
Results are reported as the cumulative amount of drug released at
30, 60, 120, 240 and 360 minutes and are expressed in units of
mg/cm.sup.2.
[0035] In Vitro Skin Penetration Test Method
[0036] The skin penetration data given in the examples below was
obtained using the following test method.
[0037] A modified Franz diffusion cell 20 of the type shown in FIG.
2 is used. Two types of skin are used, hairless mouse skin and
human cadaver skin. As shown in FIG. 2, the skin 22 is mounted
between the upper portion 23 and the lower portion 25 of the cell,
which are held together by means of a clamp 28.
[0038] The portion of the cell below the mounted membrane is
completely filled with receptor fluid (0.1M sodium acetate buffer,
pH 4.0) such that the receptor fluid is in contact with the skin.
The receptor fluid is stirred by means of a magnetic stir bar 27
and a magnetic stirrer (not illustrated). The sampling port 29 is
covered except when in use.
[0039] When a gel formulation is evaluated, the skin is placed
across the opening of the lower portion of the diffusion cell. A
300 mg portion of formulation is spread evenly over the skin. The
diffusion cell is assembled and the lower portion is filled with 10
mL of warm (32.+-.1.degree. C.) receptor fluid.
[0040] The sampling port is covered and the cell is placed in a
constant temperature (32.+-.1.degree. C.) and humidity (45%.+-.15%
relative humidity) chamber. The receptor fluid is stirred
throughout the experiment. The entire volume of receptor fluid is
withdrawn at intervals of 3, 6, 12, 24, 48 and 72 hours elapsed
time and immediately replaced with fresh fluid. The first 5 mL of
withdrawn fluid is filtered through a 0.45.mu. Acrodisc CRPTFE 25
mm filter (Miltex Instument Company, Ohio) and discarded. Then a 1
mL portion is filtered and placed in a high performance liquid
chromatography vial. The vial is capped then refrigerated until
analysis. The sample is analyzed for
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol using high performance liquid chromatography (Column:
15 cm.times.0.46 cm Supelcosil LC-8-DB (Supelco, Inc., Bellefonte,
Pa., USA), 5 .mu.m particle size; Mobile phase: acetonitrile/75 mM
ammonium phosphate aqueous buffer with 5 mM triethyl amine, pH 2.5,
19%/81% v/v; Flow rate: 2.0 L/min; Detector: uv at 245 nm). Results
are reported as the cumulative amount of drug penetrating at 3, 6,
12, 24, 48 and 72 hours and are expressed in units of .mu.g/mL.
EXAMPLE 1
[0041]
4-Amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]qu-
inoline-1-ethanol (0.75 g) was added to triacetin (272.25 g) in a
600 mL glass beaker. The resulting mixture was heated (about
55.degree. C.) with stirring until all of the drug was dissolved.
Colloidal silicon dioxide (27.0 g, AEROSIL.RTM. 200 from Degussa,
Frankfurt, Germany) was added to the solution and mixed with a
spatula until wetted. The mixture was sheared on a high speed
propeller mixer until a homogeneous gel was formed. The gel
contained 0.25% 4-amino-2-ethoxymethyl-.alpha.,.alpha.-di-
methyl-1H-imidazo[4,5-c]quinoline-1-ethanol, 9.0% colloidal silicon
dioxide, and 90.75% triacetin.
EXAMPLE 2
[0042] Propylene glycol (20.0 g), triacetin (343.0 g) and
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol (1.0 g) were placed in a 600 mL glass beaker then
heated (50.degree..+-.5.degree. C.) with stirring until all of the
drug was dissolved. Colloidal silicon dioxide (36.0 g) was added to
the solution and mixed with a spatula until wetted. The mixture was
sheared on a high speed propeller mixer until a homogeneous gel was
formed. The gel contained 0.25%
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidaz-
o[4,5-c]quinoline-1-ethanol, 5.0% propylene glycol, 9.0% colloidal
silicon dioxide, and 85.75% triacetin.
EXAMPLE 3
[0043] Propylene glycol (80.0 g) and
4-amino-2-ethoxymethyl-.alpha.,.alpha-
.-dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol (1.0 g) were placed
in a 600 mL glass beaker then heated (about 50.degree. C.) with
stirring until all of the drug was dissolved. Triacetin (283.0 g)
was added and the resulting mixture was stirred until a solution
was obtained. The heat was turned off. Colloidal silicon dioxide
(36.0 g) was added to the solution and mixed with a spatula until
wetted. The mixture was sheared on a high speed propeller mixer
until a homogeneous gel was formed. The gel contained 0.25%
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidaz-
o[4,5-c]quinoline-1-ethanol, 20.0% propylene glycol, 9.0% colloidal
silicon dioxide, and 70.75% triacetin.
[0044] The gel formulations of Examples 1-3 were tested for their
ability to induce cytokines using the test method described above.
The results shown in Table 1 below demonstrate that all three
formulations produced significant interferon and tumor necrosis
factor induction at the site of
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol application. Results are presented as the
mean.+-.standard error mean of 4 animals assayed individually. ND
means not done.
1TABLE 1 Cytokine Induction Cytokine Concentration Right Flank Left
Flank Time IFN TNF IFN TNF Formulation (hr) (U/mL) (pg/mL) (U/mL)
(pg/mL) 1 1 346 .+-. 54 699 .+-. 187 86 .+-. 37 273 .+-. 65 1 2 215
.+-. 107 496 .+-. 123 106 .+-. 98 344 .+-. 95 2 1 277 .+-. 80 340
.+-. 97 127 .+-. 71 163 .+-. 24 2 2 210 .+-. 70 610 .+-. 83 106
.+-. 105 237 .+-. 42 3 1 <1 .+-. 0 304 .+-. 39 <1 .+-. 0 165
.+-. 20 3 2 577 .+-. 210 1138 .+-. 232 105 .+-. 98 454 .+-. 175
Untreated 0 3 .+-. 1 120 .+-. 15 ND ND
EXAMPLE 4
[0045] Using the method of Example 1, a gel containing 0.05%
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol, 9.0% colloidal silicon dioxide, and 90.95% triacetin
was prepared.
EXAMPLE 5
[0046] Using the method of Example 2, a gel containing 0.05%
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol, 5.0% propylene glycol, 9.0% colloidal silicon dioxide,
and 85.95% triacetin was prepared.
EXAMPLE 6
[0047] Using the method of Example 3, a gel containing 0.05%
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol, 20.0% propylene glycol, 9.0% colloidal silicon
dioxide, and 70.95% triacetin was prepared.
[0048] The gel formulations of Examples 4-6 were tested for their
ability to induce cytokines using the test method described above.
The results shown in Table 2 below demonstrate that all three
formulations produced significant interferon and tumor necrosis
factor induction at the site of
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol application. Results are presented as the
mean.+-.standard error mean of 4 animals assayed individually. ND
means not done.
2TABLE 2 Cytokine Induction Cytokine Concentration Right Flank Left
Flank Time IFN TNF IFN TNF Formulation (hr) (U/mL) (pg/mL) (U/mL)
(pg/mL) 4 1 0.9 .+-. 0.5 412 .+-. 105 <1.0 280 .+-. 51 4 2 20
.+-. 10 345 .+-. 72 <1.0 153 .+-. 19 5 1 0.3 .+-. 0.3 348 .+-.
35 <1.0 262 .+-. 26 5 2 69 .+-. 58 346 .+-. 24 <1.0 194 .+-.
19 6 1 <1 .+-. 0 279 .+-. 45 <1.0 170 .+-. 40 6 2 43 .+-. 32
260 .+-. 47 <1.0 129 .+-. 41 Untreated 0 <1.0 220 .+-. 28 ND
ND
EXAMPLE 7
[0049] Using the method of Example 1, a gel containing 0.01%
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol, 9.0% colloidal silicon dioxide, and 90.99% triacetin
was prepared.
EXAMPLE 8
[0050] Using the method of Example 2, a gel containing 0.01%
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol, 5.0% propylene glycol, 9.0% colloidal silicon dioxide,
and 85.99% triacetin was prepared.
EXAMPLE 9
[0051] Using the method of Example 3, a gel containing 0.01%
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol, 20.0% propylene glycol, 9.0% colloidal silicon
dioxide, and 70.99% triacetin was prepared.
[0052] The gel formulations of Examples 7-9 were tested for their
ability to induce cytokines using the test method described above.
The results shown in Table 3 below demonstrate that the formulation
of Example 8 produced significant tumor necrosis factor induction
at the site of
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol application. Results are presented as the
mean.+-.standard error mean of 4 animals assayed individually. ND
means not done.
3TABLE 3 Cytokine Induction Cytokine Concentration Right Flank Left
Flank Time IFN TNF IFN TNF Formulation (hr) (U/mL) (pg/mL) (U/mL)
(pg/mL) 7 1 <1.0 455 .+-. 93 <1.0 421 .+-. 63 7 2 <1.0 382
.+-. 86 <1.0 398 .+-. 54 8 1 <1.0 406 .+-. 25 <1.0 316
.+-. 31 8 2 <1.0 296 .+-. 23 <1.0 375 .+-. 18 9 1 <1.0 380
.+-. 43 <1.0 352 .+-. 52 9 2 <1.0 296 .+-. 23 <1.0 366
.+-. 56 Untreated 0 <1.0 273 .+-. 44 ND ND
EXAMPLE 10
[0053] Using the method of Example 3, a gel containing 0.25%
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol, 20.0% propylene glycol, 8.0% colloidal silicon
dioxide, and 71.75% triacetin was prepared.
EXAMPLE 11
[0054] Using the method of Example 3, a gel containing 0.25%
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-1H-imidazo[4,5-c]quinolin-
e-1-ethanol, 20.0% propylene glycol, 10.0% colloidal silicon
dioxide, and 69.75% triacetin was prepared.
[0055] The gel formulations of Examples 2, 3, 5, 6, 8, 10 and 11
were tested for their ability to release
4-amino-2-ethoxymethyl-.alpha.,.alpha-
.-dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol using the test
method described above. The results shown in Table 4 below
demonstrate that all five formulations release
4-amino-2-ethoxymethyl-.alpha.,.alpha.-dimethyl-
-1H-imidazo[4,5-c]quinoline-1-ethanol. Each value is the mean of
the values from 6 diffusion cells.
4TABLE 4 Drug Release Cumulative Amount Released (mg/cm.sup.2)
Formulation 30 min 60 min 120 min 240 min 360 min Example 2 0.13
0.18 0.24 0.41 0.56 Example 3 0.15 0.23 0.33 0.50 0.62 Example 5
0.03 0.04 0.05 0.09 0.11 Example 6 0.03 0.04 0.07 0.12 0.15 Example
8 0.024 0.041 0.062 0.091 0.108 Example 10 0.14 0.22 0.33 0.58 0.72
Example 11 0.12 0.17 0.25 0.45 0.59
[0056] The in vitro penetration of
4-amino-2-ethoxymethyl-.alpha.,.alpha.--
dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol through hairless
mouse skin or human cadaver skin from a gel formulation of Example
3 was determined using the test method described above. The results
shown in Table 5 below demonstrate that the gel releases
4-amino-2-ethoxymethyl-.alpha.,.alpha.--
dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol to and through the
skin. Each value shown is the average of six independent
determinations with the standard deviation.
5TABLE 5 In Vitro Skin Penetration Time Cumulative Amount
Penetrating (.mu.g/mL) (hours) Hairless Mouse Skin Human Cadaver
Skin 3 0.073 .+-. 0.08 0 6 0.147 .+-. 0.133 0 12 0.413 .+-. 0.210 0
24 1.085 .+-. 0.380 0 48 2.89 .+-. 1.18 0.292 .+-. 0.088 72 5.455
.+-. 3.88 0.493 .+-. 0.158
* * * * *