U.S. patent application number 11/275699 was filed with the patent office on 2006-05-18 for method of using sulfonamide substituted imidazoquinolines.
This patent application is currently assigned to 3M Innovative Properties Company. Invention is credited to Stephen L. Crooks, Bryon A. Merrill.
Application Number | 20060106052 11/275699 |
Document ID | / |
Family ID | 26836139 |
Filed Date | 2006-05-18 |
United States Patent
Application |
20060106052 |
Kind Code |
A1 |
Crooks; Stephen L. ; et
al. |
May 18, 2006 |
METHOD OF USING SULFONAMIDE SUBSTITUTED IMIDAZOQUINOLINES
Abstract
Imidazoquinoline and tetrahydroimidazoquinoline compounds that
contain sulfonamide or sulfonamide functionality at the 1-position
are useful as immune response modifiers. The compounds and
compositions of the invention can induce the biosynthesis of
various cytokines and are useful in the treatment of a variety of
conditions including viral diseases and neoplastic diseases.
Inventors: |
Crooks; Stephen L.;
(Mahtomedi, MN) ; Merrill; Bryon A.; (River Falls,
WI) |
Correspondence
Address: |
3M INNOVATIVE PROPERTIES COMPANY
PO BOX 33427
ST. PAUL
MN
55133-3427
US
|
Assignee: |
3M Innovative Properties
Company
|
Family ID: |
26836139 |
Appl. No.: |
11/275699 |
Filed: |
January 25, 2006 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10826836 |
Apr 16, 2004 |
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11275699 |
Jan 25, 2006 |
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10027272 |
Dec 21, 2001 |
6800624 |
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10826836 |
Apr 16, 2004 |
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10166321 |
Jun 15, 2001 |
6825350 |
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10027272 |
Dec 21, 2001 |
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09589216 |
Jun 7, 2000 |
6331539 |
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10166321 |
Jun 15, 2001 |
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60138365 |
Jun 10, 1999 |
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Current U.S.
Class: |
514/292 |
Current CPC
Class: |
A61P 31/00 20180101;
A61K 9/006 20130101; A61P 37/02 20180101; A61K 31/4745 20130101;
A61P 35/00 20180101; A61P 37/00 20180101; C07D 471/04 20130101;
A61P 31/12 20180101; A61K 45/06 20130101; Y02A 50/30 20180101; A61P
37/04 20180101; A61K 9/0063 20130101; A61K 31/4745 20130101; A61K
2300/00 20130101 |
Class at
Publication: |
514/292 |
International
Class: |
A61K 31/4745 20060101
A61K031/4745 |
Claims
1. A method of inducing cytokine biosynthesis in an animal
comprising administering an effective amount of a compound of the
formula (I): ##STR241## wherein bonds represented by the dashed
lines are present or absent; R.sub.1 is
--(CH.sub.2).sub.2-4--NH--SO.sub.2--R.sub.4; R.sub.2 is hydrogen,
--C.sub.1-4 alkyl or --C.sub.1-4 alkyl-O--C.sub.1-4 alkyl; R.sub.4
is C.sub.1-4 alkyl; n is 0 and each R present is independently
selected from the group consisting of C.sub.1-10 alkyl, C.sub.1-10
alkoxy, halogen, and trifluoromethyl; or a pharmaceutically
acceptable salt thereof to the animal.
2. A method of inducing cytokine biosynthesis according to claim 1,
wherein R.sub.1 is --(CH.sub.2).sub.4--NH--SO.sub.2--C.sub.1-4
alkyl, and bonds represented by the dashed lines are present.
3. A method of inducing cytokine biosynthesis according to claim 2,
wherein R.sub.1 is --(CH.sub.2).sub.4--NH--SO.sub.2--CH.sub.3, and
R.sub.2 is --C.sub.1-4 alkyl.
Description
FIELD OF THE INVENTION
[0001] This invention relates to imidazoquinoline compounds that
have sulfonamide or sulfamide substitution at the 1-position and to
pharmaceutical compositions containing the compounds. A further
aspect of this invention relates to the use of these compounds as
immunomodulators, for inducing cytokine biosynthesis in animals and
in the treatment of diseases including viral and neoplastic
diseases.
BACKGROUND OF THE INVENTION
[0002] The first reliable report on the 1H-imidazo[4,5-c]quinoline
ring system, Backman et al., J. Org. Chem. 15, 1278-1284 (1950)
describes the synthesis of
1-(6-methoxy-8-quinolinyl)-2-methyl-1H-imidazo[4,5-c]quinoline for
possible use as an antimalarial agent. Subsequently, syntheses of
various substituted 1H-imidazo[4,5-c]quinolines were reported. For
example, Jain et al., J. Med. Chem. 11, pp. 87-92 (1968),
synthesized the compound
1-[2-(4-piperidyl)ethyl]-1H-imidazo[4,5-c]quinoline as a possible
anticonvulsant and cardiovascular agent. Also, Baranov et al.,
Chem. Abs. 85, 94362 (1976), have reported several
2-oxoimidazo[4,5-c]quinolines, and Berenyi et al., J. Heterocyclic
Chem. 18, 1537-1540 (1981), have reported certain
2-oxoimidazo[4,5-c]quinolines.
[0003] Certain 1H-imidazo[4,5-c]quinolin-4-amines and 1- and
2-substituted derivatives thereof were later found to be useful as
antiviral agents, bronchodilators and immunomodulators. These are
described in, inter alia, U.S. Pat. Nos. 4,689,338; 4,698,348;
4,929,624; 5,037,986; 5,268,376; 5,346,905; and 5,389,640, all of
which are incorporated herein by reference.
[0004] There continues to be interest in the imidazoquinoline ring
system, as seen for example in WO 98/30562, EP 894 797 and WO
00/09506. EP 894 797 discloses amide substituted imidazoquinoline
compounds that are disclosed to be useful as immune response
modifying compounds, while WO 00/09506 discloses imidazoquinoline
compounds that contain a sulfonamide substituent wherein the
sulfonamide nitrogen is part of a saturated heterocyclic ring.
Despite these efforts, however, there is a continuing need for
compounds that have the ability to modulate the immune response, by
induction of cytokine biosynthesis or other mechanisms.
SUMMARY OF THE INVENTION
[0005] We have found a new class of compounds that are useful in
inducing cytokine biosynthesis in animals. Accordingly, this
invention provides compounds of Formula I: ##STR1## wherein R,
R.sub.1 and R.sub.2 are as defined herein.
[0006] The compounds of Formula I are useful as immune response
modifiers due to their ability to induce cytokine biosynthesis and
otherwise modulate the immune reponse when administered to animals.
This makes the compounds useful in the treatment of a variety of
conditions such as viral diseases and tumors that are responsive to
such changes in the immune response.
[0007] The invention further provides pharmaceutial compositions
containing a therapeutically effective amount of a compound of
Formula I and methods of inducing cytokine biosynthesis in an
animal, treating a viral infection and/or treating a neoplastic
disease in an animal by administering a effective amount of a
compound of Formula I to the animal.
[0008] In addition, methods of synthesizing compounds of Formula I
and intermediates useful in the synthesis of these compounds are
provided.
DETAILED DESCRIPTION OF THE INVENTION
[0009] As mentioned earlier, the invention provides compounds of
Formula I: ##STR2## wherein
[0010] R.sub.1 is -alkyl-NR.sub.3--SO.sub.2--X--R.sub.4 or
-alkenyl-NR.sub.3--SO.sub.2--X--R.sub.4;
[0011] X is a bond or --NR.sub.5--;
[0012] R.sub.4 is aryl, heteroaryl, heterocyclyl, alkyl or alkenyl,
each of which may be unsubstituted or substituted by one or more
substituents selected from the group consisting of: [0013] -alkyl;
[0014] -alkenyl; [0015] -aryl; [0016] -heteroaryl; [0017]
-heterocyclyl; [0018] -substituted aryl; [0019] -substituted
heteroaryl; [0020] -substituted heterocyclyl; [0021] --O-alkyl;
[0022] --O-(alkyl).sub.0-1-aryl; [0023]
--O-(alkyl).sub.0-1-substituted aryl; [0024]
--O-(alkyl).sub.0-1-heteroaryl; [0025]
--O-(alkyl).sub.0-1-substituted heteroaryl; [0026]
--O-(alkyl).sub.0-1-heterocyclyl; [0027]
--O-(alkyl).sub.0-1-substituted heterocyclyl; [0028] --COOH; [0029]
--CO--O-alkyl; [0030] --CO-alkyl; [0031] --S(O).sub.0-2-alkyl;
[0032] --S(O).sub.0-2-(alkyl).sub.0-1-aryl; [0033]
--S(O).sub.0-2-(alkyl).sub.0-1-substituted aryl; [0034]
--S(O).sub.0-2-(alkyl).sub.0-1-heteroaryl; [0035]
--S(O).sub.0-2-(alkyl).sub.0-1-substituted heteroaryl; [0036]
--S(O).sub.0-2-(alkyl).sub.0-1-heterocyclyl; [0037]
--S(O).sub.0-2-(alkyl).sub.0-1-substituted heterocyclyl; [0038]
-(alkyl).sub.0-1-NR.sub.3R.sub.3; [0039]
-(alkyl).sub.0-1-NR.sub.3--CO--O-alkyl; [0040]
-(alkyl).sub.0-1-NR.sub.3--CO-alkyl; [0041]
-(alkyl).sub.0-1-NR.sub.3--CO-aryl; [0042]
-(alkyl).sub.0-1-NR.sub.3--CO-substituted aryl; [0043]
-(alkyl).sub.0-1-NR.sub.3--CO-heteroaryl; [0044]
-(alkyl).sub.0-1-NR.sub.3--CO-substituted heteroaryl; [0045]
--N.sub.3; [0046] -halogen; [0047] -haloalkyl; [0048] -haloalkoxy;
[0049] --CO-haloalkoxy; [0050] --NO.sub.2; [0051] --CN; [0052]
--OH; [0053] --SH; and in the case of alkyl, alkenyl, or
heterocyclyl, oxo;
[0054] R.sub.2 is selected from the group consisting of: [0055]
-hydrogen; [0056] -alkyl; [0057] -alkenyl; [0058] -aryl; [0059]
-substituted aryl; [0060] -heteroaryl; [0061] -substituted
heteroaryl; [0062] -alkyl --O-alkyl; [0063] -alkyl-O-- alkenyl; and
[0064] -alkyl or alkenyl substituted by one or more substituents
selected from the group consisting of: [0065] --OH; [0066]
-halogen; [0067] --N(R.sub.3).sub.2; [0068] --CO--N(R.sub.3).sub.2;
[0069] --CO--C.sub.1-10 alkyl; [0070] --CO--O--C.sub.1-10 alkyl;
[0071] --N.sub.3; [0072] -aryl; [0073] -substituted aryl; [0074]
-heteroaryl; [0075] -substituted heteroaryl; [0076] -heterocyclyl;
[0077] -substituted heterocyclyl; [0078] --CO-aryl; [0079]
--CO-(substituted aryl); [0080] --CO-heteroaryl; and [0081]
--CO-(substituted heteroaryl);
[0082] each R.sub.3 is independently selected from the group
consisting of hydrogen and C.sub.1-10 alkyl;
[0083] R.sub.5 is selected from the group consisting of hydrogen
and C.sub.1-10 alkyl, or R.sub.4 and R.sub.5 can combine to form a
3 to 7 membered heterocyclic or substituted heterocyclic ring;
[0084] n is 0 to 4 and each R present is independently selected
from the group consisting of C.sub.1-10 alkyl, C.sub.1-10 alkoxy,
halogen and trifluoromethyl,
or a pharmaceutically acceptable salt thereof.
Preparation of the Compounds
[0085] Imidazoquinolines of the invention can be prepared according
to Reaction Scheme I where R, R.sub.1, R.sub.2 and n are as defined
above.
[0086] In step (1) of Reaction Scheme I a 4-chloro-3-nitroquinoline
of Formula II is reacted with an amine of Formula R.sub.1NH.sub.2
where R.sub.1 is as defined above to provide a
3-nitroquinolin-4-amine of Formula III. The reaction can be carried
out by adding amine to a solution of a compound of Formula II in a
suitable solvent such as chloroform or dichloromethane and
optionally heating. Many quinolines of Formula II are known
compounds (see for example, U.S. Pat. No. 4,689,338 and references
cited therein).
[0087] In step (2) of Reaction Scheme I a 3-nitroquinolin-4-amine
of Formula III is reduced to provide a quinoline-3,4-diamine of
Formula IV. Preferably, the reduction is carried out using a
conventional heterogeneous hydrogenation catalyst such as platinum
on carbon or palladium on carbon. The reaction can conveniently be
carried out on a Parr apparatus in a suitable solvent such as
isopropyl alcohol or toluene.
[0088] In step (3) of Reaction Scheme I a quinoline-3,4-diamine of
Formula IV is reacted with a carboxylic acid or an equivalent
thereof to provide a 1H-imidazo[4,5-c]quinoline of Formula V.
Suitable equivalents to carboxylic acid include acid halides,
orthoesters, and 1,1-dialkoxyalkyl alkanoates. The carboxylic acid
or equivalent is selected such that it will provide the desired
R.sub.2 substituent in a compound of Formula V. For example,
triethyl orthoformate will provide a compound where R.sub.2 is
hydrogen and triethyl orthoacetate will provide a compound where
R.sub.2 is methyl. The reaction can be run in the absence of
solvent or in an inert solvent such as toluene. The reaction is run
with sufficient heating to drive off any alcohol or water formed as
a byproduct of the reaction.
[0089] In step (4) of Reaction Scheme I a
1H-imidazo[4,5-c]quinoline of Formula V is oxidized to provide a
1H-imidazo[4,5-c]quinoline-5N-oxide of Formula VI using a
conventional oxidizing agent that is capable of forming N-oxides.
Preferred reaction conditions involve reacting a solution of a
compound of Formula V in chloroform with 3-chloroperoxybenzoic acid
at ambient conditions.
[0090] In step (5) of Reaction Scheme I a
1H-imidazo[4,5-c]quinoline-5N-oxide of Formula VI is aminated to
provide a 1H-imidazo[4,5-c]quinolin-4-amine of Formula VII which is
a subgenus of Formula I. Step (5) involves (i) reacting a compound
of Formula VI with an acylating agent and then (ii) reacting the
product with an aminating agent. Part (i) of step (5) involves
reacting an N-oxide of Formula VI with an acylating agent. Suitable
acylating agents include alkyl- or arylsulfonyl chlorides (e.g.,
benezenesulfonyl chloride, methanesulfonyl chloride,
p-toluenesulfonyl chloride). Arylsulfonyl chlorides are preferred.
Para-toluenesulfonyl chloride is most preferred. Part (ii) of step
(5) involves reacting the product of part (i) with an excess of an
aminating agent. Suitable aminating agents include ammonia (e.g.,
in the form of ammonium hydroxide) and ammonium salts (e.g.,
ammonium carbonate, ammonium bicarbonate, ammonium phosphate).
Ammonium hydroxide is preferred. The reaction is preferably carried
out by dissolving the N-oxide of Formula VI in an inert solvent
such as dichloromethane, adding the aminating agent to the
solution, and then slowly adding the acylating agent. The product
or a pharmaceutically acceptable salt thereof can be isolated using
conventional methods.
[0091] Alternatively, step (5) may be carried out by (i) reacting
an N-oxide of Formula VI with an isocyanate and then (ii)
hydrolyzing the resulting product. Part (i) involves reacting the
N-oxide with an isocyanate wherein the isocyanato group is bonded
to a carbonyl group. Preferred isocyanates include trichloroacetyl
isocyanate and aroyl isocyanates such as benzoyl isocyanate. The
reaction of the isocyanate with the N-oxide is carried out under
substantially anhydrous conditions by adding the isocyanate to a
solution of the N-oxide in an inert solvent such as chloroform or
dichloromethane. Part (ii) involves hydrolysis of the product from
part (i). The hydrolysis can be carried out by conventional methods
such as heating in the presence of water or a lower alkanol
optionally in the presence of a catalyst such as an alkali metal
hydroxide or lower alkoxide. ##STR3##
[0092] Compounds of the invention where the R.sub.1 substituent
contains a sulfonamide can also be prepared according to Reaction
Scheme II where R, R.sub.2, R.sub.4 and n are as defined above and
m is 1-20.
[0093] In Reaction Scheme II an aminoalkyl substituted
1H-imidazo[4,5-c]quinolin-4-amine of Formula VIII is reacted with a
sulfonyl chloride of Formula IX to provide a compound of Formula X
which is a subgenus of Formula I. The reaction can be run at
ambient temperature in an inert solvent such as dichloromethane in
the presence of a base such as pyridine or
N,N-diisopropylethylamine. Many 1H-imidazo[4,5-c]quinolin-4-amines
of Formula VIII are known compounds, see for example U.S. Pat. No.
6,069,149 (Namba); others can be readily prepared using known
synthetic methods. Many sulfonyl chlorides of Formula IX are
commercially available; others can be readily prepared using known
synthetic methods. The product or a pharmaceutically acceptable
salt thereof can be isolated using conventional methods.
##STR4##
[0094] Compounds of the invention where the R.sub.1 substituent
contains a sulfonamide can also be prepared according to Reaction
Scheme III where R, R.sub.2, R.sub.4 and n are as defined above and
m is 1-20.
[0095] In Reaction Scheme III an aminoalkyl substituted
1H-imidazo[4,5-c]quinolin-4-amine of Formula VIII is reacted with a
sulfonic anhydride of Formula XI to provide a compound of Formula X
which is a subgenus of Formula I. The reaction can be run at
ambient temperature in an inert solvent such as dichloromethane in
the presence of a base such as pyridine or
N,N-diisopropylethylamine. Alternatively, the reaction can be run
at ambient temperature in acetonitrile. Many sulfonic anhydrides of
Formula XI are commercially available; others can be readily
prepared using known synthetic methods. The product or a
pharmaceutically acceptable salt thereof can be isolated using
conventional methods. ##STR5##
[0096] Tertiary sulfonamides of the invention can be prepared
according to Reaction Scheme IV where R, R.sub.2, R.sub.3, R.sub.4
and n are as defined above and m is 1-20.
[0097] In Reaction Scheme IV a 1H-imidazo[4,5-c]quinolinyl
sulfonamide of Formula X is reacted with a halide of Formula XII to
provide a compound of Formula XIII which is a subgenus of Formula
I. The reaction can be carried out at ambient temperature by adding
sodium hydride to a solution of a compound of Formula X in
N,N-dimethylformamide and then adding the halide. Many halides of
Formula XII are commercially available; others can be readily
prepared using known synthetic methods. The product or a
pharmaceutically acceptable salt thereof can be isolated using
conventional methods. ##STR6##
[0098] Compounds of the invention where R.sub.1 contains a
sulfamide group can be prepared according to Reaction Scheme V
wherein R, R.sub.2, R.sub.4, R.sub.5 and n are as defined above and
m is 1-20.
[0099] In step (1) of Reaction Scheme V an aminoalkyl substituted
1H-imidazo[4,5-c]quinolin-4-amine of Formula VIII is reacted with
sulfuryl chloride to generate in situ a sulfamoyl chloride of
Formula XIV. The reaction can be carried out by adding a solution
of sulfuryl chloride in dichloromethane to a solution of a compound
of Formula VIII in dichloromethane in the presence of one
equivalent of 4-(dimethylamino)pyridine. The reaction is preferably
carried out at a reduced temperature (-78.degree. C.). Optionally,
after the addition is complete the reaction mixture can be allowed
to warm to ambient temperature.
[0100] In step (2) of Reaction Scheme V an amine of Formula
R.sub.5R.sub.4NH is reacted with the sulfamoyl chloride of Formula
XIV to provide a 1H-imidazo[4,5-c]quinolinyl sulfamide of Formula
XV which is a subgenus of Formula I. The reaction can be carried
out by adding a solution containing 2 equivalents of the amine and
2 equivalents of triethylamine in dichloromethane to the reaction
mixture from step (1). The addition is preferably carried out at a
reduced temperature (-78.degree. C.). After the addition is
complete the reaction mixture can be allowed to warm to ambient
temperature. The product or a pharmaceutically acceptable salt
thereof can be isolated using conventional methods. ##STR7##
[0101] Tetrahydroimidazoquinolines of the invention can be prepared
according to Reaction Scheme VI where R.sub.2, R.sub.3, R.sub.4,
and R.sub.5 are as defined above and m is 1-20.
[0102] In step (1) of Reaction Scheme VI an aminoalkyl substituted
1H-imidazo[4,5-c]quinolin-4-amine of Formula XVI is reduced to
provide an aminoalkyl substituted
6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-4-amine of Formula
XVII. Preferably the reduction is carried out by suspending or
dissolving the compound of Formula XVI in trifluoroacetic acid,
adding a catalytic amount of platinum (IV) oxide, and then
subjecting the mixture to hydrogen pressure. The reaction can
conveniently be carried out on a Parr apparatus. The product or a
salt thereof can be isolated using conventional methods.
[0103] In step (2a) of Reaction Scheme VI an aminoalkyl substituted
6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-4-amine of Formula
XVII is reacted to provide a compound of Formula XVIII which is a
subgenus of Formula I. When R.sub.3 is hydrogen, the reaction can
be carried out in one step according to the methods described in
Reaction Schemes II and III above using a
tetrahydroimidazoquinoline of Formula XVII in place of the
imidazoquinoline of Formula VIII. When R.sub.3 is other than
hydrogen, the reaction can be carried out in two steps with step
one being carried out according to the methods of Reaction Schemes
II and III and step two being carried out according to the method
of Reaction IV using the tetrahydroimidazoquinoline analog of the
imidazoquinoline. The product or a pharmaceutically acceptable salt
thereof can be isolated using conventional methods.
[0104] In step (2b) of Reaction Scheme VI an aminoalkyl substituted
6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-4-amine of Formula
XVII is reacted to provide a compound of Formula XIX which is a
subgenus of Formula I. The reaction can be carried out according to
the method described in Reaction Scheme V using a
tetrahydroimidazoquinoline of Formula XVII in place of the
imidazoquinoline of Formula VIII. The product or a pharmaceutically
acceptable salt thereof can be isolated using conventional methods.
##STR8##
[0105] Tetrahydroimidazoquinolines of the invention can also be
prepared according to Reaction Scheme VII where R, R.sub.2,
R.sub.3, R.sub.4, R.sub.5 and n are as defined above and m is
1-20.
[0106] In step (1) of Reaction Scheme VII a
6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolinyl tert-butylcarbamate
of Formula XX is hydrolyzed to provide an aminoalkyl substituted
6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-4-amine of Formula
XXI. The reaction can be carried out dissolving the compound of
Formula XX in a mixture of trifluoroacetic acid and acetonitrile
and stirring at ambient temperature. Alternatively, the compound of
Formula XX can be combined with dilute hydrochloric acid and heated
on a steam bath. Tetrahydro-1H-imidazo[4,5-c]quinolinyl
tert-butylcarbamates of Formula XX can be prepared using the
synthetic route disclosed in U.S. Pat. No. 5,352,784 (Nikolaides).
The product or a salt thereof can be isolated using conventional
methods.
[0107] Steps (2a) and (2b) can be carried out in the same manner as
in Reaction Scheme VI. ##STR9##
[0108] Some compounds of Formula I can be readily prepared from
other compounds of Formula I. For example, compounds wherein the
R.sub.4 substituent contains a chloroalkyl group can be reacted
with an amine to provide an R.sub.4 substituent substituted by a
secondary or teriary amino group; compounds wherein the R.sub.4
substituent contains a nitro group can be reduced to provide a
compound wherein the R.sub.4 substituent contains a primary
amine.
[0109] As used herein, the terms "alkyl", "alkenyl", "alkynyl" and
the prefix "-alk" are inclusive of both straight chain and branched
chain groups and of cyclic groups, i.e. cycloalkyl and
cycloalkenyl. Unless otherwise specified, these groups contain from
1 to 20 carbon atoms, with alkenyl and alkynyl groups containing
from 2 to 20 carbon atoms. Preferred groups have a total of up to
10 carbon atoms. Cyclic groups can be monocyclic or polycyclic and
preferably have from 3 to 10 ring carbon atoms. Exemplary cyclic
groups include cyclopropyl, cyclopentyl, cyclohexyl and
adamantyl.
[0110] The term "haloalkyl" is inclusive of groups that are
substituted by one or more halogen atoms, including groups wherein
all of the available hydrogen atoms are replaced by halogen atoms.
This is also true of groups that include the prefix "haloalk-".
Examples of suitable haloalkyl groups are chloromethyl,
trifluoromethyl, and the like.
[0111] The term "aryl" as used herein includes carbocyclic aromatic
rings or ring systems. Examples of aryl groups include phenyl,
naphthyl, biphenyl, fluorenyl and indenyl. The term "heteroaryl"
includes aromatic rings or ring systems that contain at least one
ring hetero atom (e.g., O, S, N). Suitable heteroaryl groups
include furyl, thienyl, pyridyl, quinolinyl, tetrazolyl, imidazo,
pyrazolo, thiazolo, oxazolo, and the like.
[0112] "Heterocyclyl" includes non-aromatic rings or ring systems
that contain at least one ring hetero atom (e.g., O, S, N).
Exemplary heterocyclic groups include pyrrolidinyl,
tetrahydrofuranyl, morpholinyl, thiomorpholinyl, piperidinyl,
piperazinyl, thiazolidinyl, imidazolidinyl, and the like.
[0113] Unless otherwise specified, the terms "substituted
cycloalkyl", "substituted aryl", "substituted heteroaryl" and
"substituted heterocyclyl" indicate that the rings or ring systems
in question are further substituted by one or more substituents
independently selected from the group consisting of alkyl, alkoxy,
alkylthio, hydroxy, halogen, haloalkyl, haloalkylcarbonyl,
haloalkoxy (e.g., trifluoromethoxy), nitro, alkylcarbonyl,
alkenylcarbonyl, arylcarbonyl, heteroarylcarbonyl, aryl, arylalkyl,
heteroaryl, heteroarylalkyl, heterocyclyl, heterocycloalkyl,
nitrile, alkoxycarbonyl, alkanoyloxy, alkanoylthio, and in the case
of cycloalkyl and heterocyclyl, oxo.
[0114] In structural formulas representing compounds of the
invention certain bonds are represented by dashed lines. These
lines mean that the bonds represented by the dashed line can be
present or absent. Accordingly, compounds of Formula I can be
either imidazoquinoline compounds or tetrahydroimidazoquinoline
compounds.
[0115] The invention is inclusive of the compounds described herein
in any of their pharmaceutically acceptable forms, including
isomers such as diastereomers and enantiomers, salts, solvates,
polymorphs, and the like.
Pharmaceutical Compositions and Biological Activity
[0116] Pharmaceutical compositions of the invention contain a
therapeutically effective amount of a compound of Formula I in
combination with a pharmaceutically acceptable carrier.
[0117] As used herein, the term "a therapeutically effective
amount" means an amount of the compound sufficient to induce a
therapeutic effect, such as cytokine induction, antitumor activity
and/or antiviral activity. Although the exact amount of active
compound used in a pharmaceutical composition of the invention will
vary according to factors known to those of skill in the art, such
as the physical and chemical nature of the compound as well as the
nature of the carrier and the intended dosing regimen, it is
anticipated that the compositions of the invention will contain
sufficient active ingredient to provide a dose of about 100 ng/kg
to about 50 mg/kg, preferably about 10 .mu.g/kg to about 5 mg/kg of
the compound to the subject. Any of the conventional dosage forms
may be used, such as tablets, lozenges, parenteral formulations,
syrups, creams, ointments, aerosol formulations, transdermal
patches, transmucosal patches and the like.
[0118] The compounds of the invention have been shown to induce the
production of certain cytokines in experiments performed according
to the tests set forth below. These results indicate that the
compounds are useful as immune response modifiers that can modulate
the immune response in a number of different ways, rendering them
useful in the treatment of a variety of disorders.
[0119] Cytokines that may be induced by the administration of
compounds according to the invention generally include
interferon-.alpha. (IFN-.alpha.) and tumor necrosis factor-.alpha.
(TNF-.alpha.) as well as certain interleukins (IL). Cytokines whose
biosynthesis may be induced by compounds of the invention include
IFN-.alpha., TNF-.alpha., IL-1, 6, 10 and 12, and a variety of
other cytokines. Among other effects, cytokines inhibit virus
production and tumor cell growth, making the compounds useful in
the treatment of viral diseases and tumors.
[0120] In addition to the ability to induce the production of
cytokines, the compounds of the invention affect other aspects of
the innate immune response. For example, natural killer cell
activity may be stimulated, an effect that may be due to cytokine
induction. The compounds may also activate macrophages, which in
turn stimulates secretion of nitric oxide and the production of
additional cytokines. Further, the compounds may cause
proliferation and differentiation of B-lymphocytes.
[0121] Compounds of the invention also have an effect on the
acquired immune response. For example, although there is not
believed to be any direct effect on T cells or direct induction of
T cell cytokines, the production of the T helper type 1 (Th1)
cytokine IFN-.gamma. is induced indirectly and the production of
the T helper type 2 (Th2) cytokines IL-4, IL-5 and IL-13 are
inhibited upon administration of the compounds. This activity means
that the compounds are useful in the treatment of diseases where
upregulation of the Th1 response and/or downregulation of the Th2
response is desired. In view of the ability of compounds of Formula
Ia to inhibit the Th2 immune response, the compounds are expected
to be useful in the treatment of atopic diseases, e.g., atopic
dermatitis, asthma, allergy, and allergic rhinitis; and systemic
lupus erythematosis; as a vaccine adjuvant for cell mediated
immunity; and possibly as a treatment for recurrent fungal diseases
and chlamydia.
[0122] The immune response modifying effects of the compounds make
them useful in the treatment of a wide variety of conditions.
Because of their ability to induce the production of cytokines such
as IFN-.alpha. and/or TNF-.alpha., the compounds are particularly
useful in the treatment of viral diseases and tumors. This
immunomodulating activity suggests that compounds of the invention
are useful in treating diseases such as, but not limited to, viral
diseases including genital warts; common warts; plantar warts;
Hepatitis B; Hepatitis C; Herpes Simplex Virus Type I and Type II;
molluscum contagiosum; HIV; CMV; VZV; intraepithelial neoplasias
such as cervical intraepithelial neoplasia; human papillomavirus
(HPV) and associated neoplasias; fungal diseases, e.g. candida,
aspergillus, and cryptococcal meningitis; neoplastic diseases,
e.g., basal cell carcinoma, hairy cell leukemia, Kaposi's sarcoma,
renal cell carcinoma, squamous cell carcinoma, myelogenous
leukemia, multiple myeloma, melanoma, non-Hodgkin's lymphoma,
cutaneous T-cell lymphoma, and other cancers; parasitic diseases,
e.g. pneumocystis carnii, cryptosporidiosis, histoplasmosis,
toxoplasmosis, trypanosome infection, leishmaniasis; and bacterial
infections, e.g., tuberculosis, mycobacterium avium. Additional
diseases or conditions that can be treated using the compounds of
the invention include eczema; eosinophilia; essential
thrombocythaemia; leprosy; multiple sclerosis; Ommen's syndrome;
discoid lupus; Bowen's disease; Bowenoid papulosis; and to enhance
or stimulate the healing of wounds, including chronic wounds.
[0123] Accordingly, the invention provides a method of inducing
cytokine biosynthesis in an animal comprising administering an
effective amount of a compound of Formula I to the animal. An
amount of a compound effective to induce cytokine biosynthesis is
an amount sufficient to cause one or more cell types, such as
monocytes, macrophages, dendritic cells and B-cells to produce an
amount of one or more cytokines such as, for example, IFN-(, TNF-(,
IL-1,6, 10 and 12 that is increased over the background level of
such cytokines. The precise amount will vary according to factors
known in the art but is expected to be a dose of about 100 ng/kg to
about 50 mg/kg, preferably about 10 .mu.g/kg to about 5 mg/kg. The
invention also provides a method of treating a viral infection in
an animal, and a method of treating a neoplastic disease in an
animal, comprising administering an effective amount of a compound
of Formula I to the animal. An amount effective to treat or inhibit
a viral infection is an amount that will cause a reduction in one
or more of the manifestations of viral infection, such as viral
lesions, viral load, rate of virus production, and mortality as
compared to untreated control animals. The precise amount will vary
according to factors known in the art but is expected to be a dose
of 100 ng/kg to about 50 mg/kg, preferably about 10 .mu.g/kg to
about 5 mg/kg. An amount of a compound effective to treat a
neoplastic condition is an amount that will cause a reduction in
tumor size or in the number of tumor foci. Again, the precise
amount will vary according to factors known in the art but is
expected to be a dose of about 100 ng/kg to about 50 mg/kg,
preferably about 10 kg/kg to about 5 mg/kg.
[0124] The invention is further described by the following
examples, which are provided for illustration only and are not
intended to be limiting in any way.
EXAMPLE 1
N.sup.1-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-5-(dimet-
hylamino)-1-naphthalenesulfonamide
[0125] ##STR10##
[0126] 5-Dimethylamino-1-naphthalenesulfonyl chloride (1.82 g, 6.74
mmol) was added to a mixture of N,N-diisopropylethylamine (1.23 mL,
7.06 mmol), dichloromethane (15 mL) and
1-(4-aminobutyl)-2-butyl-1H-imidazo[4,5-c]quinolin-4-amine (2.0 g,
6.42 mmol). The reaction mixture was allowed to stir at ambient
temperature overnight. Methanol was added to the reaction mixture
until a clear solution was obtained. Silica gel was added to the
reaction mixture and then the solvents were removed. The silica gel
was placed in a column and then eluted with chloroform in a
stepwise gradient to 9:1 chloroform:methanol. The resulting product
was recrystallized from N,N-dimethylformamide and deionized water
to provide 2.5 g of
N.sup.1-[4-(4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-5-(dime-
thylamino)-1-naphthalenesulfonamide as a yellow crystalline solid,
m.p. 223-224.degree. C. Analysis: Calculated for
C.sub.30H.sub.36N.sub.6O.sub.2S: % C, 66.15; % H, 6.66; % N, 15.43;
Found: % C, 66.36; % H, 6.34; % N, 15.23.
EXAMPLE 2
N.sup.1-[4-(4-Amino-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-5-(dimethylamino-
)-1-naphthalenesulfonamide
[0127] ##STR11##
[0128] A suspension of
1-(4-aminobutyl)-1H-imidazo[4,5-c]quinolin-4-amine (0.5 g, 2.0
mmol) in pyridine (250 mL) was warmed to 60.degree. C. to dissolve
the amine. The solution was allowed to cool to about 30.degree. C.
and then 5-dimethylamino-1-naphthalenesulfonyl chloride (0.5 g, 1.8
mmol) was slowly added. After 1 hour 0.3 g of
5-dimethylamino-1-naphthalenesulfonyl chloride was added. The
reaction mixture was warmed to 60.degree. C. and maintained at that
temperature overnight. The reaction mixture was concentrated under
vacuum. The residue was recrystallized from propyl acetate to
provide
N.sup.1-[4-(4-amino-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-5-(dimethylamin-
o)-1-naphthalenesulfonamide as a solid, m.p. 200-201.degree. C.
EXAMPLE 3
N.sup.2-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-2-thioph-
enesulfonamide
[0129] ##STR12##
[0130] 2-Thiophenesulfonyl chloride (0.3 g in 10 ml
dichloromethane, 1.6 mmol) was added dropwise to a stirring
solution of
1-(4-aminobutyl)-2-butyl-1H-imidazo[4,5-c]quinoline-4-amine (0.5 g,
1.6 mmol), dichloromethane (40 ml), and pyridine (0.8 ml). The
reaction was maintained at room temperature for a few hours and
then an additional portion of 2-thiophenesulfonyl chloride (0.1 g,
0.6 mmol) was added. The reaction was maintained overnight and then
concentrated in vacuo. The resulting residue was purified by flash
column chromatography (silica gel, 9:1 dichloromethane\methanol)
and the fractions containing product were washed with saturated
aqueous sodium bicarbonate. The organic layer was dried
(MgSO.sub.4), filtered, and concentrated to provide 0.2 g of
N.sup.2-[4-(4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-2-thiop-
henesulfonamide as an off white powder, m.p. 137.5-141.5.degree. C.
.sup.1H NMR (300 MHz, DMSO-d.sub.6) .delta. 8.00 (d, J=8.0 Hz, 1H),
7.89 (dd, J=5.0, 1.3 Hz, 1H), 7.83 (broad s, 1H), 7.61 (dd, J=8.3,
1.1 Hz, 1H), 7.54 (dd, J=3.7, 1.3 Hz, 1H), 7.42 (t, J=7.2 Hz, 1H),
7.25 (m, 1H), 7.15 (m, 1H), 6.44 (broad s, 2H), 4.47 (t, J=7.4 Hz,
2H), 2.87 (m, 4H), 1.80 (m, 4H), 1.58-1.38 (m, 4H), 0.96 (t, J=7.4
Hz, 3H); IR (KBr) 3467, 3361, 3167, 3091, 2957, 2933, 2870, 1644,
1617, 1585, 1533, 1478, 1405, 1336, 1154, 1095, 1014, 854, 761, 733
cm.sup.-1; MS (E1) m/e 457.1606 (457.1606 calcd for
C.sub.22H.sub.27N.sub.5O.sub.2S.sub.2); Anal calcd for
C.sub.22H.sub.27N.sub.5O.sub.2S.sub.2: C, 57.74; H, 5.95; N, 15.30.
Found: C, 57.50; H, 5.98; N, 15.15.
EXAMPLE 4
N-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]
phenylmethanesulfonamide
[0131] ##STR13##
[0132] .alpha.-Toluenesulfonyl chloride (0.5 g in 10 ml
dichloromethane, 2.7 mmol) was added dropwise to a stirring
solution of
1-(4-aminobutyl)-2-butyl-1H-imidazo[4,5-c]quinoline-4-amine (0.75
g, 2.4 mmol), dichloromethane (115 ml), and pyridine (1 ml). The
reaction was maintained at room temperature for 4 hours and then
concentrated in vacuo. The residue was purified by flash column
chromatography (silica gel, 9:1 dichloromethane\methanol, R.sub.f
0.16). The fractions containing product were combined and washed
with saturated aqueous bicarbonate. The organic layer was dried
(MgSO4), filtered, and concentrated. A final recrystallization from
dichloromethane\diethyl ether provided 0.65 g of
N-[4-(4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]phenylmethanes-
ulfonamide as a white crystalline solid, m.p. 197.0-199.5.degree.
C. .sup.1H NMR (300 MHz, DMSO-d.sub.6) .delta. 8.02 (d, J=7.6 Hz,
1H), 7.62 (dd, J=8.3, 1.1 Hz, 1H), 7.42 (dt, J=7.5, 1.1 Hz, 1H),
7.35-7.23 (m, 7H), 7.12 (t, J=5.4 Hz, 1H), 6.46 (broad s, 2H), 4.49
(t, J=7.5 Hz, 2H), 4.29 (s, 2H), 2.91 (m, 4H), 1.83-1.42 (m, 8H),
0.96 (t, J=7.4 Hz, 3H); IR (KBr) 3460, 3293, 3226, 3158, 2955,
2931, 2867, 1632, 1586, 1534, 1482, 1437, 1389, 1331, 1152, 1094,
752, 700 cm.sup.-1; MS (E1) m/e 465.2204 (465.2198 calcd for
C.sub.25H.sub.31N.sub.5O.sub.2S); Anal calcd for
C.sub.25H.sub.31N.sub.5O.sub.2S: C, 64.49; H, 6.71; N, 15.04.
Found: C, 64.15; H, 6.71; N, 15.00.
EXAMPLE 5
N.sup.1-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-1-benzen-
esulfonamide
[0133] ##STR14##
[0134] Benzenesulfonyl chloride (0.45 ml in 10 ml dichloromethane,
3.5 mmol) was added dropwise to a stirring solution of
1-(4-aminobutyl)-2-butyl-1H-imidazo[4,5-c]quinoline-4-amine (1.0 g,
3.2 mmol), dichloromethane (140 ml), and pyridine (0.8 ml). The
reaction was maintained at room temperature for four hours and then
concentrated in vacuo. The residue was purified by flash column
chromatography (silica gel, 9:1 dichloromethane\methanol, R.sub.f
0.28) followed by recrystallization from dichloromethane\diethyl
ether to provide 1.14 g of
N.sup.1-[4-(4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-1-benze-
nesulfonamide as a white powder, m.p. 75.5-79.0.degree. C. .sup.1H
NMR (500 MHz, DMSO-d.sub.6) .delta. 7.99 (d, J=7.7 Hz, 1H), 7.76
(d, J=7.2, 2H), 7.63-7.53 (m, 5H), 7.42 (m, 1H), 7.25 (m, 1H), 6.43
(broad s, 2H), 4.45 (t, J=7.6 Hz, 2H), 2.87 (t, J=7.7 Hz, 2H), 2.78
(m, 2H), 1.79 (m, 4H), 1.55-1.40 (m, 4H), 0.95 (t, J=7.4 Hz, 3H);
MS (E1) m/e 451.2036 (451.2042 calcd for
C.sub.24H.sub.29N.sub.5O.sub.2S); Anal calcd for
C.sub.24H.sub.29N.sub.5O.sub.2S: C, 63.83; H, 6.47; N, 15.51.
Found: C, 63.89; H, 6.42; N, 15.30.
EXAMPLE 6
N-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]methanesulfonam-
ide
[0135] ##STR15##
[0136] Methanesulfonic anhydride (0.6 g, 3.4 mmol) was added
dropwise to a stirring solution of
1-(4-aminobutyl)-2-butyl-1H-imidazo[4,5-c]quinoline-4-amine (1.0 g,
3.2 mmol) and acetonitrile (200 ml). A precipitate formed within a
few minutes. The solvent was removed in vacuo and the residue was
partitioned between dichloromethane and saturated aqueous sodium
bicarbonate. The fractions were separated and the organic fraction
was dried (MgSO.sub.4), filtered and concentrated to yield the
crude product as a white solid. Recrystallization from methyl
acetate provided
N-[4-(4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]methanesulfona-
mide as a white crystalline solid, m.p. 195.1-196.0.degree. C.
.sup.1H NMR (300 MHz, DMSO-d.sub.6) .delta. 8.04 (d, J=7.4 Hz, 1H),
7.61 (dd, J=8.3, 1.2 Hz, 1H), 7.50 (dt, J=7.5, 1.1 Hz, 1H), 7.26
(dt, J=7.5, 1.2 Hz, 1H), 6.99 (t, J=5.7 Hz, 1H), 6.44 (broad s,
2H), 4.52 (t, J=7.5 Hz, 2H), 3.02-2.86 (m, 7H), 1.82 (m, 4H), 1.62
(m, 2H), 1.46 (q, J=7.4 Hz, 2H), 0.96 (t, J=7.4 Hz, 3H); IR (KBr)
3348, 3299, 3152, 2952, 2931, 2869, 1642, 1584, 1530, 1480, 1323,
1155, 1142, 1094, 982, 765 cm.sup.-1; MS (E1) m/e 389.1889
(389.1885 calcd for C.sub.19H.sub.27N.sub.5O.sub.2S); Anal calcd
for C.sub.19H.sub.27N.sub.5O.sub.2S: C, 58.59; H, 6.99; N, 17.98.
Found: C, 58.26; H, 6.64; N, 17.69
EXAMPLE 7
N.sup.1-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-3-nitro--
1-benzenesulfonamide Hydrochloride
[0137] ##STR16##
[0138] According to the general method of Example
5,3-nitrobenzenesulfonyl chloride and
1-(4-aminobutyl)-2-butyl-1H-imidazo[4,5-c]quinoline-4-amine were
combined.
N.sup.1-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)buty-
l]-3-nitro-1-benzenesulfonamide was isolated as the hydrochloride
salt (white solid), m.p. 176.0-178.2.degree. C. .sup.1H NMR (300
MHz, DMSO-d.sub.6) .delta. 8.70 (very broad s, 2H), 8.49-8.42 (m,
2H), 8.21-8.17 (m, 2H), 8.06 (t, J=5.7 Hz, 1H), 7.88-7.81 (m, 2H),
7.71 (t, J=7.7 Hz, 1H), 7.57 (t, J=7.7 Hz, 1H), 4.56 (t, J=7.3 Hz,
2H), 2.94 (t, J=7.7 Hz, 2H), 2.86 (m, 2H), 1.81 (m, 4H), 1.60-1.42
(m, 4H), 0.96 (t, J=7.3 Hz, 3H); IR (KBr) 3096, 2954, 2869, 2771,
1671, 1607, 1528, 1351, 1335, 1163, 1128, 1083, 879, 758, 735, 672,
661 cm.sup.-1; MS (E1) m/e 496.1897 (496.1893 calcd for
C.sub.24H.sub.28N.sub.6O.sub.4S). Anal calcd for
C.sub.24H.sub.28N.sub.6O.sub.4S*HCl*H.sub.2O: C, 52.31; H, 5.67; N,
15.25. Found: C, 52.26; H, 5.46; N, 15.09.
EXAMPLE 8
N.sup.1-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-3-amino--
1-benzenesulfonamide Hydrochloride
[0139] ##STR17##
[0140] A solution of
N.sup.1-[4-(4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-3-nitro-
-1-benzenesulfonamide hydrochloride (0.4 g) in methanol (250 ml)
was charged with a catalytic amount of 10% palladium on carbon
(0.085 g). The reaction was placed under an atmosphere of hydrogen
(50 psi; 3.44.times.10.sup.5 Pa) and shaken on a Parr apparatus for
2 hours. The reaction mixture was filtered and the solvent removed
in vacuo. The solid product was recrystallized from 2-propanol to
provide 0.18 g of
N.sup.1-[4-(4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-3-amino-
-1-benzenesulfonamide hydrochloride as an off white crystalline
solid, m.p. 110.2.degree. C. (decomposition). .sup.1H NMR (300 MHz,
DMSO-d.sub.6) .delta. 8.70 (very broad s, 2H), 8.22 (d, J=8.2 Hz,
1H), 7.83 (d, J=7.8 Hz, 1H), 7.72 (t, J=7.6 Hz, 1H), 7.59 (t, J=7.7
Hz, 1H), 7.43 (t, J=5.9 Hz, 1H), 7.15 (t, J=7.9 Hz, 1H), 6.95 (t,
J=1.9 Hz, 1H), 6.84 (d, J=7.7 Hz, 1H), 6.73 (dd, J=8.0, 1.5 Hz,
1H), 5.63 (broad s, 2H), 4.56 (t, J=7.5 Hz, 2H), 2.96 (t, J=7.7 Hz,
2H), 2.77 (q, J=6.3 Hz, 2H), 1.83 (m, 4H), 1.60-1.40 (m, 4H), 0.97
(t, J=7.3 Hz, 3H); IR (KBr) 3313, 3135, 2957, 2870, 2782, 1671,
1599, 1485, 1454, 1313, 1155, 1084, 754, 686 cm.sup.-1; MS (EI) m/e
466.2150 (466.2151 calcd for C.sub.24H.sub.30N.sub.6O.sub.2S). Anal
calcd for C.sub.24H.sub.30N.sub.6O.sub.2S*HCl*0.25H.sub.2O: C,
56.79; H, 6.26; N, 16.56; Cl, 6.98. Found: C, 56.87; H, 6.22; N,
16.19; Cl, 7.22.
EXAMPLE 9
N.sup.1-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-4-nitro--
1-benzenesulfonamide Hydrochloride
[0141] ##STR18##
[0142] According to the general method of Example
5,4-nitrobenzenesulfonyl chloride and
1-(4-aminobutyl)-2-butyl-1H-imidazo[4,5-c]quinoline-4-amine were
combined.
N.sup.1-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)buty-
l]-4-nitro-1-benzenesulfonamide was isolated as the hydrochloride
salt (white solid), m.p. 96.0.degree. C. (decomposition). .sup.1H
NMR (300 MHz, DMSO-d.sub.6) .delta. 8.70 (very broad s, 2H),
8.38-8.34 (m, 2H), 8.19 (d, J=8.2 Hz, 1H), 8.09 (t, J=5.6 Hz, 1H),
8.03-7.99 (m, 2H), 7.80 (d, J=7.4 Hz, 1H), 7.68 (t, J=7.4 Hz, 1H),
7.54 (t, J=7.2 Hz, 1H), 4.55 (t, J=7.4 Hz, 2H), 2.94 (t, J=7.7 Hz,
2H), 2.86 (q, J=6.2 Hz, 2H), 1.80 (m, 4H), 1.58 (m, 2H), 1.45 (q,
J=7.5 Hz, 2H), 0.96 (t, J=7.3 Hz, 3H); IR (KBr) 3283, 3100, 2957,
2870, 2782, 1670, 1606, 1528, 1347, 1311, 1162, 1092, 854, 746,
737, 686 cm.sup.-1; MS (E1) m/e 496.1902 (496.1893 calcd for
C.sub.24H.sub.28N.sub.6O.sub.4S). Anal calcd for
C.sub.24H.sub.28N.sub.6O.sub.4S*HCl*0.85H.sub.2O: C, 52.57; H,
5.64; N, 15.33. Found: C, 52.57; H, 5.46; N, 15.33.
EXAMPLE 10
N.sup.1-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-4-amino--
1-benzenesulfonamide Hydrochloride
[0143] ##STR19##
[0144] A solution of
N.sup.1-[4-(4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-4-nitro-
-1-benzenesulfonamide hydrochloride (0.38 g) in methanol (250 ml)
was charged with a catalytic amount of 10% palladium on carbon
(0.085 g). The reaction was placed under an atmosphere of hydrogen
(50 psi; 3.44.times.10.sup.5 Pa)) and shaken on a Parr apparatus
for 2 hours. The reaction mixture was filtered and the solvent
removed in vacuo. The solid product was recrystallized from
2-propanol to provide 0.34 g of
N.sup.1-[4-(4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-4-amino-
-1-benzenesulfonamide hydrochloride as an off white powder, m.p.
203.1-205.0.degree. C. .sup.1H NMR (300 MHz, DMSO-d.sub.6) .delta.
8.65 (very broad s, 2H), 8.21(d, J=8.0 Hz, 1H), 7.82 (m, 1H), 7.71
(t, J=7.7 Hz, 1H), 7.58 (t, J=7.7 Hz, 1H), 7.38 (d, J=8.7 Hz, 2H),
7.13 (t, J=5.9 Hz, 1H), 6.60 (d, J=8.7 Hz, 2H), 5.92 (broad s, 2H),
4.55 (t, J=7.6 Hz, 2H), 2.96 (t, J=7.6 Hz, 2H), 2.70 (q, J=6.4 Hz,
2H), 1.81 (m, 4H), 1.58-1.43 (m, 4H), 0.96 (t, J=7.4 Hz, 3H); IR
(KBr) 3430, 3316, 3215, 3046, 2955, 2868, 2679, 1671, 1594, 1334,
1157, 1091, 851, 776, 759 cm.sup.-1; MS (E1) m/e 466.2145 (466.2151
calcd for C.sub.24H.sub.30N.sub.6O.sub.2S). Anal calcd for
C.sub.24H.sub.30N.sub.6O.sub.2S*HCl: C, 57.30; H, 6.21; N, 16.71.
Found: C, 57.36; H, 6.31; N, 16.21.
EXAMPLE 11
N.sup.5-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-5-isoqui-
nolinesulfonamide
[0145] ##STR20##
[0146] A suspension of isoquinoline-5-sulfonyl chloride
hydrochloride (0.83 g in 50 ml of pyridine, 3.1 mmol) was added
dropwise to a stirring solution of
1-(4-aminobutyl)-2-butyl-1H-imidazo[4,5-c]quinoline-4-amine (1.0 g,
3.2 mmol) and dichloromethane (175 ml). The solution turned a
bright yellow color and was maintained at room temperature for 4
hours. An additional 0.18 g of isoquinoline-5-sulfonyl chloride
hydrochloride was added and the reaction was maintained an
additional 60 hours. The yellow solution was concentrated in vacuo,
dissolved in dichloromethane, and washed sequentially with
saturated aqueous sodium bicarbonate and water. The organic
fraction was dried (MgSO.sub.4), filtered, and concentrated in
vacuo. The residue was purified by flash column chromatography
(silica gel, 9:1 dichloromethane\methanol) to provide 0.7 g of
N.sup.5-[4-(4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-5--
isoquinolinesulfonamide as a white crystalline solid, m.p.
96.0.degree. C. (decomposition). .sup.1H NMR (300 MHz,
DMSO-d.sub.6) .delta. 9.44 (d, J=0.7 Hz, 1H), 8.64(d, J=6.1 Hz,
1H), 8.41-8.35 (m, 2H), 8.30 (dd, J=7.4, 1.2 Hz, 1H), 8.11 (t,
J=5.6 Hz, 1H), 7.92 (d, J=7.6 Hz, 1H), 7.75 (t, J=7.7 Hz, 1H), 7.61
(dd, J=8.3, 1.2 Hz, 1H), 7.41 (dt, J=7.7, 1.2 Hz, 1H), 7.22 (dt,
J=7.6, 1.2 Hz, 1H), 6.47 (broad s, 2H), 4.38 (t, J=7.5 Hz, 2H),
2.86-2.74 (m, 4H), 1.78-1.63 (m, 4H), 1.50-1.34 (m, 4H), 0.94 (t,
J=7.4 Hz, 3H); MS (E1) m/e 502.2151 (502.2151 calcd for
C.sub.27H.sub.30N.sub.6O.sub.2S). Anal calcd for
C.sub.27H.sub.30N.sub.6O.sub.2S: C, 64.52; H, 6.02; N, 16.72.
Found: C, 64.03; H, 6.03; N, 16.55.
EXAMPLE 12
N-[4-(4-Amino-2-(4-methoxybenzyl)-1H-imidazo[4,5-c]quinolin-1-yl]butyl]met-
hanesulfonamide
[0147] ##STR21##
[0148] Methanesulfonic anhydride (0.19 g, 1.1 mmol) was added to a
stirring solution of
1-(4-aminobutyl)-2-(4-methoxybenzyl)-1H-imidazo[4,5-c]quinolin-4-amine
(0.4 g, 1.07 mmol), dichloromethane (75 ml) and acetonitrile (100
ml). The reaction was maintained at room temperature for 60 hours.
The solvent was removed in vacuo and the residue was purified by
flash column chromatography (silica gel, 9:1
dichloromethane\methanol). The fractions containing product were
combined, washed with saturated aqueous sodium bicarbonate, dried
(MgSO.sub.4), filtered, and concentrated to provide 0.3 g of
N-[4-(4-amino-2-(4-methoxybenzyl)-1H-imidazo[4,5-c]quinolin-1-yl-
]butyl]methanesulfonamide as a white solid, m.p. 78.1-79.5.degree.
C. .sup.1H NMR (300 MHz, DMSO-d.sub.6) .delta. 7.99 (d, J=7.6 Hz,
1H), 7.62 (dd, J=8.3, 1.2 Hz, 1H), 7.42 (m, 1H), 7.27-7.21 (m, 3H),
6.98 (t, J=5.7 Hz, 1H), 6.89 (d, J=8.7 Hz, 2H), 6.58 (broad s, 2H),
4.45 (broad s, 2H), 4.33 (s, 2H), 3.72 (s, 3H), 2.87 (m, 5H), 1.55
(broad s, 2H); MS (CI) m/e 454 (M+H).
EXAMPLE 13
N.sup.1-[4-(4-Amino-1H-imidazo[4,5-c]quinolin-1-y)butyl]-1-butanesulfonami-
de
[0149] ##STR22##
[0150] A solution of
1-(4-aminobutyl)-1H-imidazo[4,5-c]quinolin-4-amine (9.3 mg, 36
.mu.mol) in 10 mL of dichloromethane in a screw-capped test tube
was cooled down to -5.degree. C. Butanesulfonyl chloride (45
.mu.mol) was added as a 0.3 M solution in dichloromethane, with
argon bubbling through the mixture during addition and for an
additional 15 seconds. The mixture was allowed to stand at
-5.degree. C. overnight. Aminomethyl polystyrene resin (ca. 90 mg,
0.62 meq/g, 100-200 mesh, Bachem) was added and the mixture was
warmed to reflux and shaken at about 600 rpm for 3 hours. The
mixture was filtered through a Poly-Prep column (Bio-Rad #731-1550)
to remove resin. Solvent was removed in vacuo and the residue was
purified by semi-preparative hplc on a Gilson system (Rainin
Microsorb C18 column, 21.4.times.250 mm, 8 micron particle size,
60A pore, 10 mL/min., gradient elution from 2-95% B in 25 min.,
hold at 95% B for 5 min., where A=0.1% trifluoroacetic acid/water
and B=0.1% trifluoroacetic acid/acetonitrile, peak detection at 254
nm for triggering fraction collection). The semi-prep hplc
fractions were analyzed by LC-APCI/MS and the appropriate fractions
were combined and lyophilized. The solid was dissolved in ca. 3 mL
of 2:1 dichloromethane-methanol and shaken with ca. 80 mg (300
.mu.mol) of diisopropylaminomethyl-polystyrene resin (Argonaut
PS-DIEA, 3.86 mmol/g) for .about.2 h to liberate the free amine,
and then filtered and dried in vacuo to give the product as a
solid. MS (APCI) m/e 376.16 (M+H).
EXAMPLE 14
N.sup.1-{4-[4-Amino-2-(2-methoxyethyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c-
]quinolin-1-yl]butyl}-4-fluoro-1-benzenesulfonamide
[0151] ##STR23##
[0152] According to the general method of Example
5,1-(4-aminobutyl)-2-(2-methoxyethyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c-
]quinolin-4-amine and 4-fluorobenzenesulfonyl chloride were
combined. Recrystallization from 4:1 n-propyl acetate\methanol
provided
N.sup.1-{4-[4-amino-2-(2-methoxyethyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5--
c]quinolin-1-yl]butyl}-4-fluoro-1-benzenesulfonamide as a white
crystalline solid, m.p. 191.0-193.0.degree. C. .sup.1H NMR (300
MHz, DMSO-d.sub.6) .delta. 7.86-7.81 (m, 2H), 7.67 (broad s, 1H),
7.45-7.39 (m, 2H), 5.65 (broad s, 2H), 4.15 (m, 2H), 3.76 (t, J=6.7
Hz, 2H), 3.27 (s, 3H), 3.00 (t, J=6.7 Hz, 2H), 2.90 (broad s, 2H),
2.78 (m, 2H), 2.65 (broad s, 2H), 1.75 (broad s, 4H), 1.61 (m, 2H),
1.43 (m, 2H); MS (CI) m/e 476 (M+H). Analysis: Calculated for
C.sub.23H.sub.30FN.sub.5O.sub.3S: % C, 58.09; % H, 6.36; % N,
14.73; Found: % C, 58.37; % H, 6.35; % N, 14.60.
EXAMPLE 15
N-[4-(4-Amino-2-phenyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-4-fluoro-1-be-
nzenesulfonamide
[0153] ##STR24## Part A
[0154] A solution of benzoyl chloride (5.3 g, 37.7 mmol) in
dichloromethane (100 mL) was slowly added to a solution of
tert-butyl N-{4-[(3-aminoquinolin-4-yl)amino]butyl}carbamate (12.5
g, 37.7 mmol) in dichloromethane (250 mL) at ambient temperature.
The reaction mixture was maintained at ambient temperature
overnight. The resulting precipitate was isolated by filtration and
dried to provide 11.0 g of tert-butyl
N-(4-{[3-(benzoylamino)quinolin-4-yl]amino}butyl)carbamate
hydrochloride as a white solid.
Part B
[0155] Triethylamine (7.26 g, 71.7 mmol) was added to a solution of
the material from Part A in ethanol (200 mL) and heated at reflux
for 2 days. The reaction mixture was concentrated to provide an
orange syrup. HPLC mass spec analysis showed that the syrup
contained the desired product and starting material. The syrup was
taken up in dichloromethane (100 mL) and then cooled in an ice
bath. Triethylamine (5 mL) and benzoyl chloride (1.9 mL) were
added. The reaction mixture was maintained at ambient temperature
for 2 days at which time analysis by HPLC indicated that the
reaction was not complete. The reaction mixture was concentrated
under vacuum. The residue was taken up in isopropyl alcohol (150
mL). Triethylamine (5 mL) was added and the reaction mixture was
heated at reflux overnight. The reaction mixture was concentrated
under vacuum. The residue was purified by flash chromatography
(silica gel; eluting with 10% methanol in dichloromethane). The
fractions containing product were combined and concentrated under
vacuum. The residue was recrystallized from acetonitrile to provide
6.7 g of tert-butyl
N-[4-(2-phenyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]carbamate as a
solid, m.p. 158-159.degree. C.
Part C
[0156] 3-Chloroperoxybenzoic acid (1.05 eq of 65%) was slowly added
in small portions to a solution of tert-butyl
N-[4-(2-phenyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]carbamate (6.56
g, 15.75 mmol) in dichloromethane (120 mL). After 3 hours the
reaction was quenched with 1% aqueous sodium bicarbonate (200 mL).
The layers were separated. The aqueous layer was extracted with
dichloromethane (2.times.50 mL). The organic fractions were
combined, dried over magnesium sulfate and then concentrated under
vacuum to provide a pale orange syrup. The syrup was triturated
with diethyl ether to provide 6.8 g of
1-[4-(tert-butylcarbamyl)butyl]-2-phenyl-1H-imidazo[4,5-c]quinoline--
5N-oxide as a pale tan solid, m.p. 178-181.degree. C.
Part D
[0157] A solution of
1-[4-(tert-butylcarbamyl)butyl]-2-phenyl-1H-imidazo[4,5-c]quinoline-5N-ox-
ide (6.8 g, 15.75 mmol) in dichloromethane (100 mL) was chilled in
an ice bath. Concentrated ammonium hydroxide (30 mL) was added.
Tosyl chloride (3.0 g, 15.75 mmol) was added in small portions over
a period of 30 minutes. The reaction mixture was allowed to warm to
ambient temperature overnight. The reaction was quenched with water
(350 mL). The layers were separated. The aqueous layer was
extracted with dichloromethane. The organic fractions were
combined, dried over magnesium sulfate and then concentrated under
vacuum to provide a tan solid. This material was purified by flash
chromatography (silica gel eluting with 10% methanol in
dichloromethane) to provide 4.8 g of product. The bulk of the
material was carried on to the next step. A small portion was
recrystallized from toluene to provide tert-butyl
N-[4-(4-amino-2-phenyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]carbamate
as a solid, m.p. 182-183.degree. C. Analysis: Calculated for
C.sub.25H.sub.29N.sub.5O.sub.2: % C, 69.58; % H, 6.77; % N, 16.22;
Found: % C, 69.86; % H, 6.95; % N, 15.80.
Part E
[0158] The material from Part D was dissolved in methanol (15 mL)
and 1 N hydrochloric acid (100 mL) and then heated at reflux for 2
hours. The reaction mixture was concentrated under vacuum to a
volume of about 50 mL. Addition of concentrated ammonium hydroxide
to pH 12 did not produce a precipitate. The pH was adjusted to 7
with 1 N hydrochloric acid. The mixture was extracted with
dichloromethane and then with ethyl acetate. The aqueous layer was
concentrated to dryness. The residue was dissolved in water (50 mL)
and then extracted continuously with refluxing chloroform for 36
hours. The chloroform extract was concentrated under vacuum to
provide a light tan solid. This material was recrystallized from
acetonitrile to provide 2.5 g of
1-(4-aminobutyl)-2-phenyl-1H-imidazo[4,5-c]quinolin-4-amine as an
off white solid, m.p. 175-177.degree. C. Analysis: Calculated for
C.sub.20H.sub.21N.sub.5: % C, 72.48; % H, 6.39; % N, 21.13; Found:
% C, 72.72; % H, 6.32; % N, 20.71.
Part F
[0159] 1-(4-Aminobutyl)-2-phenyl-1H-imidazo[4,5-c]quinolin-4-amine
(0.331 g, 1.0 mmol) was dissolved in anhydrous acetonitrile (35 mL)
and the solution was cooled to 4.degree. C. A solution of
4-fluorobenzenesulfonyl chloride (0.194 g, 1.0 mmol) in anhydrous
dichloromethane (10 mL) was slowly added. The reaction was allowed
to slowly warm to ambient temperature over the weekend. The
reaction was quenched by the addition of aqueous saturated sodium
bicarbonate solution. The layers were separated and the organic
layer was concentrated to provide a pale yellow solid. This
material was recrystallized from isopropyl alcohol and then further
purified by flash chromatography (silica gel eluting with 10%
methanol in dichloromethane). The pure fractions were combined and
concentrated. The residue was recrystallized from isopropyl alcohol
to provide 0.2 g of
N-[4-(4-amino-2-phenyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-4-fluoro-1-b-
enzenesulfonamide as a pale yellow solid, m.p. 214-216.degree. C.
Analysis: Calculated for C.sub.26H.sub.24FN.sub.5O.sub.2S: % C,
63.79; % H, 4.94; % N, 14.30; Found: % C, 63.19; % H, 4.85; % N,
13.90. Mass spec M+1=490.2
EXAMPLE 16
N-[4-(4-Amino-2-phenyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]methanesulfona-
mide
[0160] ##STR25##
[0161] Using the general method of Example 15 Part F,
1-(4-aminobutyl)-2-phenyl-1H-imidazo[4,5-c]quinolin-4-amine (0.331
g, 1.0 mmol) was reacted with methanesulfonic anhydride to provide
0.14 g of
N-[4-(4-Amino-2-phenyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]methanesulfon-
amide as a white solid, m.p. 234-235.degree. C. Mass spec
M+1=410.2.
EXAMPLES 17-33
[0162] The compounds shown in the Table below were prepared using
the synthetic method described in Reaction Scheme II above.
[0163] 1-(2-Aminoethyl)-2-butyl-1H-imidazo[4,5-c]quinolin-4-amine
(25 mg) was placed in a 2 dram (7.4 mL) vial. Diisopropylethylamine
(11 .mu.L, 1.2 eq), dichloromethane (1 mL) and the sulfonyl
chloride (1.1 eq) were added in order. The vial was placed on a
shaker for about 2 hours and then on a sonicator for about 0.5
hours. The reaction mixture was allowed to stand at ambient
temperature overnight and analyzed by LC/MS to confirm the
formation of the desired product. The solvent was removed and the
residue was purified by semi-preparative HPLC (Capcell Pak C18
column, 35 mm.times.20 mm, 5 micron particle size, 20 mL/min.,
gradient elution from 5-95% B in 10 min., hold at 95% B for 2 min.,
where A=0.1% trifluoroacetic acid/water and B=0.1% trifluoroacetic
acid/acetonitrile, peak detection at 254 nm for triggering fraction
collection). The semi-prep HPLC fractions were analyzed by
LC-APCI/MS and the appropriate fractions were combined and
lyophilized to provide the trifluoroacetate salt of the desired
sulfonamide. TABLE-US-00001 Example # Structure of the Free Base
Observed Mass 17 ##STR26## 390.2 18 ##STR27## 460.2 19 ##STR28##
430.1 20 ##STR29## 424.1 21 ##STR30## 504.0 22 ##STR31## 492.0 23
##STR32## 438.1 24 ##STR33## 534.0 25 ##STR34## 480.2 26 ##STR35##
466.2 27 ##STR36## 454.1 28 ##STR37## 438.1 29 ##STR38## 450.1 30
##STR39## 475.1 31 ##STR40## 474.2 32 ##STR41## 474.1 33 ##STR42##
517.2
EXAMPLE 34
N-[2-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)
ethyl]methanesulfonamide Trifluoroacetate
[0164] ##STR43##
[0165] This compound was prepared using the method of Examples
17-33 above except that 1.1 eq of methanesulfonic anhydride was
used in place of the sulfonyl chloride. (Observed Mass=362.2)
EXAMPLE 35
N-[2-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)ethyl]
trifluoromethanesulfonamide Trifluoroacetate
[0166] ##STR44##
[0167] This compound was prepared using the method of Examples
17-33 above except that 1.1 eq of trifluoromethanesulfonic
anhydride was used in place of the sulfonyl chloride. (Observed
Mass=416.1)
EXAMPLES 36-48
[0168] The compounds shown in the Table below were prepared using
the synthetic method described in Reaction Scheme II above.
[0169] 1-(4-Aminobutyl)-2-butyl-1H-imidazo[4,5-c]quinolin-4-amine
(25 mg) was placed in a 2 dram (7.4 mL) vial. Diisopropylethylamine
(14 .mu.L, 1.0 eq), dichloromethane (1 mL) and the sulfonyl
chloride (1.0 eq) were added in order. The vial was placed on a
shaker for about 30 minutes at which time almost everything was in
solution. Some time later a precipitate formed. A small amount of
methanol was added and the precipitate dissolved. The reaction
mixture was left on the shaker for an additional hour and then it
was put on a sonicator for about 0.5 hours. The reaction mixture
was analyzed by LC/MS to confirm the formation of the desired
product. The solvent was removed and the residue was purified by
semi-preparative HPLC (Capcell Pak C18 column, 35 mm.times.20 mm, 5
micron particle size, 20 mL/min., gradient elution from 5-95% B in
10 min., hold at 95% B for 2 min., where A=0.1% trifluoroacetic
acid/water and B=0.1% trifluoroacetic acid/acetonitrile, peak
detection at 254 nm for triggering fraction collection). The
semi-prep HPLC fractions were analyzed by LC-APCI/MS and the
appropriate fractions were combined and lyophilized to provide the
trifluoroacetate salt of the desired sulfonamide. TABLE-US-00002
Example # Structure of Free Base Observed Mass 36 ##STR45## 390.1
37 ##STR46## 482.1 38 ##STR47## 418.1 39 ##STR48## 452.1 40
##STR49## 466.1
EXAMPLES 41-52
[0170] The compounds shown in the Table below were prepared using
the synthetic method described in Reaction Scheme II above.
[0171] 1-(4-Aminobutyl)-2-butyl-1H-imidazo[4,5-c]quinolin-4-amine
(25 mg) was placed in a 2 dram (7.4 mL) vial. Diisopropylethylamine
(14 .mu.L, 1.0 eq), dichloromethane (1 mL) and the sulfonyl
chloride (1.0 eq) were added in order. The vial was placed on a
sonicator at ambient temperature for about 60 minutes. The reaction
mixture was analyzed by LC/MS to confirm the formation of the
desired product. The solvent was removed and the residue was
purified by semi-preparative HPLC (Capcell Pak C18 column, 35
mm.times.20 mm, 5 micron particle size, 20 mL/min., gradient
elution from 5-95% B in 10 min., hold at 95% B for 2 min., where
A=0.1% trifluoroacetic acid/water and B=0.1% trifluoroacetic
acid/acetonitrile, peak detection at 254 nm for triggering fraction
collection). The semi-prep HPLC fractions were analyzed by
LC-APCI/MS and the appropriate fractions were combined and
lyophilized to provide the trifluoroacetate salt of the desired
sulfonamide. TABLE-US-00003 Example # Structure of Free Base
Observed Mass 41 ##STR50## 502.1 42 ##STR51## 502.1 43 ##STR52##
503.2 44 ##STR53## 458.1 45 ##STR54## 494.2 46 ##STR55## 578.2 47
##STR56## 508.3 48 ##STR57## 520.1 49 ##STR58## 466.2 50 ##STR59##
478.2 51 ##STR60## 418.2 52 ##STR61## 560.1
EXAMPLE 53
N-[4-(4-Amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)butyl]
trifluoromethanesulfonamide Trifluoroacetate
[0172] ##STR62##
[0173] This compound was prepared using the method of Examples
41-52 above except that 1.0 eq of trifluoromethanesulfonic
anhydride was used in place of the sulfonyl chloride. (Observed
Mass=444.1)
EXAMPLES 54-71
[0174] The compounds shown in the Table below were prepared using
the synthetic method described in Reaction Scheme IV above.
Part A
[0175] A catalytic amount of platinum (IV) oxide was added to a
solution of 1-(4-aminobutyl)-1H-imidazo[4,5-c]quinolin-4-amine
(2.75 g, 10.8 mmol) in trifluoroacetic acid (150 mL). The reaction
mixture was placed under a hydrogen atmosphere at 50 psi
(3.44.times.10.sup.5 Pa). After 1 week analysis by mass
spectroscopy indicated the presence of both starting material and
the tetrahydro product. Fresh catalyst was added to the reaction
mixture and hydrogenation was continued at 50 psi
(3.44.times.10.sup.5 Pa). After 2 weeks the reaction mixture was
filtered to remove the catalyst. The filtrate was concentrated
under vacuum. The residue was dissolved in 1N hydrochloric acid
(120 mL) and the solution was stirred at ambient temperature for 1
hour. The solution was made basic (pH 10) by the addition of 50%
sodium hydroxide and then extracted with dichloromethane
(5.times.100 mL). The extracts were combined and concentrated under
vacuum to provide 2.08 g of
1-(4-aminobutyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-4-amine
as a white solid.
Part B
[0176]
1-(4-Aminobutyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-4-am-
ine (25 mg) was placed in a 2 dram (7.4 mL) vial.
Diisopropylethylamine (11 .mu.L, 1.2 eq), dichloromethane (1 mL)
and the sulfonyl chloride (1.1 eq) were added in order. The vial
was placed on a shaker for about 6 hours. The reaction mixture was
allowed to stand at ambient temperature overnight and was analyzed
by LC/MS to confirm the formation of the desired product. The
solvent was removed and the residue was purified by
semi-preparative HPLC (Capcell Pak C18 column, 35 mm.times.20 mm, 5
micron particle size, 20 mL/min., gradient elution from 5-95% B in
10 min., hold at 95% B for 2 min., where A=0.1% trifluoroacetic
acid/water and B=0.1% trifluoroacetic acid/acetonitrile, peak
detection at 254 nm for triggering fraction collection). The
semi-prep HPLC fractions were analyzed by LC-APCI/MS and the
appropriate fractions were combined and lyophilized to provide the
trifluoroacetate salt of the desired sulfonamide. TABLE-US-00004
Example # Structure of Free Base Observed Mass 54 ##STR63## 366.2
55 ##STR64## 366.1 56 ##STR65## 436.2 57 ##STR66## 406.1 58
##STR67## 400.1 59 ##STR68## 434.0 60 ##STR69## 468.0 61 ##STR70##
526.0 62 ##STR71## 456.1 63 ##STR72## 442 64 ##STR73## 414 65
##STR74## 430 66 ##STR75## 508.0 67 ##STR76## 414.1 68 ##STR77##
426.1 69 ##STR78## 451.1 70 ##STR79## 450.1 71 ##STR80## 493.1
EXAMPLE 72
N-[4-(4-Amino-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)butyl]meth-
anesulfonamide Trifluoroacetate
[0177] ##STR81##
[0178] This compound was prepared using the method of Examples
54-71 above except that 1.1 eq of methanesulfonic anhydride was
used in place of the sulfonyl chloride. (Observed Mass=338.2)
EXAMPLES 73-201
[0179] The compounds in the table below were prepared according the
synthetic method of Reaction Scheme II above using the following
general method.
[0180] The 1H-imidazo[4,5-c]quinolin-4-amine (50 mg) was placed in
a 2 dram (7.4 mL) vial. Diisopropylethylamine (1.2 eq) in
dichloromethane (.about.1 mL) was added. A solution containing the
sulfonyl chloride (1.1 eq) in dichloromethane (.about.1 mL) was
added. The vial was placed on a shaker for about 2-16 (usually 2)
hours at ambient temperature. The reaction mixture was analyzed by
LC/MS to confirm the formation of the desired product. The solvent
was removed and the residue was purified by semi-preparative HPLC
(Capcell Pak C18 column, 35 mm.times.20 mm, 5 micron particle size,
20 mL/min., gradient elution from 5-95% B in 10 min., hold at 95% B
for 2 min., where A=0.1% trifluoroacetic acid/water and B=0.1%
trifluoroacetic acid/acetonitrile, peak detection at 254 nm for
triggering fraction collection). The semi-prep HPLC fractions were
analyzed by LC-APCI/MS and the appropriate fractions were combined
and lyophilized to provide the trifluoroacetate salt of the desired
sulfonamide. TABLE-US-00005 Example # Structure of the Free Base
APCI-MS m/e 73 ##STR82## 526.2 74 ##STR83## 432.2 75 ##STR84##
600.3 76 ##STR85## 578.2 77 ##STR86## 530.1 78 ##STR87## 530, 532.0
79 ##STR88## 565.0 80 ##STR89## 520.1 81 ##STR90## 512.1 82
##STR91## 452.1 83 ##STR92## 497.1 84 ##STR93## 496.1 85 ##STR94##
536.1 86 ##STR95## 531.0, 533.0 87 ##STR96## 470.1 88 ##STR97##
497.1 89 ##STR98## 526.2 90 ##STR99## 542.0 91 ##STR100## 536.1 92
##STR101## 520.0, 522.0 93 ##STR102## 488.1 94 ##STR103## 471.1 95
##STR104## 470.1 96 ##STR105## 528.1 97 ##STR106## 511.1 98
##STR107## 508.1 99 ##STR108## 537.9 100 ##STR109## 516.0, 518.0
101 ##STR110## 492.0, 494.0 102 ##STR111## 603.1 103 ##STR112##
520.1 104 ##STR113## 482.1 105 ##STR114## 560.0, 562 106 ##STR115##
484.1 107 ##STR116## 522.1 108 ##STR117## 364.1 109 ##STR118##
432.0 110 ##STR119## 519.1 111 ##STR120## 392.2 112 ##STR121##
460.1 113 ##STR122## 468.2 114 ##STR123## 547.3 115 ##STR124##
406.1 116 ##STR125## 420.1 117 ##STR126## 434.1 118 ##STR127##
454.1 119 ##STR128## 468.1 120 ##STR129## 472.1 121 ##STR130##
472.1 122 ##STR131## 472.1 123 ##STR132## 473.1 124 ##STR133##
484.1 125 ##STR134## 484.1 126 ##STR135## 488.1 127 ##STR136##
488.1 128 ##STR137## 488.0 129 ##STR138## 490.1 130 ##STR139##
490.1 131 ##STR140## 494.0 132 ##STR141## 496.2 133 ##STR142##
496.1 134 ##STR143## 496.2 135 ##STR144## 499.1 136 ##STR145##
499.1 137 ##STR146## 508.1 138 ##STR147## 513.1 139 ##STR148##
514.1 140 ##STR149## 514.1 141 ##STR150## 518.0 142 ##STR151##
522.1 143 ##STR152## 522.0, 524.0 144 ##STR153## 522.0, 524.0 145
##STR154## 522.0, 524.0 146 ##STR155## 522.0, 524.0 147 ##STR156##
522.0, 524.0 148 ##STR157## 528.2 149 ##STR158## 528.0, 530.0 150
##STR159## 528.0, 530.0 151 ##STR160## 532, 534.0 152 ##STR161##
532, 534.0 153 ##STR162## 538.1 154 ##STR163## 538.1 155 ##STR164##
538, 540.0 156 ##STR165## 580.0 157 ##STR166## 605.1 158 ##STR167##
454.2 159 ##STR168## 468.2 160 ##STR169## 479.2 161 ##STR170##
532.2 162 ##STR171## 479.1 163 ##STR172## 486.1 164 ##STR173##
490.2 165 ##STR174## 498.1 166 ##STR175## 498.1 167 ##STR176##
502.1 168 ##STR177## 502.1 169 ##STR178## 504.2 170 ##STR179##
504.1 171 ##STR180## 505.2 172 ##STR181## 506.1 173 ##STR182##
506.2 174 ##STR183## 506.2 175 ##STR184## 510.3 176 ##STR185##
510.2 177 ##STR186## 513.2 178 ##STR187## 513.2 179 ##STR188##
513.2 180 ##STR189## 524.2 181 ##STR190## 526.2 182 ##STR191##
530.2 183 ##STR192## 532.2 184 ##STR193## 534.1 185 ##STR194##
533.1 186 ##STR195## 536.1, 538.1 187 ##STR196## 544.1
188 ##STR197## 546.3 189 ##STR198## 556, 558.1 190 ##STR199## 556,
558.1 191 ##STR200## 556, 558.1 192 ##STR201## 562, 564.1 193
##STR202## 567.2 194 ##STR203## 580.3 195 ##STR204## 593.2 196
##STR205## 606.0, 608.0, 609.9 197 ##STR206## 610.0, 612.0, 614.0
198 ##STR207## 616, 618.1 199 ##STR208## 616.0, 617.9, 620.0 200
##STR209## 528.3 201 ##STR210## 522.2
EXAMPLES 202-213
[0181] The examples in the table below were prepared according to
the synthetic method of Reaction Scheme VI above.
Part A
[0182] The tetrahydroquinoline amine starting materials were
prepared as follows.
[0183] A catalytic amount of platinum (IV) oxide was added to a
solution of
1-(4-aminobutyl)-2-butyl-1H-imidazo[4,5-c]quinolin-4-amine (2.2 g,
7.06 mmol) in trifluoroacetic acid (200 mL). The reaction mixture
was hydrogenated at 50 psi (3.44.times.10.sup.5 Pa) on a Parr
apparatus for 6 days. The reaction mixture was filtered to remove
the catalyst and the filtrate was concentrated under vacuum. The
residue was combined with 1 N hydrochloric acid (100 mL) and heated
on a steam bath for 2 hours. The mixture was cooled, made basic
with ammonium hydroxide and then extracted with dichloromethane.
The extract was concentrated under vacuum to provide of
1-(4-aminobutyl)-2-butyl-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-4-a-
mine as a solid, m.p. 63-67.degree. C.
[0184] A catalytic amount of platinum (IV) oxide was added to a
solution of
1-(4-aminobutyl)-2-methoxyethyl-1H-imidazo[4,5-c]quinolin-4-amine
(7.7 g, 24.5 mmol) in trifluoroacetic acid (250 mL). The reaction
mixture was hydrogenated at 50 psi (3.44.times.10.sup.5 Pa) on a
Parr apparatus. The progress of the reaction was monitored by
LC/MS. Additional catalyst was added 7, 11, and 17 days after the
start of the reaction. After 25 days the reaction was complete. The
reaction mixture was filtered through a layer of Celite.RTM. filter
aid to remove the catalyst and the filtrate was concentrated under
vacuum. The residue was combined with 1 N hydrochloric acid (100
mL) and stirred overnight. The mixture was made basic (pH=11) with
ammonium hydroxide and then extracted with dichloromethane
(3.times.300 mL). The extracts were combined and concentrated under
vacuum to provide 3.5 g of
1-(4-aminobutyl)-6,7,8,9-tetrahydro-2-methoxyethyl-1H-imidazo[4,5c]quinol-
in-4-amine as a solid.
Part B
[0185] The tetrahydroimidazoquinoline amines from Part A were
reacted with the appropriate sulfonyl chloride using the method of
Examples 73-201 above to provide the desired sulfonamide.
TABLE-US-00006 Example # Structure of the Free Base APCI-MS m/e 202
##STR211## 394.20 203 ##STR212## 422.1 204 ##STR213## 462.1 205
##STR214## 470.1 206 ##STR215## 549.2 207 ##STR216## 410.2 208
##STR217## 424.2 209 ##STR218## 438.2 210 ##STR219## 458.1 211
##STR220## 472.2 212 ##STR221## 532.2 213 ##STR222## 551.2
EXAMPLE 214
N-[4-(4-Amino-6,7,8,9-tetrahydro-2-(2-methoxyethyl)-1H-imidazo[4,5-c]quino-
lin-1-yl)butyl]methanesulfonamide Trifluoroacetat
[0186] ##STR223##
[0187] This compound was prepare using the method of Examples
202-213 above except that methanesulfonic anhydride was used in
place of the sulfonyl chloride.
EXAMPLE 215
N-[4-(4-Amino-2-(2-methoxyethyl)-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-N-m-
ethyl-3,5-dimethylisooxazolo-4-sulfonamide Trifluoroacetate
[0188] ##STR224## Part A
[0189] Using the general method of Example DC001,
1-(4-aminobutyl)-2-(2-methoxyethyl)-1H-imidazo[4,5-c]quinolin-4-amine
was reacted with 3,5-dimethyloxazole-4-sulfonyl chloride to provide
N-[4-(4-amino-2-(2-methoxyethyl)-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-3,-
5-dimethylisooxazolo-4-sulfonamide trifluoroacetate.
Part B
[0190] Sodium hydride (5.8 mg) was added to a solution of the
material from Part A (25.4 mg) in dimethylformamide. Iodomethane
(3.2 .mu.L) was added and the reaction mixture was shaken at
ambient temperature for 2 hours. The reaction mixture was analyzed
by LC/MS to confirm the formation of the desired product. The
solvent was removed and the residue was purified by
semi-preparative HPLC (Capcell Pak C18 column, 35 mm.times.20 mm, 5
micron particle size, 20 mL/min., gradient elution from 5-95% B in
10 min., hold at 95% B for 2 min., where A=0.1% trifluoroacetic
acid/water and B=0.1% trifluoroacetic acid/acetonitrile, peak
detection at 254 nm for triggering fraction collection). The
semi-prep HPLC fractions were analyzed by LC-APCI/MS and the
appropriate fractions were combined and lyophilized. The
lyophilized material was purified a second time by semi-preparative
HPLC using the same conditions except that the gradient elution
from 5-95% B was run for 60 minutes instead of 10 minutes. The
semi-prep HPLC fractions were analyzed by LC-APCI/MS and the
appropriate fractions were combined and lyophilized to provide the
trifluoroacetate salt of the desired amide.
EXAMPLE 216
N-[4-(4-Amino-2-(2-methoxyethyl)-1H-imidazo[4,5-c]quinolin-1-yl)butyl]-N-m-
ethyltrifluoromethanesulfonamide Trifluoroacetate
[0191] ##STR225##
[0192] This compound was prepared using the general method of
Example 215 above, except that trifluoromethanesulfonic anhydride
was used in place of the sulfonyl chloride in Part A.
EXAMPLES 217-221
[0193] The examples in the table below were prepare using the
following general method. The 1H-imidazo[4,5-c]quinolin-4-amine or
the 6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-4-amine (50 mg)
was placed in a 2 dram (7.4 mL) vial. Dichloromethane (2 mL) and
diisopropylethylamine (1.2 eq) were added. Dimethylsulfamoyl
chloride (1.1 eq) was added. The vial was placed on a shaker for
about 2-4 hours at ambient temperature. The reaction mixture was
analyzed by LC/MS to confirm the formation of the desired product.
The solvent was removed and the residue was purified by
semi-preparative HPLC (Capcell Pak C18 column, 35 mm.times.20 mm, 5
micron particle size, 20 mL/min., gradient elution from 5-95% B in
10 min., hold at 95% B for 2 min., where A=0.1% trifluoroacetic
acid/water and B=0.1% trifluoroacetic acid/acetonitrile, peak
detection at 254 nm for triggering fraction collection). The
semi-prep HPLC fractions were analyzed by LC-APCI/MS and the
appropriate fractions were combined and lyophilized to provide the
trifluoroacetate salt of the desired sulfamide. TABLE-US-00007
Example # Structure of the Free Base APCI-MS m/e 217 ##STR226##
393.1 218 ##STR227## 421.2 219 ##STR228## 483.3 220 ##STR229##
423.2 221 ##STR230## 425.1
EXAMPLES 222-228
[0194] The examples in the table below were prepared according to
the synthetic method shown in Reaction Scheme V above.
[0195]
1-(4-Aminobutyl)-2-(2-methoxyethyl)-1H-imidazo[4,5-c]quinolin-4-am-
ine (50 mg) was placed in a 2 dram (7.4 mL) vial.
4-(Dimethylamino)pyridine (19 mg, 1.0 eq) and dichloromethane (800
.mu.L) were added. The vial was sealed and cooled to -78.degree. C.
in a dry ice/acetone bath. Sulfuryl chloride (186 .mu.L of 1 M in
dichloromethane) was added. The vial was put on a shaker for about
30 minutes and then cooled back down to -78.degree. C. A separate
vial was charged with the amine of formula R.sub.4R.sub.5NH (2.0
eq), triethylamine (2.0 eq) and dichloromethane (1 mL) and cooled
to -78.degree. C. The amine/triethylamine solution was added to the
first vial. The vial was placed on a shaker at ambient temperature
for about 1 hour. The reaction mixture was analyzed by LC/MS to
confirm the formation of the desired product. The solvent was
removed and the residue was purified by semi-preparative HPLC
(Capcell Pak C18 column, 35 mm.times.20 mm, 5 micron particle size,
20 mL/min., gradient elution from 5-95% B in 10 min., hold at 95% B
for 2 min., where A=0.1% trifluoroacetic acid/water and B=0.1%
trifluoroacetic acid/acetonitrile, peak detection at 254 nm for
triggering fraction collection). The semi-prep HPLC fractions were
analyzed by LC-APCI/MS and the appropriate fractions were combined
and lyophilized to provide the trifluoroacetate salt of the desired
sulfamide. TABLE-US-00008 Example # Structure of the Free Base
APCI-MS m/e 222 ##STR231## 449.2 223 ##STR232## 475.3 224
##STR233## 469.1 225 ##STR234## 490.2 226 ##STR235## 497.1 227
##STR236## 533.2 228 ##STR237## 479.1
EXAMPLES 229-231
[0196] The examples in the table below were prepared using the
method of Examples 222-228 above except that the amine of formula
R.sub.4R.sub.5NH was reacted with the sulfuryl chloride to provide
the sulfamoyl chloride intermediate which was then reacted with 2.0
eq of
1-(4-aminobutyl)-2-(2-methoxyethyl)-1H-imidazo[4,5-c]quinolin-4-amine.
TABLE-US-00009 Example # Structure of the Free Base APCI-MS m/e 229
##STR238## 447.1 230 ##STR239## 449.2 231 ##STR240## 483.2
Cytokine Induction in Human Cells
[0197] An in vitro human blood cell system was used to assess
cytokine induction by compounds of the invention. Activity is based
on the measurement of interferon and tumor necrosis factor
(.alpha.) (IFN and TNF, respectively) secreted into culture media
as described by Testerman et. al. In "Cytokine Induction by the
Immunomodulators Imiquimod and S-27609", Journal of Leukocyte
Biology, 58, 365-372 (September, 1995).
Blood Cell Preparation for Culture
[0198] Whole blood is collected by venipuncture into EDTA
vacutainer tubes from healthy human donors. Peripheral blood
mononuclear cells (PBMCs) are separated from whole blood by density
gradient centrifugation using Histopaque.RTM.-1077 (Sigma
Chemicals, St. Louis, Mo.). The PBMCs are suspended at
3-4.times.10.sup.6 cells/mL in RPMI 1640 medium containing 10%
fetal bovine serum, 2 mM L-glutamine and 1% penicillin/streptomycin
solution (RPMI complete). The PBMC suspension is added to 48 well
flat bottom sterile tissue culture plates (Costar, Cambridge, Mass.
or Becton Dickinson Labware, Lincoln Park, N.J.) containing an
equal volume of RPMI complete media containing test compound.
Compound Preparation
[0199] The compounds are solubilized in dimethyl sulfoxide (DMSO).
The DMSO concentration should not exceed a final concentration of
1% for addition to the culture wells.
Incubation
[0200] The solution of test compound is added at 60 .mu.M to the
first well containing RPMI complete and serial (three fold or ten
fold) dilutions are made. The PBMC suspension is then added to the
wells in an equal volume, bringing the test compound concentrations
to the desired range. The final concentration of PBMC suspension is
1.5-2.times.10.sup.6 cells/mL. The plates are covered with sterile
plastic lids, mixed gently and then incubated for 18 to 24 hours at
37.degree. C. in a 5% carbon dioxide atmosphere.
Separation
[0201] Following incubation the plates are centrifuged for 5-10
minutes at 1000 rpm (.about.200.times.g) at 4.degree. C. The cell
culture supernatant is removed with a sterile polypropylene pipet
and transferred to sterile polypropylene tubes. Samples are
maintained at -30 to -70.degree. C. until analysis. The samples are
analyzed for interferon (.alpha.) and tumor necrosis factor
(.alpha.) by ELISA
Interferon (.alpha.) and Tumor Necrosis Factor (.alpha.) Analysis
by ELISA
[0202] Interferon (.alpha.) concentration is determined by ELISA
using a Human Multi-Species kit from PBL Biomedical Laboratories,
New Brunswick, N.J.
[0203] Tumor necrosis factor (.alpha.) (TNF) concentration is
determined using ELISA kits available from Genzyme, Cambridge,
Mass.; R&D Systems, Minneapolis, Minn.; or Pharmingen, San
Diego, Calif.
[0204] The table below lists the lowest concentration found to
induce interferon and the lowest concentration found to induce
tumor necrosis factor for each compound. A "**" indicates that no
induction was seen at any of the tested concentrations (0.12, 0.37,
1.11, 3.33, 10 and 30 .mu.M). A "***" indicates that no induction
was seen at any of the tested concentrations (0.0001, 0.001, 0.01,
0.1, 1 and 10 .mu.M). TABLE-US-00010 Cytokine Induction in Human
Cells Lowest Effective Example Concentration (.mu.M) Number
Interferon Tumor Necrosis Factor 1 0.12 3.33 2 ** ** 3 0.01 ** 6
0.00017 1.11 7 0.01 ** 9 0.04 ** 11 0.01 1.11 13 10 ** 17 1.11 3.33
18 3.33 ** 19 0.12 3.33 20 0.12 3.33 21 1.11 30 22 0.37 ** 23 0.12
10 24 0.12 30 25 3.33 ** 26 10 ** 27 1.11 30 28 1.11 30 29 0.37 10
30 1.11 ** 31 1.11 ** 32 1.11 ** 33 1.11 10 34 0.04 0.37 35 1.11 10
36 0.0015 3.33 37 0.01 1.11 38 0.0015 0.37 40 0.0015 3.33 41 0.01
** 42 0.01 ** 43 0.04 ** 44 0.0015 1.11 45 0.37 ** 46 0.37 ** 47
0.37 ** 48 0.37 10 50 0.12 ** 51 0.0015 0.37 52 0.12 10 53 0.01
3.33 54 10 ** 55 3.33 ** 56 ** ** 57 3.33 ** 58 3.33 ** 59 3.33 **
60 ** ** 61 3.33 ** 62 ** ** 63 ** ** 64 3.33 ** 65 3.33 ** 66 **
30 67 10 ** 68 10 ** 69 10 ** 70 ** ** 71 ** 30 72 3.33 ** 73 0.001
0.1 74 0.001 0.01 75 *** *** 76 *** *** 77 0.001 1 78 0.001 0.1 79
0.01 1 80 1 10 81 0.001 1 82 0.001 1 83 0.001 1 84 1 10 85 1 *** 86
0.01 1 87 0.001 1 88 0.01 1 89 0.001 1 90 0.01 1 91 0.01 1 92 0.1
10 93 0.001 0.1 94 0.001 1 95 0.001 1 96 1 *** 97 0.1 10 98 1 ***
99 0.1 10 100 0.01 10 101 0.01 10 102 0.001 10 103 0.1 10 104 0.01
*** 105 1 10 106 1 1 107 1 *** 108 0.1 10 109 1 10 110 10 *** 111
0.001 10 112 0.0001 *** 113 0.0001 *** 114 0.01 *** 116 0.001 1 117
0.0001 1 120 0.0001 1 121 0.0001 10 122 0.0001 1 123 0.0001 10 127
0.0001 10 128 0.0001 1 131 0.0001 1 138 0.0001 10 148 0.0001 1 152
0.0001 10 154 0.001 10 158 0.0001 1 159 0.0001 0.1 160 0.001 1 161
0.01 10 184 0.0001 1 200 0.01 0.1 202 0.0001 1 203 0.0001 1 204
0.0001 1 205 0.0001 1 206 1 *** 207 0.001 1 208 0.0001 1 209 0.0001
0.1 210 0.0001 1 211 0.0001 1 212 0.0001 0.01 213 0.0001 1 214 0.01
10 215 0.01 1 217 1 *** 218 0.0001 1 220 0.0001 1 221 0.0001 1 224
0.0001 10 226 0.0001 0.1 227 0.001 *** 229 0.0001 0.1 230 0.0001 1
231 0.0001 1
[0205] The present invention has been described with reference to
several embodiments thereof. The foregoing detailed description and
examples have been provided for clarity of understanding only, and
no unnecessary limitations are to be understood therefrom. It will
be apparent to those skilled in the art that many changes can be
made to the described embodiments without departing from the spirit
and scope of the invention. Thus, the scope of the invention should
not be limited to the exact details of the compositions and
structures described herein, but rather by the language of the
claims that follow.
* * * * *