Note : Les descriptions sont présentées dans la langue officielle dans laquelle elles ont été soumises.
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HYDRAZONE DERIVATIVES AND THEIR USE AS BETA SECRETASE INHIBITORS
The present invention relates to novel hydrazone derivatives and the
use of such hydrazone derivatives as beta-secretase inhibitors.
Alzheimer's disease (AD) is a very common form of dementia in humans
and affects parts of the brain that control inter alia memory and
language.
The occurrence of AD is associated with protein clusters in the
brain, also termed "amyloid plaques" and "neurofibrillary tangles".
These protein clusters comprise the (3-amyloid precursor protein ((3-
APP). (3-APP is degraded by the (3-Amyloid Converting Enzyme (BACE,
also known under the name of beta-secretase) to produce (3-amyloid
peptide which is the main component of these clusters of amyloid
plaques. It has been found out that the activity of BACE is an early
step in the pathogenesis pathway for AD.
It has therefore been an object of the present invention to provide
new compounds which serve as inhibitors for BACE and are thus thera-
peutically useful in the treatment of AD.
This object is solved by providing hydrazone derivatives which act
as beta-secretase inhibitors having the general formula I
R3 H
Z1 NN\Z 2 I
wherein R3 is selected from the group consisting of H, methyl
and hydroxyalkyl,
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and wherein Z1 and Z2 are selected independently from one an-
other from the group consisting of substituted or unsubstituted
phenyl, naphtyl, pyridinyl, pyrazolyl, pyrimidinyl, pyrazid-
inyl, quinolinyl, iso-quinolinyl, coumarinyl, indolyl, thia-
zolyl and thiophenyl groups bearing substituents n R1 and m R2,
wherein R,_ and R2 which may be the same or are different from
one another are selected from the group consisting of H, alkyl,
cycloalkyl, -C02R4, -CONHR4, -CR40, -S02R4, -NR4-CO-R4, alkoxy,
alkylthio, -OH, -0-aryl, -0-cycloalkyl, -S-aryl, -S-cycloalkyl,
hydroxyalkyl, halogen, haloalkyl, haloalkoxy, -CN, -NOZ, hy-
droxyalkylamine, aminoalkyl, alkylamine, aryl, heteroaryl and
sulfonamide,
wherein R4 is selected from the group consisting of H, C1 to C6
branched or unbranched alkyl, aryl, cycloalkyl, alkoxy, hetero-
cycloalkyl, -OH, -0-aryl, -0-alkyl, -0-cycloalkyl, aminoalkyl,
alkylamine, aryl and heteroaryl,
wherein n and m are the numbers of substituents R1 and R2 in Z1
and Z2, respectively, which are in a range between 0 and 5,
with the proviso that if n and m are integers higher than 2, R1
and/or R2 may form a fused aromatic or a carbocyclic or hetero-
cyclic ring system,
or a salt, or a physiologically functional derivative, or a
prodrug thereof,
whereas the compounds
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H
CI ,N \
OH
CI
H O~N+;O
I
CI
O", + / F
OH N
CI p F
H CI
CI ,N
I
COH CI
CI
are not comprised within formula I.
The object is further solved by providing hydrazone derivatives
which act as beta-secretase inhibitors having the general formula I
H
R3
z Z2
wherein
Z1 and Z2 are selected independently from one another from the
group consisting of substituted or unsubstituted cyclic and
acyclic alkyl, phenyl, naphthyl, pyridinyl, pyrrolyl, pyra-
zolyl, pyrimidinyl, pyrazidinyl, quinolinyl, iso-quinolinyl,
coumarinyl, indolyl, triazinyl (comprising 1,2,4 and 1,3,5 tri-
azinyl), imidazolyl, thiazolyl, thiophenyl and oxazolyl groups
bearing substituents (R1)n and (R2)m,
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wherein the number of substituents in Zl and Z2 ,(Rl)n and (R2)m
is in a range between 0 and 5 and
R1, R2are selected from the group consisting of H, alkyl,
cycloalkyl, -C02R4, -CONHR4, -CR40, -S02R4, -NR4-CO-R4, alkoxy,
alkylthio, -OH, -SH, -0-aryl, -0-cycloalkyl, -S-aryl, -S-
cycloalkyl, hydroxyalkyl, halogen, haloalkyl, haloalkoxy, CN,
NOz, hydroxyalkylamine, aminoalkyl, alkylamine, aryl or het-
eroaryl, sulfone and sulfonamide
whereby, R, and R2 may be the same or different from one another
and wherein in the case that n and m are integers higher than
2, R1 and/or R2 may form a fused aromatic or a carbocyclic or
heterocyclic ring system
R3 is selected from the group consisting of H, C1 to C6
branched or, unbranched alkyl, aryl, heteroaryl; haloalkyl,
haloalkoxy, hydroxyalkyl, cycloalkyl and heterocycloalkyl.
R4 is selected from the group consisting of H, C1 to C6
branched or unbranched alkyl, aryl, cycloalkyl, alkoxy, hetero-
cycloalkyl, -OH, -0-aryl, -0-alkyl, -0-cycloalkyl, aminoalkyl,
alkylamine, aryl or heteroaryl;
and wherein compound
H
CI
OH
CI
is not comprised within formula I,
or a salt, or a physiologically functional derivative, or a
prodrug thereof.
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The object is further solved by providing hydrazone derivatives
which act as beta-secretase inhibitors having the general formula I
R3 H
I
Z/ 'N"' N~Z
1 2 I
wherein R3 is selected from the group consisting of H, methyl
and hydroxyalkyl,
and wherein Zs, and Z2 are selected independently from one an-
other from the group consisting of substituted or unsubstituted
phenyl, naphtyl, pyridinyl, pyrazolyl, pyrimidinyl, pyrazid-
inyl, quinolinyl, iso-quinolinyl, coumarinyl, indolyl, thia-
zolyl and thiophenyl groups bearing substituents n R1 and m R2,
wherein Rl and R2 which may be the same or are different from
one another are selected from the group consisting of H, alkyl,
cycloalkyl, -C02R4, -CONHR4, -CR40, -S02R4, -NR4-CO-R4, alkoxy,
alkylthio, -OH, -0-aryl, -0-cycloalkyl, -0-alkyl-aryl, -S-aryl,
-S-cycloalkyl, hydroxyalkyl, halogen, haloalkyl, haloalkoxy, -
CN, -NOz, hydroxyalkylamine, aminoalkyl, alkylamine, aryl, het-
eroaryl and sulfonamide,
wherein R4 is selected from the group consisting of H, C1 to C6
branched or unbranched alkyl, aryl, cycloalkyl, alkoxy, hetero-
cycloalkyl, -OH, -0-aryl, -0-alkyl, -0-cycloalkyl, aminoalkyl,
alkylamine, aryl and heteroaryl,
wherein n and m are the numbers of substituents Rl and R2 in Za,
and Z2, respectively, which are in a range between 0 and 5,
with the proviso that if n and m are integers higher than 2, R,
and/or R2 may form a fused aromatic or a carbocyclic or hetero-
cyclic ring system,
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or a salt, or a physiologically functional derivative, or a
prodrug thereof,
whereas the compounds
H
N \I /
CI *O'HN
CI
H O 'N+'O
I
CI N,N
COH O. N + I
CI p F F
H CI
V
CI N~N
( I iN
OH CI
CI
are not comprised within formula I.
It is understood that general formula I comprises the corresponding
isomeric forms, i.e. the E and the Z form thereof.
Examples of such salts are, for example, alkali metal salts, in par-
ticular sodium and potassium salts, or ammonium salts but not lim-
ited thereto.
It is understood that the compounds according to the invention may
contain one or more asymmetric carbon atoms in R or S configuration
and thus the compounds may be present as racemates or racemic mix-
tures, pure enantiomers, diastereomic mixtures etc.
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In a preferred embodiment, R4 is selected from the group consisting
of H, C1 to C4 branched or unbranched alkyl, aryl, cycloalkyl,
alkoxy, heterocycloalkyl, -OH, -0-aryl, -0-alkyl, -0-cycloalkyl,
aminoalkyl, alkylamine, aryl and heteroaryl.
In another preferred embodiment, Z, is a substituted phenyl group
which is in a still more preferred embodiment substituted by at
least one halogen atom or at least one hydroxyl group like in the
examples no. 3, 18, 19, 20, 21, 23-26, 28-31, 33, 35, 39, 40, 46,
47, 55, 57-60, 63, 66-68, 75-77, 86-88, 91-96, 98, 100-104, 108,
111, 113, 114, 118 and 119.
In a further preferred embodiment of the invention Z. is a substi-
tuted phenyl group which is substituted by at least two halogen at-
oms and one hydroxyl group and in another preferred embodiment of
the invention the hydroxyl group is in o-position relative to the
bonding position of the hydrazone motif.
Aditionally, in a preferred embodiment of the invention Z1 is a sub-
stituted phenyl group which bears at least a substituted or unsub-
stituted aryl or heteroaryl ring or one or two terbutyl groups.
In another preferred embodiment, the phenyl group (Z1) is substi-
tuted by another phenyl or heteroaryl group which are directly
linked to the phenyl group (Z1) or via an ether bond. In a further
preferred embodiment, the phenyl group is part of a fused ring sys-
tem.
In preferred embodiments, Z1 is a substituted phenyl group which is
in a still more preferred embodiment substituted by at least one
halogen atom or at least one hydroxy group.
Further, in another preferred embodiment of the invention Z1 bears
two or three substituents which may be the same or different from
one another, preferentially one substituent is in o-position rela-
tive to the bonding position of the hydrazone motif.
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In another preferred embodiment Z2 is selected from the group con-
sisting of substituted or unsubstituted phenyl, pyridinyl and
pyrimidinyl groups.
In a further preferred embodiment, Z2 is phenyl group which is pref-
erably substituted by a benzyl group that is linked to the phenyl
group via an ether bond like in the examples no. 4-6, 18-21 and 65.
Further, in a preferred embodiment of the invention the benzyl group
which is linked to the phenyl group via an ether bond is a substi-
tuted or unsubstituted benzyloxy group.
In an additional preferred embodiment, Z2 is pyrimidyl ring bearing a
substituted or unsubstitued cycloalkyl group or a disubstituted
amino group like in the examples no. 3, 31, 74, 76-84, 86-102, 105-
110 and 112.
In a further preferred embodiment, Z2 is a pyridyl ring bearing a
trifluoromethyl group, like in the examples no. 114-118.
Additionally, in another preferred embodiment Z2 is an unsubstituted
phenyl group.
In another preferred embodiment Z2 is a phenyl group bearing at least
one halogen substituent.
In a further preferred embodiment of the invention Z2 is a phenyl
group bearing at least two halogen substituents, which may be the
same or different from one another.
In another preferred embodiment of the invention Z2 is a phenyl group
bearing at least one substituent selected from the group consisting
of methyl, trifluoromethyl, hydroxy, carboxyl or an ether group
which is in a still more preferred embodiment a substituted or un-
,substituted benzyloxy group.
In another preferred embodiment one substituent in Z2 is in o-
position relative to the bonding position of the hydrazone motif.
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Additionally, in a further preferred embodiment the compound has one
of the following formulas
H2Ny NH Oy NH2
N NN N N N
ZN" N I / Z,N, N
H H
L0
The invention further relates to a compound of the formula
GI
OH
N\
CI N
H
L5 Other preferred compounds are those listed in table 1.
Other preferred compounds are those listed in table 2.
The invention further relates to the use of the compounds according
20 to the invention as a medicament.
The invention further relates to the use of the compounds according
to the invention for the manufacture of a pharmaceutical composition
for the treatment or prevention of a condition or disorder which is
25 mediated by beta-secretase. The compounds according to the invention
are useful as a medicament, especially as beta-secretase inhibitors.
Beta-secretase is inhibited very effectively and the compounds ac-
cording to the invention have IC50 values of less than 120 M, more
preferred of less than 100 M, still more preferred less than 50 M
30 and most preferred less than 20 pM. The most potent compounds ac-
cording to the invention have IC50 values of less than 10 M, more
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5 preferred of less than 8 M and most preferred less than 6 M like
for example compounds no. 60, 61, 74-81.
The invention further relates to the use of the compounds according
to the invention for the manufacture of a pharmaceutical composition
10 to inhibit the formation of beta-amyloid peptides from the amyloid
precursor protein APP. The compounds according to the invention are
well suited for the inhibition of the formation of beta-amyloid pep-
tides from the amyloid precursor protein APP. Therefore, the com-
pounds according to the invention are suitable for the manufacture
of a pharmaceutical composition for the treatment or prevention of
conditions or diseases relying on beta-amyloid peptide formation as
Alzheimer's disease, Down syndrome, cerebral amyloid angiopathy, He-
reditary Cerebral Haemorrhage with Amylosis of the Dutch type
(HCHWA-D) and other degenerative dementia.
The invention further relates to the use of the compounds according
to the invention for the manufacture of a pharmaceutical composition
for the treatment or prevention of Alzheimer's disease, neurodegen-
eration, memory and attention deficits, dysfunction of cellular pro-
liferation, or other disorders associated with or responsive to the
inhibition of beta-amyloid peptides.
The invention further relates to the use of the compounds according
to the invention for the manufacture of a pharmaceutical composition
for the treatment or prevention of conditions or diseases selected
from the group consisting of Alzheimer's disease, Down syndrome,
cerebral amyloid angiopathy, Hereditary Cerebral Haemorrhage with
Amylosis of the Dutch type (HCHWA-D) and other degenerative dementia
characterized by beta-amyloid deposits.
The invention further relates to a pharmaceutical composition com-
prising a compound according to the invention or a pharmaceutically
acceptable salt thereof. In one preferred embodiment the pharmaceu-
.tical composition comprises a pharmaceutically acceptable carrier,
diluent, and/or excipient.
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The compounds according to the invention may be administered alone
or in the form of a pharmaceutically acceptable salt thereof. A
pharmaceutical composition comprising a compound according to the
invention or a pharmaceutically active salt thereof may further com-
prise at least one pharmaceutically acceptable carrier, diluent or
excipient. It is understood that in specific embodiments also fur-
ther active compounds are contained within the composition.
The compounds according to the invention may be formulated for topi-
cal, oral, transdermal, parenteral, sublingual, intranasal, in-
trathecal, rectal, inhalative or intravenous administration in form
of e.g. tablets, gel, capsules, patches, ointments, creams. Par-
enteral delivery can be carried out by depot, syringe, ampoule or
vial.
The compounds of the invention, together with a conventional adju-
vant, carrier, or diluent, may thus be placed into the form of phar-
maceutical compositions and unit dosages thereof, and in such form
may be employed as solids, liquids or in the form of sterile in-
jectable solutions. If a solid carrier is used, the preparation may
be tableted, placed in a hard gelatine capsule in powder or pellet
form, or in form of a troche or lozenge. The solid carrier may con-
tain conventional excipients such as binding agents, tableting lu-
bricants, fillers, disintegrants, wetting agents and the like. Tab-
lets may be film coated by conventional techniques. If a liquid
carrier is employed, the preparation may be in form of syrup, emul-
sion, soft gelatine capsule, sterile vehicle for injection, an aque-
ous or non-aqueous liquid suspension, or may be a dry product for
reconstitution with water or other suitable vehicles before use.
Liquid preparations may contain conventional additives such as sus-
pending agents, emulsifying agents, wetting agents, non-aqueous ve-
hicle (including edible oils), preservatives, as well as flavouring
and /or colouring agents. For parenteral administration, a vehicle
normally will comprise sterile water, at least in large part, al-
though saline solutions, glucose solutions and like may be utilized.
Injectable suspensions also may be used, in which case conventional
suspending agents may be employed. Conventional preservatives, buff-
ering agents and the like also may be added to the parenteral dosage
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forms. Administration, however, can also be carried out rectally,
e.g., in the form of suppositories, or vaginally, e.g. in the form
of pessaries, tampons, creams, or percutaneously, e.g., in the form
of ointments, creams or tinctures.
A suitable dose of compounds or pharmaceutical compositions accord-
ing to the invention for a mammal, especially humans, suffering
from, or likely to suffer from any condition as described herein is
an amount of active ingredient from about 0.1jig/kg (NB: there are
few example of compounds that are delivered at lower dosis, eg some
hormone) to 500mg/kg body weight. For parenteral administration, the
dose may be in the range of 0.1}a.g/kg to 100mg/kg body weight for in-
travenous administration. The active ingredient will preferably be
administered in equal doses from one to four times daily. The com-
pounds of Formula (I) can also be used in the form of a precursor
(prodrug) or a suitably modified form that releases the active com-
pound in vivo. Normally, the administered dose will be gradually in-
creased until the optimal effective dosage for the treated host is
determined. The optimal administered dosage will be determined by a
physician or others skilled in the art, depending on the relevant
circumstances including the condition to be treated, the choice of
compound to be administered, the route of administration, the sex,
age, weight, and the specific response of the treated individual in
respect to the severity of the individual's symptoms.
The pharmaceutical compositions are prepared by conventional tech-
niques appropriate to the desired preparation containing appropriate
amounts of the active ingredient, i.e., the compounds of the present
invention. Such pharmaceutical compositions and unit dosage forms
thereof may comprise conventional ingredients in conventional pro-
portions, with or without additional active compounds or principles,
and such unit dosage forms may contain any suitable effective amount
of the active ingredient commensurate with the intended daily dosage
range to be employed.
The compounds of the invention are 'prepared by processes known in
the art for the synthesis of hydrazones. Especially preferred is the
reaction of an aldehyde or ketone of Formula II with a compound of
Formula III to obtain the hydrazone derivative of general formula I
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according to the invention, whereby the substituents Zi, Z2 and R3
are defined as in the foregoing.
H H
R3 H I R3 I
+ NZ
Z1 Q ~N Z2 Zi 2
H
II III I
A hydrazine of general formula III was reacted under appropriate
conditions with an aldehyde or a ketone of general formula II under
ion exchange resin catalysis and preferably at, but not limited to,
room temperature to yield compound I.
In the compounds I and III of the reaction equation Z2 is preferably
selected from the group consisting of substituted or unsubstituted
phenyl, pyridinyl and pyrimidinyl groups.
The terms as used herein for the chemical entities are defined as
follows:
An alkyl group, if not stated otherwise, denotes a linear or
branched C1-C6-alkyl, preferably a linear or branched chain of C1-C6 -
atoms, a linear or branched C1-C6-alkenyl or a linear or branched C1-
C6-alkinyl group, which can optionally be substituted by one or more
substituents R3, wherein R3 and R4 are defined as above. C1-C6 alkyl
denotes in particular methyl, ethyl, propyl, butyl, pentyl, hexyl or
the corresponding alkene and alkine homologues. Preferably, an alkyl
group denotes a linear or branched C1-C4-alkyl, a linear or branched
C1-C4-alkenyl group, which can be optionally be substituted by one or
more substituents, which are defined as the residues R3 or R4 as men-
tioned above. C1-C4 alkyl denotes in particular methyl, ethyl, pro-
pyl, isopropyl, butyl, isobutyl, t-butyl or the corresponding alkene
homologues.
The Cl-C4-alkyl, C1-C4-alkenyl and C1-C4-alkinyl residue may include
but is not limited to the following groups
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-CH3, -C2H5 , -CH=CH2, -C=CH, -C3H7, -CH ( CH3 ) 2, -CH2-CH=CH2, -C ( CH3 )
=CHZ ,
-CH=CH-CH3, -C=C-CH3 , -CH2-C=CH , -C4H9 , -CH2-CH ( CH3 ) 2, -CH ( CH3 ) -
C2H5 ,
-CH2-CH ( CH3 ) -CH3 , -C ( CH3 ) 3 , -C2H4-CH=CH2 , -CH=CH-C2H5, -CH=C ( CH3
) 2, -CHz -
CH=CH-CH3 , -CH2-C ( CH3 ) =CH2, -C ( CH3 ) =CH-CH3, -C ( CH3 ) -CH=CH2, -
CH=CH-
CH=CH2, -C2H4-C=CH, -C=C-C2H5 , -CH2-C=C-CH3, -C=C-CH=CH2, -CH=CH-C=CH,
-C=C-C=CH.
Preferentially, the Cl-C4-alkyl and C1-C4-alkenyl residue may include
not limited to the following groups
-CH3, -C2H5, -CH=CH2, -C3H7, -CH ( CH3 ) Z , -CH2-CH=CH2, -C4H9 , -CH2-CH (
CH3 ) 2 ,
-CH ( CH3 ) -C2H5 , -CHz-CH ( CH3 ) -CH3 , -C ( CH3 ) 3 =
A cycloalkyl group denotes a non-aromatic ring system containing
three to eight carbon atoms, preferably four to seven carbon atoms,
wherein one or more of the carbon atoms in the ring can be substi-
tuted by a group X, wherein X is selected from the group consisting
of N, S, 0, SO, SOz, NR4 and CO; the C4-C7-cycloalkyl residue may be
selected from the group comprising -cyclo-C3H5, -cyclo-C4H7, -cyclo-
C5H9, -cyclo-C6H11i -cyclo-C7H13, the cycloalkyl group can optionally
be substituted by one or more substituents R3, wherein R3 and R4 are
defined as in the foregoing. Preferentially, a cycloalkyl group de-
notes a non-aromatic ring system containing three to six carbon at-
oms, wherein one or more of the carbon atoms in the ring can be sub-
stituted by a group X, wherein X is selected from the group consist-
ing of N, S, 0, SO, SOz, NR4 and CO; the C4-C6-cycloalkyl residue may
be selected from the group comprising -cyclo-C3H5, -cyclo-C4H7, -
cyclo-C5H9, -cyclo-C6H11, the cycloalkyl group can optionally be sub-
stituted by one or more substituents which are optionally defined as
the residues R3 and/or R4 as stated in the foregoing.
An alkoxy group denotes an 0-alkyl group, the alkyl group being de-
fined as above; the alkoxy group is preferably a methoxy, ethoxy,
isopropoxy, t-butoxy group or a pentoxy group.
A haloalkyl group denotes an alkyl group which is substituted by one
to five halogen atoms, the alkyl group being as defined above; the
haloalkyl group is preferably a -C(R5) 3, -CR5 (R5) 2, -CR5 (R5) R5, -
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or -C2H4-
C(R5) 3, wherein R5 represents F, Cl, Br or I, preferably F or Cl and
wherein R5 may be the same or different. Preferentially, a haloalkyl
group denotes an alkyl group which is substituted by one to three
halogen atoms, the alkyl group being as defined above; the haloalkyl
10 group is preferably a -C(R5) 3, -CH (R5) a, -CH2-CH (R5) 2, -CH2-C (Rs) 3, -
C2H4-CH (R5) Z or -C2H4-C (R5) 3, wherein R5 represents F, Cl, Br or I,
preferably F or Cl and wherein R5 may be the same or different from
one another.
15 A hydroxyalkyl group denotes an HO-alkyl group, the alkyl group be-
ing defined as above.
A haloalkyloxy group denotes an alkoxy group which is substituted by
one to five halogen atoms, the alkyl group being as defined above;
the haloalkyloxy group is pre f erably a -OC( R5 ) 3, -OCR5 (R5)2, -
OCR5 (R5) R5, -OC2 (R5) 5, -OCH2-C (R5) 3, -OCH2-CR5 (R5) 2, -OCH2-CR5 (R5)
R5, -
OC3 (R5) 7 or -OC2H4-C (R5) 3, wherein R5 represents F, Cl, Br or I, pref-
erably F or Cl and wherein R5 may be the same or different. Preferen-
tially, a haloalkoxy group denotes an alkoxy group which is substi-
tuted by one to three halogen atoms, the alkyl group being as de-
fined above; the haloalkyloxy group is preferably a -OC(R5)3, -OCH2-
C(R5) 3, -OC2H4-C (R5) 3, wherein R5 represents F, Cl, Br or I and wherein
R5 may be the same or different from one another.
A hydroxyalkylamino group denotes an (HO-alkyl)2-N- group or
HO-alkyl-NH- group, the alkyl group being as defined above. Prefer-
entially, a hydroxylakylamino group denotes an (OH-alkyl)2-N- group,
HO-alkyl-NH- group or HO-alkyl-NR4- group, the alkyl and R4 groups
being defined as above.
A halogen group denotes chlorine, bromine, fluorine or iodine.
An aryl group denotes preferably an aromatic group having five to
ten carbon and/or heteroatoms, which can optionally be substituted
by one or more substituents wherein R, and R2 being as defined above.
Preferentially, an aryl group denotes an aromatic group having five
to six carbon and/or heteroatoms, which can optionally be substi-
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16
~ 5 tuted by one or more substituents which are optionally being defined
as the residues R1 and/or R2 as mentioned above.
A heteroaryl group denotes a 5- or 6-membered heterocyclic group
which contains at least one heteroatom like 0, N, S. This heterocyc-
lic group can be fused to another ring. For example, this group can
be selected from an oxazol-2-yl, oxazol-4-yl, oxazol-5-yl, thiazol-
2-yl, thiazol-4-yl, thiazol-5-yl, isothiazol-3-yl, isothiazol-4-yl,
isothiazol-5-yl, 1,2,4-oxadiazol-3-yl, 1,2,4-oxadiazol-5-yl, 1,2,4-
thiadiazol-3-yl, 1,2,4-thiadiazol-5-yl, 1,2,5-oxadiazol-3-yl,
1,2,5-thiadiazol-3-yl, 1-imidazolyl, 2-imidazolyl, 4-imidazolyl,
1-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl, 2-furanyl, 3-furanyl, 2-thienyl,
3-thienyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidinyl,
4-pyrimidinyl, 5-pyrimidinyl, 3-pyridazinyl, 4-pyridazinyl,
2-pyrazinyl, 1-pyrazolyl, 3-pyrazolyl, 4-pyrazolyl, indolyl, in-
dolinyl, benzo-[b]-furanyl, benzo[b]thiophenyl, benzimidazolyl, ben-
zothiazolyl, quinazolinyl, quinoxazolinyl, indolizin-yl, isoindolyl,
3H-indolyl, 1H-indazolyl, purinyl, 4H-quinolizinyl, isoxazol-3-yl,
isoxazol-4-yl, isoxazol-5-yl, quinolinyl, tetrahydroquinolinyl, iso-
quinolinyl, tetrahydroisoquinolinyl, phthalazinyl, 1,8-
naphthyridinyl, pteridinyl, carbazolyl, acridinyl, phenazinyl, phe-
nothiazinyl, phenoxazinyl, indenyl, coumarinyl, naphthalinyl, fluo-
renyl, anthracenyl group. This heterocyclic group can optionally be
substituted by one or more substituents R, or R2 which are defined as
above.
Preferentially, a heteroaryl group denotes a 5- or 6-membered het-
erocyclic group which contains at least one heteroatom like 0, N, S
and is selected from the group consisting of a thiazol-2-yl, thia-
zol-4-yl, thiazol-5-yl, 1,2,4-thiadiazol-3-yl, 1,2,4-thiadiazol-5-
yl, 1,2,5-thiadiazol-3-yl, 1-imidazolyl, 2-imidazolyl, 4-imidazolyl,
2-furanyl, 3-furanyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-
pyrimidinyl, 4-pyrimidinyl, 5-pyrimidinyl, 3-pyridazinyl, 4-
pyridazinyl, 2-pyrazinyl, 1-pyrazolyl, 3-pyrazolyl, 4-pyrazolyl, in-
dolyl, benzimidazolyl, benzothiazolyl, quinazolinyl, isoindolyl, 3H-
indolyl, quinolinyl, isoquinolinyl, phtalazinyl, 1,8-naphtyridinyl,
coumarinyl and naphtalinyl group. This heterocyclic group can op-
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17
tionally be substituted by one or more substituents which are de-
fined as the residues R1 and/or R2 as mentioned above.
The invention is further described by way of examples in a merely
illustrative manner without limiting the scop.e of the invention.
Examples
Example 1: General synthetic procedure for the preparation of com-
pounds of formula I
A hydrazine compound according to formula III (-100mg), an alde-
hyde according to general formula II (1 equivalent), ca. 100 mg of
Amberlyst 15 ion-exchange resin and molecular sieves in tetrahydro-
furan (4m1) were placed in a G-12 flask. The reaction mixture was
heated to reflux for 4 h. A TLC analysis (Hexane/Et0Ac:1/1) showed
that all hydrazine was reacted. The reaction mixture was filtered
and the solution concentrated to a small volume (ca. 0.5 mL). The
resulting residue was suspended in methanol (ca. 1.5 mL), sonicated,
centrifuged and decanted. This process was repeated a second time.
The white-yellowish solid formed was dried under vacuum.
This methodology is applicable for all compounds of general formula
I according to the invention. However, other synthetic pathways are
not excluded and are also within the knowledge of a person skilled
in the art.
Specific compounds are shown in the following table 1.
Further specific compounds are shown in the following table 2.
Mass was determined by mass spectrometry. Compounds in Tables 1 and
2 show in-vitro activity below 50 M. In-Vitro activities were de-
termined as described in example 2.
Table 1: List of specific compounds according to general formula I
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18
HPLC/MS
N Structure (ESI)
Br
~ I CI 388
1 I
Br N, N \ [M+H] +
H
/ CI
254
2 Cl o 'N\ ~
N [M+H]+
H
CI
&-H-N426
CI H H N Q [M+H]+
v N"-~OH
/
/ O \ I 395
4 a,_ O N' A ~ [M+H] +
i N
H
a-- O / I
N~N ~ O 395
H [M+H]+
H
6 I \ N'N 381
[M+H]+
N
CI
7 O I\ / I 324
/ / N~N \ [M+H]+
H
H
N/N )acl 307 8 [M+H]+
N
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CI
I \ ~
9 N H n. d.
O
H
\NN
CI
340
[M+H]+
CI
/
11 ~ \ ( \ I 289
N"N \ [M+H]+
H
O
12 N I/ \ N~ o 383
N
0 H [M+H]+
H
NN
306
13 [M+H]+
CI
\ O \ /
I 322
14 CI I/ I/ N.N \ [M+H]+
H
H
N / 235
~
N~N \ [M+H] +
H
NI / I CI 297
16
N, \ [M+H]
H
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17 ( 280
N / \ [M+H] +
H
/ 352
18 JD~! N\ \ I [M+H]+
CI N
H
I CI O 352
19 H
[M+H]+
. I /
CI
OH o 389
20 I \
[M+H]+
CI / ~NN
H
CI
~ OH /
~ ~ 389
/
21 ci ~ H O
[M+H] +
~ \
/
CI
\. ~cI 299
22 NN [M+H]+
CI
H
OH / CI
277
~
23 O N'N [M+H]+
H
CI
CI
/ I 328
24 jt cH
\ [M+H]+
CI , N
H
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21
CI
\ OH / ~ CI 315
25 I
\ [M+H]+
CI ~ N,N
H
CI
26 L I \ CI 282
N~N / / [M+H]+
H
OH
CI
27 265
\ , N\ I / [M+H] +
CI N
H
j:D[ H jo
243
28 N. O N [M+H]+
H
CI
OH \ 283
29
CI \ ~ N'N ~ [M+H] +
H
OH \
246
30 CI \ I N\N /
[M+H]+
H
CI
Ni 'N \
31 N NN I/ CI
H
OJ OH
OH H Br
32 N'N
\ \ \
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22
CI
33 N~ N ~
~1 I
CI NJN~ / CI
H
OH
H
N
3 4 N
+
0:5 la
I-
O
- HO
35 HN -N~
CI
F
~
- Br
36 F H-N\
-O
CI
CI ~ ~
37 Br
0
H-N~
-O
CI
Br
38 H-N~
-O
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23
F
F
39 Br
H_N~
HO
F
F
40 Br
H-%
HO Br
C(IH OZN 41 N
H
N O2
0
OH /
I OH
42 OCN
N ~H
CI
H
43 N-N
-O OH
CI
- N-N
44 OH O-Cl
CI
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24
CI tN-N
45 0=N OH
~-
O b
CI
CI ~N
I N
46 6cl
OH CI CI
H
47 CI N
OH
CI H
~N
48
OH
O -N"O
H
O
49 OYOHN N+0
I I
0
H
N~N I \ _
OH N+.O
CI 0
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O~~N+.O
H
51 QO ~ -
O
N+0
I I
O
O,Z~, N+.O
H
/ ~N \
52 N
I I -
OH N+.O
II
O
OH
H
53 N~N
\ \ I /
O
H
54 NN
O 3r
CI
HO
55 OH O HN I N
\ N N~ CI
H
56 N 1:?, N H
;10
O ,N,, O
CI
N~N ~
57 N~N ~ I NN~ ~ I CI
H
OH
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26
ci
58 N~ I N
N N ci
H OH
Br
ci
59 ci
N N -N H OH
a
i
60 I n. d.
a / /
H
OH
ci
390
61 I I
[M-H]
H
0
H I \
ci 386
62
[M+H]+
I \ I
H
ci
HO 328
63
N\ [M+H]+
N~ CI
H
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F
F
-" 370
64 S /
[M+H]+
N
H \ / I
H
Y 370
65 0
[M+H]+
0
a
/ 326
66
NiN~ \ ( Br [M+H]+
H
OH
a a
\ / I
67 351
a / N-, [M+H]+
H
OH
H 0
CI
I \/ 358
68
/ [M-H] +
4 H
OH CI
H
a
I I I
69 392
\ \ /
[M+H]+
a
CI
70 I \ / ( n.d.
H
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CI
71 338
N~ CI [ M+H ]+
H
OH
72 325
N~ [M+H]+
H
OH
73 269
N ~ \ I [ M+H ] +
H
OH
74 N)-"N 412
\ I W~ N [ M+H ] +
H
OH
Br
/ N I N~ I /
H
Br \ I OH
G
76 \ ~ \N 355
CI I [ M+H ] +
H
OH
CI
77 N-)"N / 400
~~ \ I [M+H]+
H Br
OH
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29
cl
78 408
N--~N [M+H] +
H
a
79 395
I \ ri~N [ M+H ] +
H
CI
80 381
I \ ri \N [M+H]
/ i N
H
CL,a0
81 ~N n. d.
I / /N I
H I
O
440
82 j
N [M+H] +
H
C+1
CI--aO
83 438
N ~ \N [M+H]+
H
C
\ I 0
84 397
~ \ I \N [M+H] +
s N
H
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F
F
85 N~ I I \
H G-I
a Q
86 \ ~N 352
[M+H]
G
H
O-I
a ~
87 Nl)-,N 367
[M+H]+
H
CH
CI
88 N)'~'N 369
N N ~ \ I CI [ M+H ] +
H
OH
89 N)'~'N 356
I N"N\ \ I [M+H]+
H
OH
90 426
~ N I \ [M+H]+
H
OH
C~ a 418
91
ri,N [M+H] +
a ri'N~ ~ I a
H
a-I
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a
92 390
/
N \ I [ M+H ] +
H G
G-I
N Y I a
404
N
93 OH N'NA
[M+H]+
a
94 a 382
'N [M+H]+
a I/ U, l i
H
a-I
~ ~
95 c369
I ~ [M+H]+
/ ~
c
H
OH
C~
ci I 383
96 ~
OI N N [M+H]+
\
H
OH
/\ ~ F F
97 N/\N 420
I eF [M+H]+
AF
H F
Br
98 Ni 'N 423
N [ M+H ] +
H
OH
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32
403
99 ~
r~'~nl i I [M+H]+
H
a-I
a 427
100 I
N~N / [M+H]+
H N \ \ Br
a-I
101 cl 414
~
\ \ [M+H]+
H Br
OH
a 396
102
N~'N [M+H] +
H
aH
103 ~N 398
N a [M+H]+
H
CH
104 CI 426
[M+H]+
N~ CI
H
CH
CI
105 N)~'N 377
,
N~N ~ I [M+H]+
H
0-<
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33
a
106 N"~kN 353
~ N~ N \ \ I [M+H]+
H
383
107
~N [M+H]+
H
CH
~ a
108 388
I [M+H] +
II N a
ci /
~ ~
H
CI-I
Ni 'N 343
109
N [M+H] +
H
& 437
110
\N [M+H] +
H
OH
a I 411
111 I \ NJi\N +
[M+H]
H
OH
112 ~ N' ~N 399
I ~~
\ ~NiJ\% \ [ M+H ] +
F H
F'- F
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a
Ni 'N 397
113 ~ N [M+H]+
H
OH
CI
114 Br / I \ N, N y F n.d.
OH H F F
O, N+-,O
42
1
aN---
115
N\ F [M+H]+
Br N
O H F F
F F
F
\ OH 353
116 I
[M+H]+
/ N~N
H
OH
O
400
117
N [M+H]+
N~N F
H F F
CI
\ N 365
~ I F
118 N
CI / ~ ~N [M+x] +
0 H H F F
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I \ I
119 N 312
N-_ / [M+H]+
H
OH
N)--'N N 335
120 ~,NeK N \ \ ~ [ M+H ] +
H
5
Table 2: List of further specific compounds according to general
formula I
Structure HPLC/MS
N (ESI)
1 Br 388
ci [M+H] +
I \ I \
Br N~N
H
2 Ci 254
[M+H]+
I \
CI j ~N /
H
4 H 395
N
+H] +
[M
I I N )ao
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36
395
~
H N O [M+Hl
N
6 381
H [M+H]+
N O
N
~
N-
7 H 324
\ N [M+H]+
O CI
8 307
\ / \ N LM+H]+
N~
CI
N-
9 H n. d.
~ ~N
T N
O CI
O
N
\ I /
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H 340
N [M+Hl +
I \
CI
CI
11 H 289
N [M+Hl +
12 H 383
N~N [M+Hl +
O
N
13 H 306
~ N~N [M+H]+
CI
14 H 322
[M+Hl +
Ci
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38
15 H 235
N \ ~ [M+H]+
N
H
16 297
N~N Ci [M+Hl
N
H
17 S 280
1 [M+Hl
N-<\
N I \ I \
N
H
18 Cl 352
0 [M+H]-
{ \ \ I
j ~,
H
OH
19 ci 352
[M+H]
I / N\ \
N O
H
OH
20 ci 389
Q [M+H]+
I \
CI / ~ ",
N
H
OH
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39
21 CI 389
[M+H]+
~ ~
CI / N 0
4 H
OH
22 ci 299
CI [M+H]
,
CI N N
H
23 277
[M+H]+
CI
N
H
OH
24 ci 328
cl [M+H]+
( \ I \
CI ,,N
H
OH
25 ci 315
CI [M+H]
\ ( \
CI / ~ N
H
OH
26 ci 282
CI [M+Hl
-
I \ N I
H
OH
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27 ci 265
[M+H]+
1 \ I ~
Cl / ~ N.,N
H
28 243
0 [M+H}+
j ~N /
c ~ \
H
OH
29 ci 283
[M+H]+
~ \
N., /
Cl ~ H
OH
30 cl 246
[M+H]
( \
N\ /
N
H
OH
121
H OH
S
N,,N
N ~ \
I /
~.
~~N
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41
122
S
L N
N
HN
N
OH
123
H
N\ ~ N
O\
N N NH
N
OH O
O
124
-N H
~~-N \
HO N N- OH
O
31
O
H Cf
~~Ny,,,yN"N,
N~ N I /
HO
CI
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42
32
\ \ ~
OH ,IN
HN
Br
33 /
CI
N N 4 iN ( /
CI H ~ CI
OH
34 OH H
\N~N
O
\ \ N
0
CI
HO
N
a N
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43
125
0
+
p=N
~_
+
N ~ Np
N
O
O
126 F
NN' N NH ~
O~ ~ N / N /
~
H
OH
127
H
N
N- H
N J f N'N_ p-
O'N N
-
0
~
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128
o
HO N{-0
1 ~ ~ H N-N ~ O
- ~ N
N N 0-
0 H/\
N~-,N
129
O/ N H
N-
~
N NH
N
~ O
CI \
130 NH2 Chira!
N IN IN
N N
H
HO,,11
O
I::: OH
HO~~,=.
131
H / I
N-N O
S~ \
\ \\N CI
I I
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36 F
F
Br
N-N
0
0
~
37 CI
ci j ~
- Br
H-N
0
38 ci
0 Br
N-N
0
~
39 F
F
Br
H-N15
HO
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46
40 F
F
Br
N-N
H 1--o
HO Br
132
NH
N ~
O\N N NH CI
N O
133 H
N N N, \
~ ~ ~ N
O\N I
N NH O- I
N /
H
134
NH
N N\1
N) N ~
HO
1 Ir 7
O-N
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135
p N
H H N---N~
F N 1N O~
N / ~N
O'
136 F
a NH
N ~ N\
N N
1 ~
O-N
~
137
N N NH O
O / :,, N ~ ~ >
N N N~
H
138 N
O ~ ~ S
~ ~_' ~ H
N-
~
-'O
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139 HO
HN~ ~
N
S i
N
41
,O
ON
HO -N
H
+
O-N
0
140
I
N N NH O
~
'D\ N; , OH
N NH I \
N O CI
141 OH
O O
F ~ ~ H H-N / \ I OH
- // \\
N N
// \\
N", "IN
0
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49
142 \
/
x, OH
N
HN
N
N
143 NH2 Chiral
N~ 'N N N
~\ ~ \ \
N H
HO,,,,= O
HO OH
42 Q
~ I OH
N~N ~
H
OH
144
/ N,N, CI
S 1 H
N\ N
\/ O
CI
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145
O
HN
---' N
S
146
H OH
S
N,,N Br
N-"/ I N
Br
43 CI
N-N
O OH 0
CI
44 OH
N-N
O-Cl
CI
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51
45 c!
H
N~N
OH
+
0 O
147 OH
/ ~
H
- N-N
N _N
-
N
S
46 C!
c! Ci
C! N\
N
OH H
Cl
47 /
OH NNH
CI
48
O OH
I+ H
O%N N,,N
\ l \ I
ci
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52
. 148
O -
O~N+,
N 10
OH NI-INH 0
HO
CI
4g OH
H
N'
N+;O
1_
O
50 OH
CI
H
N'
N+'O
I_
O
51 \ OH
O~
+-O
N N
6
N'
N+,O
1-
O
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53
52 -
O1, N+.O
N+ ,O
OH NNH 0
53 OH
H
\ \ I I
54
H
1N
N
0 Br
149
~ \ \
HN
OH
S N)
55 HO CI
N-N
N_
N CI
OH
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54
56
OH
N
-N / +
O-N
0
57
N
N
H
N~--N \N-
HO CI
CI
58 cI
N
I I \
N N~N CI
H
Br OH
59
?N\ CI
CI
~ N,~
N OH
N H
150 298
O / I
[M+H]+
\
O
H
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151 312
0 a [M+H]+
~~'p j ~N o \ /
152 242
O
[M+H]+
N
H
153 256
O
[M+H]+
0 j N
H
5
Example 2: Inhibition of Beta-Secretase Activity - FRET assay
Assays for determining APP cleavage at the beta-secretase cleavage
site are well known in the art. Exemplary assays are described, for
10 example, in U.S. Patent Nos. 5,744,346 and 5,942,400.
One useful assay to test the inhibitory acitivity of the beta-
secretase is based on the fluorescence resonance energy transfer
(FRET) technology to monitor cleavage of a peptide substrate. 5
15 l/well of substrate (0.2 mM in 0.1 M Na-Ac pH=4.5; Bachem No. M-
2470) is pipetted into a black 96 well plate and stored in the dark.
5 l/well of DMSO containing inhibitor compound (final concentration
of compounds: between 1 nanomolar and 500 micromolar) are pipetted
into 96 well transparent V-form plate, 70~ of 0.1 M Na-Ac pH=4.5
20 and 20 l rhBACE1 (10 g/ml in 0.1 M Na-Ac pH=4.5; R&D Systems No
931-AS) are added and incubated for 10 minutes at RT. 94 ml of each
well are then transferred to the substrate in the black plate to
start the reaction. Immediately after mixing time (T)=0 is measured
with a Wallac1420 plate reader (excitation 355nm, emission 486nm).
25 The reactions are measured again after incubation for 60 min at 37 C
with gentle shaking. Relative compound inhibition potency is deter-
,mined by calculating the concentration of compound that showed a
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56
fifty percent reduction in detected signal (IC50) compared to the en-
zyme reaction signal in the control with no added compound.
Example 3: Inhibition of Beta-Secretase Activity - Cellular Assay
An exemplary assay for the analysis of the inhibition of beta-
secretase activity utilizes human embryonic kidney cell line HEK293
(ATCC Accession No. CRL-1573) transfected with APP751 containing the
naturally occurring double mutation Lys651Met652 to Asn651Met652
(numbered for APP751), commonly called the Swedish mutation and
shown to overproduce A beta (Citron et al., 1992, Nature 360:672-
674), as described in U.S. Patent No. 5,604,102. However, other
transgenic cell lines, for example CHO or SH-SY5Y could be used as
well.
The cells are incubated in the presence/absence of the inhibitory
compound (diluted in DMSO; final concentration of DMSO: 0.05%) at
the desired concentration, generally up to 100 M. At the end of the
treatment period (24h), conditioned media is analyzed for beta-
secretase activity, for example, by analysis of cleavage fragments.
A beta can be analyzed by immunoassay, using specific detection an-
tibodies (HS40 ABeta kit from The Genetics Company). The enzymatic
activity is measured in the presence and abs ence of the compound in-
hibitors to demonstrate specific inhibition of beta-secretase medi-
ated cleavage of APP substrate. Relative compound inhibition potency
is determined by calculating the concentration of compound that
showed a fifty percent reduction in detected signal (IC50) compared
to the A beta 40 signal in the control with no added compound.
The following compounds showed IC50 values below 50 M:
Nl, N8, N10, N13, N15-N21, N23-N30, N40, N43, N58-N66, N69-N70, N76-
N81, N83-N88, N90-N94, N96, N98-N100, N102, N108-N109, N114, N119.
Example 4: Determination of compound in the brain
C57/b16 mice, designated "wt" mice, were obtained. The mice were
treated with the phenylhydrazone Nr. 29. Dosing was administered ei-
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ther intraperitoneally or orally. The oral dose was administered via
oral gavage. Intraperitoneally-treated animals received a single in-
jection, or were dosed once or twice a day up to 8 days. In addi-
tion, a group of age-matching wt mice were treated with the carrier
vehicle only.
'Animals were sacrified between 0.5 and 24 h after the final dose.
The brains were removed and stored at -80 C. The brain was cut into
200 mg pieces, 800 l of a Acetonitrile:Water solution (80:20; v:v)
were added and the brain samples were sonicated for 2 min. Samples
were vortexed for 2 min and insoluble particles were removed by cen-
trifugation. The compound in the samples was detected by High Per-
formance Liquid Chromatography at 254 or 360 nm. Concentrations were
determined by adding an internal standard to the brain samples.
Following intraperitoneal administration of a single dose of 55
mg/kg, compound levels were detectable in the brain within 30 min,
persist at this level for at least 5 h post administration and dis-
play peak levels 2.5 h after injection. Upon treatment of mice once
a day with the same dose for three consecutive days, the amount of
the compound detected in the brain 24 h after the last injection was
twice the peak level measured after single dosing. Similar levels of
compound could be detected after treatment twice a day for 8 con-
secutive days, indicating a saturation effect for the accumulation
of the compound in the brain.
After single oral dose administration (100 mg/kg) the compound could
be detected in the brain of "wt" mice arguing for oral absorption of
the compound.
Example 5: Inhibition of Beta-Secretase in Animal Models of AD
,Various animal models can be used to screen for inhibition of beta-
secretase activity. Examples of animal models useful in the inven-
'tion include, but are not limited to, mouse, guinea pig, dog, and
the like. The animals used can be wild type, transgenic, or knockout
models. In addition mammalian models can express mutation in APP,
õsuch as APP695-SW and the like described herein. Examples of trans-
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- 5 genic non-human mammalian models are described in Hsiao et al.,
1996, Science 274, 99-102 and Kawarabayshi et al., 2001, J Neuroscie
21, 372-381.
For instance, Tg2576 mice, prepared as described in Hsiao et al.,
1996, Science 274, 99-102 are useful to analyze in vivo suppression
of A beta release in the presence of putative inhibitory compounds.
Transgenic animals are administered an amount of the BACE inhibitor
formulated in a carrier suitable for the chosen mode of administra-
tion, as for example described in Chang et al., 2004, J. Neurochem.
89, 1409-1416. Control animals are untreated, treated with vehicle,
or treated with an inactive compound. Administration can be acute,
i.e., single dose or multiple dose in one day, or can be chronic,
i.e., dosing is repeated daily for a period of days. Beginning at
time 0, brain tissue, cerebral fluid or blood is obtained from se-
lected animals and analyzed for the presence of APP cleavage pep-
tides, including A beta, for example, by immunoassay using specific
antibodies for A beta detection. At the end of the test period, ani-
mals are sacrificed and brain tissue, cerebral fluid or blood is
analyzed for the presence of A beta and/or beta-amyloid plaques. Al-
ternatively, treated transgenic animals (e.g. Tg2576 mice) can be
monitored for changes in the APP induced phenotypes as for example
described in Hsiao et al., 1996, Science 274, 99-102.
