Note: Descriptions are shown in the official language in which they were submitted.
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USE OF HDAC INHIBITORS FOR THE TREATMENT OF GASTROINTESTINAL CANCERS
Field of the Invention
The present invention relates to the use of an HDAC inhibitor for the
preparation of a
medicament for the treatment of gastrointestinal cancers; a method of treating
a warm-
blooded animal, especially a human, having gastrointestinal cancer, comprising
administering to said animal a therapeutically effective amount of an HDAC
inhibitor,
especially a compound of formula (I), as defined herein; and to a
pharmaceutical
composition and a commercial package.
Background of the Invention
Patients suffering from gastrointestinal cancers have low overall survival
rates. The
standard treatment of chemotherapy is not always effective. Therefore, there
is a need to
develop novel treatment methods.
Summary of the Invention
The term "gastrointestinal cancers", as used herein, includes, but is not
limited to,
hepatocellular carcinoma and/or pancreatic cancer.
The compounds of formula (I), as defined herein, are histone deacetylase
inhibitors
(HDAC inhibitors). Reversible acetylation of histones is a major regulator of
gene expression
that acts by altering accessibility of transcription factors to DNA. In normal
cells, histone
deacetylase (HDA) and histone acetyltrasferase together control the level of
acetylation of
histones to maintain a balance. Inhibition of HDA results in the accumulation
of
hyperacetylated histones, which results in a variety of cellular responses.
Surprisingly, it was now found that HDAC inhibitors, especially the compounds
of
formula (I), as defined herein, directly inhibit the proliferation of
gastrointestinal cancer, such
hepatocellular carcinoma and/or pancreatic cancer.
Hence, the invention relates to the use of an HDAC inhibitor for the
preparation of a
medicament for the treatment of gastrointestinal cancer.
-1-
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Detailed Description of the Drawings
Figure 1 illustrates that LBH589 treatment inhibits tumor growth in the HCT1
16
xenograft model.
Figure 2 illustrates co-treatment of LBH589 with 5-Fluorouracil enhances tumor
growth inhibition and tumor growth delay in Colo205 colon cancer xenograft
model.
Figure 3 illustrates the anti-proliferative and cytotoxic effects of LBH589 in
19
pancreatic cancer cell lines.
Detailed Description of the Invention
HDAC Inhibitor Compounds
HDAC inhibitor compounds of particular interest for use in the inventive
combination
are hydroxamate compounds described by the formula (I):
O Ri
Y
HONI i 2 R3 R4
H (~)
I N R5
X "s "s
wherein
R, is H; halo; or a straight-chain C,-C6alkyl, especially methyl, ethyl or n-
propyl, which
methyl, ethyl and n-propyl substituents are unsubstituted or substituted by
one or
more substituents described below for alkyl substituents;
R2 is selected from H; C,-C,oalkyl, preferably C,-Csalkyl, e.g., methyl, ethyl
or -CH2CH2-
OH; C4-C9cycloalkyl; C4-C9heterocycloalkyl; C4-C9heterocycloalkylalkyl;
cycloalkylalkyl,
e.g., cyclopropylmethyl; aryl; heteroaryl; arylalkyl, e.g., benzyl;
heteroarylalkyl, e.g.,
pyridylmethyl; -(CH2)nC(O)R6; -(CH2)nOC(O)R6; amino acyl; HON-C(O)-CH=C(R,)-
aryl-
alkyl-; and -(CH2)nR7;
R3 and R4 are the same or different and independently H, C,-C6alkyl, acyl or
acylamino, or
R3 and R4, together with the carbon to which they are bound, represent C=O,
C=S or
C=NR8, or
-2-
--
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R2, together with the nitrogen to which it is bound, and R3, together with the
carbon to
which it is bound, can form a C4-C9heterocycloalkyl, a heteroaryl, a
polyheteroaryl, a
non-aromatic polyheterocycle, or a mixed aryl and non-aryl polyheterocycle
ring;
R5 is selected from H; Cl-Csalkyl; C4-C9cycloalkyl; C4-C9heterocycloalkyl;
acyl; aryl;
heteroaryl; arylalkyl, e.g., benzyl; heteroarylalkyl, e.g., pyridylmethyl;
aromatic
polycycles; non-aromatic polycycles; mixed aryl and non-aryl polycycles;
polyheteroaryl; non-aromatic polyheterocycles; and mixed aryl and non-aryl
polyheterocycles;
n, ni, n2 and n3 are the same or different and independently selected from 0-
6, when n, is
1-6, each carbon atom can be optionally and independently substituted with R3
and/or
Ra;
X and Y are the same or different and independently selected from H; halo; Cl-
C4alkyl,
such as CH3 and CF3; NO2; C(O)Rj; OR9; SR9; CN; and NRjoR>>;
R6 is selected from H; Cl-C6alkyl; C4-C9cycloalkyl; C4-C9heterocycloalkyl;
cycloalkylalkyl,
e.g., cyclopropylmethyl; aryl; heteroaryl; arylalkyl, e.g., benzyl and 2-
phenylethenyl;
heteroarylalkyl, e.g., pyridylmethyl; OR12; and NR13R14;
R7 is selected from OR15, SR15, S(O)R16, S02R17, NR13R14 and NR12S02R6;
R8 is selected from H; OR15; NR13R14; Cl-Csalkyl; C4-C9cycloalkyl; C4-
C9heterocycloalkyl;
aryl; heteroaryl; arylalkyl, e.g., benzyl; and heteroarylalkyl, e.g.,
pyridylmethyl;
R9 is selected from Cl-C4alkyl, e.g., CH3 and CF3; C(O)-alkyl, e.g., C(O)CH3;
and
C(O)CF3;
Rlo and R>> are the same or different and independently selected from H, Cl-
C4alkyl and
-C(O)-alkyl;
R12 is selected from H; Cl-Csalkyl; C4-C9cycloalkyl; C4-C9heterocycloalkyl;
C4-C9heterocycloalkylalkyl; aryl; mixed aryl and non-aryl polycycle;
heteroaryl;
arylalkyl, e.g., benzyl; and heteroarylalkyl, e.g., pyridylmethyl;
R13 and R14 are the same or different and independently selected from H; Cl-
Csalkyl;
C4-C9cycloalkyl; C4-C9heterocycloalkyf; aryl; heteroaryl; arylalkyl, e.g.,
benzyl;
heteroarylalkyl, e.g., pyridylmethyl; amino acyl, or
R13 and R14, together with the nitrogen to which they are bound, are C4-
C9heterocycloalkyl, heteroaryl, polyheteroaryl, non-aromatic polyheterocycle
or mixed
aryl and non-aryl polyheterocycle;
-3-
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R15 is selected from H, C,-Csalkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl,
aryl, heteroaryl,
arylalkyl, heteroarylalkyl and (CH2),ZR12;
R16 is selected from C,-C6alkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl, aryl,
heteroaryl,
polyheteroaryl, arylalkyl, heteroarylalkyl and (CH2),ZR12;
R17 is selected from C,-Csalkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl, aryl,
aromatic
polycycles, heteroaryl, arylalkyl, heteroarylalkyl, polyheteroaryl and
NR13R14;
m is an integer selected from 0-6; and
Z is selected from 0; NR13; S; and S(O),
or a pharmaceutically acceptable salt thereof.
As appropriate, "unsubstituted" means that there is no substituent or that the
only
substituents are hydrogen.
Halo substituents are selected from fluoro, chloro, bromo and iodo, preferably
fluoro
or chloro.
Alkyl substituents include straight- and branched-C,-C6alkyl, unless otherwise
noted.
Examples of suitable straight- and branched-C,-Csalkyl substituents include
methyl, ethyl,
n-propyl, 2-propyl, n-butyl, sec-butyl, t-butyl and the like. Unless otherwise
noted, the alkyl
substituents include both unsubstituted alkyl groups and alkyl groups that are
substituted by
one or more suitable substituents, including unsaturation, i.e., there are one
or more double
or triple C-C bonds; acyl; cycloalkyl; halo; oxyalkyl; alkylamino; aminoalkyl;
acylamino; and
OR15, e.g., alkoxy. Preferred substituents for alkyl groups include halo,
hydroxy, alkoxy,
oxyalkyl, alkylamino and aminoalkyl.
Cycloalkyl substituents include C3-C9cycloalkyl groups, such as cyclopropyl,
cyclobutyl, cyclopentyl, cyclohexyl and the like, unless otherwise specified.
Unless otherwise
noted, cycloalkyl substituents include both unsubstituted cycloalkyl groups
and cycloalkyl
groups that are substituted by one or more suitable substituents, including C,-
Csalkyl, halo,
hydroxy, aminoalkyl, oxyalkyl, alkylamino and OR15, such as alkoxy. Preferred
substituents
for cycloalkyl groups include halo, hydroxy, alkoxy, oxyalkyl, alkylamino and
aminoalkyl.
The above discussion of alkyl and cycloalkyl substituents also applies to the
alkyl
portions of other substituents, such as, without limitation, alkoxy, alkyl
amines, alkyl ketones,
arylalkyl, heteroarylalkyl, alkylsulfonyl and alkyl ester substituents and the
like.
-4-
----- - - - -- -
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Heterocycloalkyl substituents include 3- to 9-membered aliphatic rings, such
as 4- to
7-membered aliphatic rings, containing from 1-3 heteroatoms selected from
nitrogen, sulfur,
oxygen. Examples of suitable heterocycloalkyl substituents include pyrrolidyl,
tetrahydrofuryl, tetrahydrothiofuranyl, piperidyl, piperazyl,
tetrahydropyranyl, morphilino,
1,3-diazapane, 1,4-diazapane, 1,4-oxazepane and 1,4-oxathiapane. Unless
otherwise
noted, the rings are unsubstituted or substituted on the carbon atoms by one
or more
suitable substituents, including C,-C6alkyl; C4-C9cycloalkyl; aryl;
heteroaryl; arylalkyl, e.g.,
benzyl; heteroarylalkyl, e.g., pyridylmethyl; halo; amino; alkyl amino and
OR15, e.g., alkoxy.
Unless otherwise noted, nitrogen heteroatoms are unsubstituted or substituted
by H,
C,-C4alkyl; arylalkyl, e.g., benzyl; heteroarylalkyl, e.g., pyridylmethyl;
acyl; aminoacyl;
alkylsulfonyl; and arylsulfonyl.
Cycloalkylalkyl substituents include compounds of the formula -(CH2)n5-
cycloalkyl,
wherein n5 is a number from 1-6. Suitable alkylcycloalkyl substituents include
cyclopentylmethyl, cyclopentylethyl, cyclohexylmethyl and the like. Such
substituents are
unsubstituted or substituted in the alkyl portion or in the cycloalkyl portion
by a suitable
substituent, including those listed above for alkyl and cycloalkyl.
Aryl substituents include unsubstituted phenyl and phenyl substituted by one
or more
suitable substituents including C,-C6alkyl; cycloalkylalkyl, e.g.,
cyclopropylmethyl;
O(CO)alkyl; oxyalkyl; halo; nitro; amino; alkylamino; aminoalkyl; alkyl
ketones; nitrile;
carboxyalkyl; alkylsulfonyl; aminosulfonyl; arylsulfonyl and OR15, such as
alkoxy. Preferred
substituents include including C,-C6alkyl; cycloalkyl, e.g.,
cyclopropylmethyl; alkoxy; oxyalkyl;
halo; nitro; amino; alkylamino; aminoalkyl; alkyl ketones; nitrile;
carboxyalkyl; alkylsulfonyl;
arylsulfonyl and aminosulfonyl. Examples of suitable aryl groups include C,-
C4alkylphenyl,
C,-C4alkoxyphenyl, trifluoromethylphenyl, methoxyphenyl, hydroxyethylphenyl,
dimethylaminophenyl, aminopropylphenyl, carbethoxyphenyl,
methanesulfonylphenyl and
tolylsulfonylphenyl.
Aromatic polycycles include naphthyl, and naphthyl substituted by one or more
suitable substituents including C,-C6alkyl; alkylcycloalkyl, e.g.,
cyclopropylmethyl; oxyalkyl;
halo; nitro; amino; alkylamino; aminoalkyl; alkyl ketones; nitrile;
carboxyalkyl; alkylsulfonyl;
arylsulfonyl; aminosulfonyl and OR15, such as alkoxy.
Heteroaryl substituents include compounds with a 5- to 7-membered aromatic
ring
containing one or more heteroatoms, e.g., from 1-4 heteroatoms, selected from
N, 0 and S.
-5-
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Typical heteroaryl substituents include furyl, thienyl, pyrrole, pyrazole,
triazole, thiazole,
oxazole, pyridine, pyrimidine, isoxazolyl, pyrazine and the like. Unless
otherwise noted,
heteroaryl substituents are unsubstituted or substituted on a carbon atom by
one or more
suitable substituents, including alkyl, the alkyl substituents identified
above, and another
heteroaryl substituent. Nitrogen atoms are unsubstituted or substituted, e.g.,
by R13;
especially useful N substituents include H, C,-C4alkyl, acyl, aminoacyl and
sulfonyl.
Arylalkyl substituents include groups of the formula -(CH2)n5-aryl, -(CH2)15-,-
(CH-aryl)-
(CH2),5-aryl or -(CH2)15_,CH(aryl)(aryl), wherein aryl and n5 are defined
above. Such
arylalkyl substituents include benzyl, 2-phenylethyl, 1-phenylethyl, tolyl-3-
propyl, 2-
phenylpropyl, diphenylmethyl, 2-diphenylethyl, 5,5-dimethyl-3-phenylpentyl and
the like.
Arylalkyl substituents are unsubstituted or substituted in the alkyl moiety or
the aryl moiety or
both as described above for alkyl and aryl substituents.
Heteroarylalkyl substituents include groups of the formula -(CH2)n5-
heteroaryl,
wherein heteroaryl and n5 are defined above and the bridging group is linked
to a carbon or
a nitrogen of the heteroaryl portion, such as 2-, 3- or 4-pyridylmethyl,
imidazolylmethyl,
quinolylethyl and pyrrolylbutyl. Heteroaryl substituents are unsubstituted or
substituted as
discussed above for heteroaryl and alkyl substituents.
Amino acyl substituents include groups of the formula -C(O)-(CH2)I-
C(H)(NR13R14)-
(CH2),-R5, wherein n, R13, R14 and R5 are described above. Suitable aminoacyl
substituents
include natural and non-natural amino acids, such as glycinyl, D-tryptophanyl,
L-lysinyl, D- or
L-homoserinyl, 4-aminobutryic acyl and -3-amin-4-hexenoyl.
Non-aromatic polycycle substituents include bicyclic and tricyclic fused ring
systems
where each ring can be 4- to 9-membered and each ring can contain zero, one or
more
double and/or triple bonds. Suitable examples of non-aromatic polycycles
include decalin,
octahydroindene, perhydrobenzocycloheptene and perhydrobenzo-[fi]-azulene.
Such
substituents are unsubstituted or substituted as described above for
cycloalkyl groups.
Mixed aryl and non-aryl polycycle substituents include bicyclic and tricyclic
fused ring
systems where each ring can be 4- to 9-membered and at least one ring is
aromatic.
Suitable examples of mixed aryl and non-aryl polycycles include
methylenedioxyphenyl,
bis-methylenedioxyphenyl, 1,2,3,4-tetrahydronaphthalene, dibenzosuberane,
dihdydroanthracene and 9H-fluorene. Such substituents are unsubstituted or
substituted by
nitro or as described above for cycloalkyl groups.
-6-
-- - -- -- - - ---- - -- - -
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Polyheteroaryl substituents include bicyclic and tricyclic fused ring systems
where
each ring can independently be 5- or 6-membered and contain one or more
heteroatom,
e.g., 1, 2, 3 or 4 heteroatoms, chosen from 0, N or S such that the fused ring
system is
aromatic. Suitable examples of polyheteroaryl ring systems include quinoline,
isoquinoline,
pyridopyrazine, pyrrolopyridine, furopyridine, indole, benzofuran,
benzothiofuran, benzindole,
benzoxazole, pyrroloquinoline and the like. Unless otherwise noted,
polyheteroaryl
substituents are unsubstituted or substituted on a carbon atom by one or more
suitable
substituents, including alkyl, the alkyl substituents identified above and a
substituent of the
formula -O-(CH2CH=CH(CH3)(CH2))1_3H. Nitrogen atoms are unsubstituted or
substituted,
e.g., by R13, especially useful N substituents include H, C,-C4alkyl, acyl,
aminoacyl and
sulfonyl.
Non-aromatic polyheterocyclic substituents include bicyclic and tricyclic
fused ring
systems where each ring can be 4- to 9-membered, contain one or more
heteroatom, e.g., 1,
2, 3 or 4 heteroatoms, chosen from 0, N or S and contain zero or one or more C-
C double
or triple bonds. Suitable examples of non-aromatic polyheterocycles include
hexitol,
cis-perhydro-cyclohepta[b]pyridinyl, decahydro-benzo[tj[1,4]oxazepinyl,
2,8-dioxabicyclo[3.3.0]octane, hexahydro-thieno[3,2-b]thiophene,
perhydropyrrolo[3,2-
b]pyrrole, perhydronaphthyridine, perhydro-lH-dicyclopenta[b,e]pyran. Unless
otherwise
noted, non-aromatic polyheterocyclic substituents are unsubstituted or
substituted on a
carbon atom by one or more substituents, including alkyl and the alkyl
substituents identified
above. Nitrogen atoms are unsubstituted or substituted, e.g., by R13,
especially useful N
substituents include H, C,-C4alkyl, acyl, aminoacyl and sulfonyl.
Mixed aryl and non-aryl polyheterocycles substituents include bicyclic and
tricyclic
fused ring systems where each ring can be 4- to 9-membered, contain one or
more
heteroatom chosen from 0, N or S, and at least one of the rings must be
aromatic. Suitable
examples of mixed aryl and non-aryl polyheterocycles include 2,3-
dihydroindole,
1,2,3,4-tetrahydroquinoline, 5,11-dihydro-1OH-dibenz[b,e][1,4]diazepine,
5H-dibenzo[b,e][1,4]diazepine, 1,2-dihydropyrrolo[3,4-b][1,5]benzodiazepine,
1,5-dihydro-
pyrido[2,3-b][1,4]diazepin-4-one, 1,2,3,4,6,11-hexahydro-benzo[b]pyrido[2,3-
e][1,4]diazepin-
5-one. Unless otherwise noted, mixed aryl and non-aryl polyheterocyclic
substituents are
unsubstituted or substituted on a carbon atom by one or more suitable
substituents including
-N-OH, =N-OH, alkyl and the alkyl substituents identified above. Nitrogen
atoms are
unsubstituted or substituted, e.g., by R13; especially useful N substituents
include H,
C,-C4alkyl, acyl, aminoacyl and sulfonyl.
-7-
-- -- - - - - -- -- - -- - -- - - - --- -- - -- ------ -
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Amino substituents include primary, secondary and tertiary amines and in salt
form,
quaternary amines. Examples of amino substituents include mono- and di-
alkylamino,
mono- and di-aryl amino, mono- and di-arylalkyl amino, aryl-arylalkylamino,
alkyl-arylamino,
alkyl-arylalkylamino and the like.
Sulfonyl substituents include alkylsulfonyl and arylsulfonyl, e.g., methane
sulfonyl,
benzene sulfonyl, tosyl and the like.
Acyl substituents include groups of formula -C(O)-W, -OC(O)-W, -C(O)-O-W or
-C(O)NR13R14, where W is R16, H or cycloalkylalkyl.
Acylamino substituents include substituents of the formula -N(R12)C(O)-W,
-N(R12)C(O)-O-W and -N(R,2)C(O)-NHOH and R12 and W are defined above.
The R2 substituent HON-C(O)-CH=C(R,)-aryl-alkyl- is a group of the formula:
O
HO~ X
H I
Y n4
Preferences for each of the substituents include the following:
R, is H, halo or a straight-chain C,-C4alkyl;
R2 is selected from H, C,-Csalkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl,
cycloalkylalkyl,
aryl, heteroaryl, arylalkyl, heteroarylalkyl, -(CH2)nC(O)R6, amino acyl and -
(CH2)nR7;
R3 and R4 are the same or different and independently selected from H and C,-
Csalkyl, or
R3 and R4, together with the carbon to which they are bound, represent C=O,
C=S or
C=NR8;
R5 is selected from H, C,-Csalkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl,
aryl, heteroaryl,
arylalkyl, heteroarylalkyl, a aromatic polycycle, a non-aromatic polycycle, a
mixed
aryl and non-aryl polycycle, polyheteroaryl, a non-aromatic polyheterocycle,
and a
mixed aryl and non-aryl polyheterocycle;
n, n,, n2 and n3 are the same or different and independently selected from 0-
6, when n, is
1-6, each carbon atom is unsubstituted or independently substituted with R3
and/or
R4;
-8-
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X and Y are the same or different and independently selected from H, halo, C,-
C4alkyl,
CF3, NO2, C(O)R,, OR9, SR9, CN and NR,oR,,;
R6 is selected from H, C,-C6alkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl,
alkylcycloalkyl,
aryl, heteroaryl, arylalkyl, heteroarylalkyl, OR12 and NR13R14;
R7 is selected from OR15, SR15, S(O)R,6, S02R17, NR13R14 and NR,2SO2R6;
R8 is selected from H, OR15, NR13R14, C,-C6alkyl, C4-C9cycloalkyl, C4-
C9heterocycloalkyl,
aryl, heteroaryl, arylalkyl and heteroarylalkyl;
R9 is selected from C,-C4alkyl and C(O)-alkyl;
R,o and Rõ are the same or different and independently selected from H, C,-
C4alkyl and
-C(O)-alkyl;
R12 is selected from H, C,-C6alkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl,
aryl,
heteroaryl, arylalkyl and heteroarylalkyl;
R13 and R14 are the same or different and independently selected from H, C,-
C6alkyl,
C4-C9cycloalkyl, C4-C9heterocycloalkyl, aryl, heteroaryl, arylalkyl,
heteroarylalkyl and
amino acyl;
R15 is selected from H, C,-C6alkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl,
aryl,
heteroaryl, arylalkyl, heteroarylalkyl and (CH2)mZR12;
R16 is selected from C,-C6alkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl, aryl,
heteroaryl,
arylalkyl, heteroarylalkyl and (CH2)mZR12;
R17 is selected from C,-C6alkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl, aryl,
heteroaryl,
arylalkyl, heteroarylalkyl and NR13R14;
m is an integer selected from 0-6; and
Z is selected from 0, NR13, S and S(O);
or a pharmaceutically acceptable salt thereof.
Useful compounds of the formula (I), include those wherein each of R,, X, Y,
R3 and
R4 is H, including those wherein one of n2 and n3 is 0 and the other is 1,
especially those
wherein R2 is H or -CH2-CH2-OH.
One suitable genus of hydroxamate compounds are those of formula (la):
-9-
-----
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0
HO~H (2 ~~a)
nn 4 N R ,
wherein
n4 is 0-3;
R2 is selected from H, C,-Csalkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl,
cycloalkylalkyl,
aryl, heteroaryl, arylalkyl, heteroarylalkyl, -(CH2)nC(O)R6, amino acyl and -
(CH2)nR7;
and
R5 is heteroaryl; heteroarylalkyl, e.g., pyridylmethyl; aromatic polycycles;
non-aromatic
polycycles; mixed aryl and non-aryl polycycles; polyheteroaryl or mixed aryl;
and
non-aryl polyheterocycles;
or a pharmaceutically acceptable salt thereof.
Another suitable genus of hydroxamate compounds are those of formula (la):
0
HO~ i
H 2 ~~a)
\ ' N
n4 R ~
5
wherein
n4 is 0-3;
R2 is selected from H, C,-Csalkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl,
cycloalkylalkyl,
aryl, heteroaryl, arylalkyl, heteroarylalkyl, -(CH2)nC(O)R6, amino acyl and -
(CH2)nR7;
R5 is aryl; arylalkyl; aromatic polycycles; non-aromatic polycycles and mixed
aryl; and
non-aryl polycycles, especially aryl, such as p-fluorophenyl, p-chlorophenyl,
p-O-C,-
C4alkylphenyl, such as p-methoxyphenyl, and p-C,-C4alkylphenyl; and arylalkyl,
such
as benzyl, ortho-, meta- orpara-fluorobenzyl, ortho-, meta- orpara-
chlorobenzyl,
ortho-, meta- or para-mono, di- or tri-O-C,-C4alkylbenzyl, such as ortho-,
meta- or
para-methoxybenzyl, m,p-diethoxybenzyl, o,m,p-triimethoxybenzyl and ortho-,
meta-
orpara-mono, di- or tri-C,-C4alkylphenyl, such as p-methyl, m,m-diethylphenyl;
or a pharmaceutically acceptable salt thereof.
Another interesting genus is the compounds of formula (Ib):
-10-
-
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O
HONI i 2
H (Ib)
N
Rõ
wherein
R2 is selected from H; C,-Csalkyl; C4-C6cycloalkyl; cycloalkylalkyl, e.g.,
cyclopropylmethyl; (CH2)2-4OR21, where R21 is H, methyl, ethyl, propyl and i-
propyl;
and
R5 is unsubstituted 1 H-indol-3-yl, benzofuran-3-yl or quinolin-3-yl, or
substituted 1 H-indol-
3-yl, such as 5-fluoro-1 H-indol-3-yl or 5-methoxy-1 H-indol-3-yl, benzofuran-
3-yl or
quinolin-3-yl;
or a pharmaceutically acceptable salt thereof.
Another interesting genus of hydroxamate compounds are the compounds of
formula (Ic):
O Ri
HOI-I X R1a
v
H R2 R3 R4 Z, (1c)
Y p q r
Al
wherein
the ring containing Z, is aromatic or non-aromatic, which non-aromatic rings
are
saturated or unsaturated,
Z, is 0, S or N-R20;
R18 is H; halo; C,-C6alkyl (methyl, ethyl, t-butyl); C3-C,cycloalkyl; aryl,
e.g., unsubstituted
phenyl or phenyl substituted by 4-OCH3 or 4-CF3; or heteroaryl, such as 2-
furanyl,
2-thiophenyl or 2-, 3- or 4-pyridyl;
R20 is H; C,-Csalkyl; C,-C6alkyl-C3-C9cycloalkyl, e.g., cyclopropylmethyl;
aryl; heteroaryl;
arylalkyl, e.g., benzyl; heteroarylalkyl, e.g., pyridylmethyl; acyl, e.g.,
acetyl, propionyl
and benzoyl; or sulfonyl, e.g., methanesulfonyl, ethanesulfonyl,
benzenesulfonyl and
toluenesulfonyl;
A, is 1, 2 or 3 substituents which are independently H; C,-Csalkyl; -OR19;
halo;
alkylamino; aminoalkyl; halo; or heteroarylalkyl, e.g., pyridylmethyl;
-11-
-
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R19 is selected from H; C,-Csalkyl; C4-C9cycloalkyl; C4-C9heterocycloalkyl;
aryl;
heteroaryl; arylalkyl, e.g., benzyl; heteroarylalkyl, e.g., pyridylmethyl and
-(CH2CH=CH(CH3)(CH2))1_3H;
R2 is selected from H, C,-Csalkyl, C4-C9cycloalkyl, C4-C9heterocycloalkyl,
cycloalkylalkyl,
aryl, heteroaryl, arylalkyl, heteroarylalkyl, -(CH2)~C(O)R6, amino acyl and -
(CH2),,R7;
v is 0, 1 or 2;
p is 0-3; and
q is 1-5 and r is 0; or
q is 0 and r is 1-5;
or a pharmaceutically acceptable salt thereof. The other variable substituents
are as defined
above.
Especially useful compounds of formula (Ic), are those wherein R2 is H, or
-(CH2)PCH2OH, wherein p is 1-3, especially those wherein R, is H; such as
those wherein R,
is H and X and Y are each H, and wherein q is 1-3 and r is 0 or wherein q is 0
and r is 1-3,
especially those wherein Z, is N-R20. Among these compounds R2 is preferably H
or -CH2-
CH2-OH and the sum of q and r is preferably 1.
Another interesting genus of hydroxamate compounds are the compounds of
formula (Id):
O Ri
HOI-I X R~$
H R2 R3 R4 Z, (Id)
Y p q r
Al
wherein
Z, is 0, S or N-R20;
R18 is H; halo; C,-C6alkyl (methyl, ethyl, t-butyl); C3-C7cycloalkyl; aryl,
e.g., unsubstituted
phenyl or phenyl substituted by 4-OCH3 or 4-CF3; or heteroaryl;
R20 is H; C,-Csalkyl, C,-C6alkyl-C3-C9cycloalkyl, e.g., cyclopropylmethyl;
aryl; heteroaryl;
arylalkyl, e.g., benzyl; heteroarylalkyl, e.g., pyridylmethyl; acyl, e.g.,
acetyl, propionyl
and benzoyl; or sulfonyl, e.g., methanesulfonyl, ethanesulfonyl,
benzenesulfonyl,
toluenesulfonyl);
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A, is 1, 2 or 3 substituents which are independently H, C,-C6alkyl, -OR19 or
halo;
R19 is selected from H; C,-C6alkyl; C4-C9cycloalkyl; C4-C9heterocycloalkyl;
aryl;
heteroaryl; arylalkyl, e.g., benzyl; and heteroarylalkyl, e.g., pyridylmethyl;
p is 0-3; and
q is 1-5 and r is 0; or
q is 0 and r is 1-5;
or a pharmaceutically acceptable salt thereof. The other variable substituents
are as defined
above.
Especially useful compounds of formula (Id), are those wherein R2 is H or
-(CH2)PCH2OH, wherein p is 1-3, especially those wherein R, is H; such as
those wherein R,
is H and X and Y are each H, and wherein q is 1-3 and r is 0 or wherein q is 0
and r is 1-3.
Among these compounds R2 is preferably H or -CH2-CH2-OH and the sum of q and r
is
preferably 1.
The present invention further relates to compounds of the formula (le):
O Ri
HO", X R18
H R2 R3 R4 " ' N-R20 (le)
Y P 4 r
Al
or a pharmaceutically acceptable salt thereof. The variable substituents are
as defined
above.
Especially useful compounds of formula (le), are those wherein R18 is H,
fluoro,
chloro, bromo, a C,-C4alkyl group, a substituted C,-C4alkyl group, a C3-
C7cycloalkyl group,
unsubstituted phenyl, phenyl substituted in the para position, or a
heteroaryl, e.g., pyridyl,
ring.
Another group of useful compounds of formula (le), are those wherein R2 is H
or
-(CH2)PCH2OH, wherein p is 1-3, especially those wherein R, is H; such as
those wherein R,
is H and X and Y are each H, and wherein q is 1-3 and r is 0 or wherein q is 0
and r is 1-3.
Among these compounds R2 is preferably H or -CH2-CH2-OH and the sum of q and r
is
preferably 1. Among these compounds p is preferably 1 and R3 and R4 are
preferably H.
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Another group of useful compounds of formula (le), are those wherein R18 is H,
methyl, ethyl, t-butyl, trifluoromethyl, cyclohexyl, phenyl, 4-methoxyphenyl,
4-trifluoromethylphenyl, 2-furanyl, 2-thiophenyl, or 2-, 3- or 4-pyridyl
wherein the 2-furanyl,
2-thiophenyl and 2-, 3- or 4-pyridyl substituents are unsubstituted or
substituted as described
above for heteroaryl rings; R2 is H or -(CH2)PCH2OH, wherein p is 1-3;
especially those
wherein R, is H and X and Y are each H, and wherein q is 1-3 and r is 0 or
wherein q is 0
and r is 1-3. Among these compounds R2 is preferably H or -CH2-CH2-OH and the
sum of q
and r is preferably 1.
Those compounds of formula (le), wherein R20 is H or C,-C6alkyl, especially H,
are
important members of each of the subgenuses of compounds of formula (le)
described
above.
N-hydroxy-3-[4-[[(2-hydroxyethyl)[2-(1 H-indol-3-yl)ethyl]-
amino]methyl]phenyl]-2E-2-
propenamide, N-hydroxy-3-[4-[[[2-(1H-indol-3-yl)ethyl]-amino]methyl]phenyl]-2E-
2-
propenamide and N-hyd roxy-3- [4-[[[2-(2-m ethyl- 1 H-indol-3-yl)-ethyl]-
amino]methyl]phenyl]-
2E-2-propenamide or a pharmaceutically acceptable salt thereof, are important
compounds
of formula (le).
The present invention further relates to the compounds of the formula (If):
O Ri
HO~1 X R~e
H RZ R3 R4 O (~f)
Y P 4 r
A
or a pharmaceutically acceptable salt thereof. The variable substituents are
as defined
above.
Useful compounds of formula (If), are include those wherein R2 is H or
-(CH2)PCH2OH, wherein p is 1-3, especially those wherein R, is H; such as
those wherein R,
is H and X and Y are each H, and wherein q is 1-3 and r is 0 or wherein q is 0
and r is 1-3.
Among these compounds R2 is preferably H or -CH2-CH2-OH and the sum of q and r
is
preferably 1.
N-hydroxy-3-[4-[[[2-(benzofur-3-yl)-ethyl]-amino]methyl]phenyl]-2E-2-
propenamide or
a pharmaceutically acceptable salt thereof, is an important compound of
formula (If).
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The compounds described above are often used in the form of a pharmaceutically
acceptable salt. Pharmaceutically acceptable salts include, when appropriate,
pharmaceutically acceptable base addition salts and acid addition salts, e.g.,
metal salts,
such as alkali and alkaline earth metal salts, ammonium salts, organic amine
addition salts
and amino acid addition salts and sulfonate salts. Acid addition salts include
inorganic acid
addition salts, such as hydrochloride, sulfate and phosphate; and organic acid
addition salts,
such as alkyl sulfonate, arylsulfonate, acetate, maleate, fumarate, tartrate,
citrate and
lactate. Examples of metal salts are alkali metal salts, such as lithium salt,
sodium salt and
potassium salt; alkaline earth metal salts, such as magnesium salt and calcium
salt,
aluminum salt and zinc salt. Examples of ammonium salts are ammonium salt and
tetramethylammonium salt. Examples of organic amine addition salts are salts
with
morpholine and piperidine. Examples of amino acid addition salts are salts
with glycine,
phenylalanine, glutamic acid and lysine. Sulfonate salts include mesylate,
tosylate and
benzene sulfonic acid salts.
Additional HDAI compounds within the scope of formula (I), and their
synthesis, are
disclosed in WO 02/22577. Two preferred compounds within the scope of WO
02/22577
are:
OH O
\ / \ N"IOH
N H (~~)
N
H
= N-hydroxy-3-[4-[(2-hydroxyethyl){2-(1 H-indol-3-yl)ethyl]-
amino]methyl]phenyl]-2E-2-
propenamide, or a pharmaceutically acceptable salt thereof; and
O
\ ~OH
N
H
N \ (III)
N
H
= N-hydroxy-3-[4-[[[2-(2-methyl-1 H-indol-3-yl)-ethyl]-amino]methyl]phenyl]-2E-
2-
propenamide, or a pharmaceutically acceptable salt thereof.
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The present invention pertains in particular to the use of HDAC inhibitors for
the
preparation of a medicament for the treatment of gastrointestinal cancer.
An HDAC inhibitor as used for the present invention displays in the assay
described
above preferably an IC50 value between 50 and 2500 nM, more preferably between
250 and
2000 nM, and most preferably between 500 and 1250 nM.
Furthermore, the invention relates to a method of treating gastrointestinal
cancer,
especially hepatocellular carcinoma or pancreatic cancer, comprising
administering a
therapeutically effective amount of an HDAC inhibitor to a warm-blooded
animal, in particular
a human, in need thereof, preferably a therapeutically effective amount of a
compound of
formula (I), as defined above, or the salt of such compound having at least
one salt-forming
group, to a warm-blooded animal, preferably a human, in need thereof.
The term "treatment", as used herein, comprises the treatment of patients
having
gastrointestinal cancer or being in a pre-stage of said cancer which effects
the delay of
progression of the disease in said patients.
The present invention provides a method of treating gastrointestinal cancer,
especially hepatocellular carcinoma or pancreatic cancer, comprising
administering an
HDAC inhibitor in an amount which is therapeutically effective against
gastrointestinal
cancer, especially hepatocellular carcinoma or pancreatic cancer, to a warm-
blooded animal
in need thereof.
The person skilled in the pertinent art is fully enabled to select relevant
test models to
prove the hereinbefore and hereinafter mentioned beneficial effects on
gastrointestinal
cancer, of a compound inhibiting the HDAC activity. The pharmacological
activity of a
compound inhibiting the HDAC activity may, e.g., be demonstrated in a suitable
clinical study
or by means of the Examples described below.
The present invention also provides the use of a compound of formula (I), as
defined
herein, and the use of a COMBINATION OF THE INVENTION for the preparation of a
medicament for the treatment of lymphoproliferative diseases.
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Examples
Example 1
Monolayer Growth Inhibition Assay
The MTT is a colorimetric assay to determine the cell proliferation rate. The
yellow
tetrazolium MTT (3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium
bromide) is reduced by
metabolically active cells, in part by the action of dehydrogenase enzymes, to
generate
reducing equivalents such as NADH and NADPH. The resulting intracellular
purple formazan
can be solubilized and quantified by spectrophotometric means. The signals
produced is
directly proportional to the cell numbers. Describing the MTT assay in detail,
experiments
were done using six-point or 9 point drug titrations in multi-well tissue
culture dishes, with
outer rows left empty. Cells were suspended in complete media at densities of
between 103
and 104 cell/mI, respectively, and added per well. The appropriate medium (200
l) was then
added. Twenty-four hours later, 10 l of MTS solution , were added to one
plates to
determine the activity at the time of compound addition (To). This plate was
incubated at
37 C for 4 hours and the optical density was measured on a Molecular Devices
Thermomax
at 490 nm using the Softmax program. The To plate served as a reference for
initial activity
at the beginning of the experiment.
Compound addition began 24 hours after seeding, the same time as the To
determination.
Serial dilutions at 4-fold 2-fold, 1-fold, 0.5-fold, 0.25-fold and 0.125-fold
of previously
determined IC50 values of each compound were made in a 96-deep well plate with
the
highest concentrations on the edge of the plate. Each of the six dilutions
were added in
triplicate and complete medium was added to the empty outer rows without
cells. The
compounds were added to the plates singly or in combination with Compound III
(LBH589).
The plates were incubated at 37 C for 72 hours from seeding. The MTS solution
was added
(as for the To plate) and read four hours later. In order to analyze the data,
the average value
of media alone (background) was subtracted from each experimental well and the
triplicate
values were averaged for each compound dilution. The following formulas were
used to
calculate percent growth.
If X> To, % Growth = 100 x((X-To)/(GC -To))
If X < To, % Growth = 100 x(X-To)Ro)
To = average value of To minus background
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GC = average value of untreated cells (in triplicate) minus background
X = average value of compound treated cells (in triplicate) minus background
IC50 the concentration of LBH589 required to inhibit cell growth by 50% and
LD50s the
concentration required to reduce cell number (kill cells) to 50% the original
innoculum were
determined. The "% Growth" was plotted against compound concentration and used
to
calculate IC50s and LD50s , employing the user-defined spline function in
Microsoft Excel.
The Anti-proliferation and cytotoxic effects of LBH589 in a large panel of 36
colon cancer cell
lines are described in the attached Table:
NVP-LBH589
Cell Line IC50 nM LD50 (nM)
SW620 0.79 1.70
SW480 1.48 6.90
SW403 2.03 9.84
SW837 0.66 11.46
SW48 1.29 13.09
KM20L2 14.62 16.95
Colo201 15.12 20.14
CL-11 3.54 25.92
SNU-C1 4.51 26.07
SW1116 2.28 26.93
HT115 3.49 31.02
T84 2.16 32.82
Co1o205 14.68 33.52
Co1o741 9.22 33.73
SW948 5.06 34.92
WiDr 4.39 41.86
Co1o678 5.60 45.52
CaCo2 13.84 45.55
MDST8 4.87 48.44
HCT116 7.11 51.72
LOVO 4.37 52.01
HT29 2.83 70.23
HCT8 4.94 77.70
Co1o320 6.50 82.90
LS174T 16.77 87.41
HCC2998 29.67 154.87
LS1034 76.70 227.83
C2Bbe-1 27.46 243.12
DLD-1 61.40 296.27
HT55 8.21 334.52
C170 62.29 382.03
CoIo320DM 29.48 1103.37
RKO 5.57 2346.13
Colo206F 10.13 >10000
CoIo320HSR 13.82 >10000
HCT116 Bax -/- 1.73 >10000
Colon cancer cell lines were treated with DMSO vehicle control or varying
concentrations of
LBH589 for 3 days. Cell proliferation was measured on the day of cell plating
and on the
third day post-treatment. IC50 and LD50 values were calculated as described
above.
LBH589 exhibits potent anti-proliferative effect on all 36 colon cancer cell
lines examined, as
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demonstrated by the low nanomolar concentrations of IC50 values. LBH589 also
exhibits
potent cytotoxic effect in the great majority of the colon cancer cell lines
tested with LD50 < 1
M (n=31).
Example 2
Female athymic nude mice were implanted subcutaneously with HCT116 colon
cancer cells.
When tumors reached a medan tumor volume of 120 mm3, mice were randomized into
groups of 8 mice. Mice were treated with LBH589 at 5, 10 or 20 mg/kg
intravenously (iv) 5
times a week for 3 weeks or 75 mg/kg of 5-Fluorouracil intravenously once a
week for 3
weeks. Animals were calipered weekly. Compound activity was determined as the
percent
change in tumor volume of treated animals over control animals (%T/C). The
percentage of
regression was determined as the percent change in the final tumor volume at
the end of the
study over the starting tumor volume. Treatment with LBH589 at 5 or 10 mg/kg
inhibited
HCT1 16 tumor growth with %T/C of 17% and 6% respectively. Treatment with
LBH589 at 20
mg/kg resulted in tumor regression of 8%. The results are described in Figure
1.
Example 3
Female athymic nude mice were implanted subcutaneously with Co1o205 colon
cancer cells.
When tumors reached a medan tumor volume of 220 mm3, mice were randomized into
groups of 10 mice. Mice were treated with LBH589 at 30 mg/kg intravenously on
Monday,
Wednesday, Friday per week for 3 weeks, 75 mg/kg of 5-Fluorouracil
intraperitoneally once
a week for 3 weeks, or combination of the two agents. For tumor growth
inhibition (Figure
2A), compound activity was measured as the percent change in tumor volume of
treated
animals over control animals (%T/C). The percentage of regression was
determined as the
percent change in the final tumor volume at the end of the study over the
starting tumor
volume. For tumor growth delay (Figure 2B), compound activity was measured as
the
change in median calculated time to the study end point (TTE), which was
predefined as
tumor volume of 1000 mm3. Median TTE for vehicle, LBH, 5-FU, and combined
treatment
was 39.5 days, 32.9 days, 44.5 days, and 57.1 days respectively. Treatment
with LBH589 or
5-FU single agent led to tumor growth inhibition (TGI) and tumor growth delay
(TGD). More
importantly, the combined treatment significantly enhanced both TGI and TGD.
Figure 2
illustrates the results.
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Example 4
Pancreatic cancer cell lines were treated with DMSO vehicle control or varying
concentrations of LBH589 for 3 days. Cell proliferation was measured on the
day of cell
plating and on the third day post-treatment. IC50 and LD50 values were
calculated as
described above. LBH589 exhibits potent anti-proliferative effect on all 12
pancreatic cancer
cell lines examined, as demonstrated by the low nanomolar concentrations of
IC50 values.
LBH589 also exhibits strong cytotoxic effect in the majority of the pancreatic
cancer cell lines
tested with LD50 < 1 M (n=10).
Table 2 describes the anti-proliferative and cytotoxic effects of LBH589 in a
panel of 12
pancreatic cancer cell lines.
NVP-LBH589
Cell Lines IC50 nM LD50 nM
MiaPaCa2 14.1 59.5
BxPC3 15.9 104.8
Hs766T 14.1 119.9
SU.86.86 51 170.4
Panc1 18.9 175.4
Capanl 55.6 191.5
Panc3.27 38.5 276.8
PanclO.05 31.8 282.4
AsPC1 25.1 372.4
Panc2.03 22.4 599.5
HPAC 122.7 2051.6
Panc4.03 20.3 >10000
A panel of 19 pancreatic cancer cell lines was independently assessed in cell
proliferation
assays. Cells were treated with DMSO vehicle control or varying concentrations
of LBH589
for 6 days. Consistent with results presented in Table 2, LBH589 exhibits
potent anti-
proliferative effect on all 19 pancreatic cancer cell lines, showing low
nanomolar
concentrations of IC50 values. LBH589 also exhibits potent cytotoxic effect in
18 of the 19
pancreatic cancer cell lines, with LD50 < 1 M. The results are described in
Figure 3.
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