Note: Descriptions are shown in the official language in which they were submitted.
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CYCLIC PEPTIDE ANTIFUNGAL AGENq'~
Back~round o~ the Invention
This invention relates to semi-~ynthetic cyclic
peptide compounds which are useful as anti~ungal and
antiparasitic agents and which have impro~ed stability and
water solubility. In particular, it relates to derivatives
of the echinocandin class of cyclic peptides, to methods
for treating fungal and parasitic infections and to
formulations useful in the methods.
The compounds provided by this invention are
semi-synthetic compounds derived from cyclic peptides which
are prcduced by culturing various microorganisms. A number
of cyc ic peptides are known in the art including
echinocandin B (A30912A), aculeacin, mulundocandin,
sporiofungin, L-671,329, and S3~794/F1.
In general, these cyclic peptides may ~e
charac~erized as a cyclic hexapeptide core (or nucleus)
with an acylated amino group on one of the core amino
acids. The amino group is typically acylated with a ~atty
acid group forming a side chain off the nucleus. For
exampl_, echinocandin B has a linoleoyl side chain while
aculeacin has a palmitoyl side chain.
The fatty acid side rh~in~ may be removed from
the cyclic peptide core to provide an amino nucleus tfor
example, a compound of formula I, below, where R2 is
hydrogen). The amino group may then be re-acylated to
provide semi-synthetic compounds such as those claimed in
the present application.
The echinocandin B nucleus has been re-acylated
with certain non-naturally occurring side chain moieties to
provide a number of antifungal agents (see, ~ebono, U.S.
Pat. Ser. No. 4,293,489). Among such antifungal agents is
~ cilofungin which is represented by a compound of formula IA
3~ where -', R" and R"' are methyl, RXl~ RX2~ RYl~ RY2 RY3 RY4
and R0 are each hydroxy and R2 is p-(octyloxy)benzoyl.
-
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Su~unarv of the Invention
The present invention provides a compound of
formula I
N RY2 O
~e O HN
R ~< o~ \ R"
OH NH OH
N~
~ RY4
~R~'3
wherein:
R' is hydrogen, methyl, -CH2CH2NH2 or
-CH2C (O)NH2;
R" and R"' are independently methyl or hydrogen;
RX1 is hydrogen, hydroxy, -NH-R, or -O-R;
R iS C1-C6 alkyl, benzyl, -(CH2)2Si(CH3) 3,
-CH2CHOHCH20H, -CH2CH=CH2, - (CH2)aCOOH, - (CH2)bNRZlRZ2,
- (CH2 ) cPoRZ3RZ4 or -~(CH2 ) 2~] d- (Cl-C6 ) alkyl;
a, b and c are independently 1, 2, 3, 4, 5 or 6;
Rzl and Rz2 are independently hydrogen,
Cl-C6 alkyl, or RZl and RZ2 combine to ~orm -CH2 (CH2 ) eCH2-;
RZ3 and RZ4 are independently hydroxy or
C1-C6 alkoxy;
d is 1 or 2;
e is 1, 2 or 3;
Rx2 Ryl, RY2, RY3 and RY4 are independently
hvdroxy or hydrogen;
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.. 3
R0 iS hydroxy, -OP(O)(OH~2 or a grou~ o~ the
formula:
O-~-Rl or - O~ ORl ;
OH OH
Rl is Cl-C6 alkyl, phenyl, p-halo-phenyl,
p-nitrophenyl, benzyl, p-halo-benzyl or p-nitro-benzyl;
R2 iS
- C ~ X ~ - Y- ~ R3
A, B, and C are independently selected from the
following groups:
N> ~ N ~ ~
X and Y are independently a bond or -C-C-:
R3 iS Cl-C12 alkyl, Cl-cl2 alkoxy or
~O-(CH2)m~~O-(CH2)n]p-O-(Cl-C 12 alkyl);
m is 2, 3 or 4;
n is 2, 3 or 4; and
p is 0 or 1;
with the proviso that A, B, and C cannot all be
or a pharmaceutically acceptable salt thereof.
The present invention also provides a compound
of formula II
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R' ~ ~ N- R2
O ~ \ R"
~ ~ ~ OH II
~ RY3 RY4
R~
wherein:
R' is hydrogen, methyl or -cH2c(o)NH2;
R" and R"' are independently methyl or hydrogen;
5 . RXl is hydrogen, hydroxy or -O-R;
R is C1-C6 alkyl, benzyl, -(CH2)2Si(CH3)3,
-CH2CHOHCH20H, -CH2CH=CH2, -(CH2)aCOOH, -(CH2)bNRZlRZ2,
-(CH2)CPoRZ3Rz4 or -[(CH2)20]d-~C1-C6)alkyl;
a, b and c are independently 1, 2, 3, 4, 5 or 6;
Rzl and Rz2 are independently hydrogen,
C1-C6 alkyl, or R~1 and Rz2 combine to ~orm -CH2(CH2;~CH2-;
R~3 and RZ4 are independen~ly hydroxy or
C1-C6 alkoxy;
d is 1 or 2;
e is 1, 2 or 3;
Rx2, Ryl, RY2, RY3 and RY4 are independently
hydroxy or hydrogen;
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R0 is hydroxy, -OP(O)(OH)2 or a croup of the
formula: .
O-~-R1 or O-~- ORl ;
OH OH
Rl is Cl-C6 alkyl, phenyl, p-halo-phenyl,
p-nitrophenyl, benzyl, p-halo-benzyl or p-nitro-benzyl;
R2 is
- C ~ x ~ -y- ~ R3
A, B, and C are independently selected from the
~ollowing groups:
~ ~ G
~N ' M=9 ' ~, 6~
X and Y are independently a bond or -C~C-;
R3 is C1-C12 alkyl, C1-C12 alkoxy or
~0-(CH2)m-[0-(CH2)n]p-0-(Cl-C12 alkyl);
m is 2, 3 or 4;
n is 2, 3 or 4; and
p is 0 or 1;
with the proviso that A, B, and C cannot all be
or a pharmaceutically acceptable salt thereof.
Also provided are pharmaceutical formulations,
methods for inhibiting parasitic or fungal activity and
methods of treating fungal or parasitic in~ections which
employ the compounds of the invention.
a 25 The present invention also provides for the use
of compounds of the invention for: inhibiting fungal
activity, treating fungal infection, inhibiting parasitic
activity, and treating or preventing the onset of
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_
Pneumocys~is pneumonia in a host suscepti~le t~
Pneumocystis pneumonia.
Detailed D~scri~tion
As used herein, the term ~ICl-cl2 alkyl'~ refers to
a straight or branched alkyl chain having from one to
twelve carbon atoms. Typical C1-C12 alkyl groups include
methyl, ethyl, propyl, isopropyl, butyl, sec-butyl, t-
butyl, pentyl, 5-methylpentyl, hexyl, heptyl, 3,3-
dimethylheptyl, octyl, 2-methyl-octyl, nonyl, decyl,
undecyl, dodecyl and the like. The term l'Cl-Cl2 alkyl~
includes within its definition the terms "C1-C6 alkyl~ and
C1-C4 alkyl.~
The term "halo" refers to chloro, fluoro, bromo
or iodo.
The term "C1-C12 alkylthio" refers to a straight
or branched alkyl chain having from one to twelve carbon
atoms attached to a sulfur atom. Typical Cl-C12 alkylthio
groups include methylthio, ethylthio, propylthio,
isopropylthio, butylthio, 3-methyl-heptylthio, octylthio,
5,5-dimethyl-hexylthio and the like.
The term IIC1-C12 alkoxy'~ refers to a straight or
branched alkyl chain having from one to twelve carbon atoms
attached to an oxygen atom. Typical Cl-C12 alkoxy groups
include methoxy, ethoxy, propoxy, butoxy, sec-butoxy,
pentoxy, ~-methyl-hexoxy, heptoxy, octyloxy, decyloxy
dodecyloxy and the like. The term "C1-C12 alkyl~ includes
within its definition the terms "C1-C6 alkoxy~ and
Cl-C4 alkoxy."
The term "hydroxy protecting group~ refers to a
substituent of an hydroxy group that is commonly employed
to block or protect the hydroxy functionality while
reactions are carried out on other functional groups on the
compound. ExampleS of such hydroxy protecting groups
include tetrahydropyranyl, 2-methoxyprop-2-yl, 1-ethoxyeth-
l-yl, methoxymethyl, ~-methoxyethoxymethyl,
methylthiomethyl, t-~utyl, t-amyl, trityl, 4-methoxytrityl,
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4,4'-dimethoxytrityl, 4,4',4"-trimethoxytrityl, benz~l,
allyl, trimethylsilyl, trimethylsilylethyl, (t-
bu~yl)dlmethylsilyl, and 2,2,2-trichloroethoxycarbonyl and
the like. The species of hydroxy protecting group is not
critical so long as the derivatized hydroxy group is stable
to the conditions of the subsequent reaction~s) and can be
removed at the appropriate point without disrupting the
remainder of the molecule. A preferred hydroxy protecting
group is trimethylsilylethyl. Further examples of hydroxy
protecting groups are described in T.W. Greene, "Protective
Groups in organic Synthesis," John Wiley and Sons, New
York, N.Y., t2nd ed., 1991) chapters 2 and 3. The term
~protected hydroxy" refers to a hydroxy group bonded to one
of the above hydroxy protecting groups.
The term '~amino protecting group~ as used in the
speci~ication refers to substituents of the amino group
commonly employed to block or protect the amino
functionality while reacting other functional groups on the
compound. Examples of such amino protecting groups include
formyl, trityl, phthalimido, trichloroacetyl, chloroacetyl,
bromoacetyl, iodoacetyl groups, or urethane-type blocking
groups such as benzyloxycarbonyl, 4-
phenylbenzyloxycarbonyl, 2-methylbenzyloxycarbonyl, 4-
methoxybenzyloxycarbonyl, 4-fluorobenzyloxycarbonyl, 4-
chlorobenzyloxycarbonyl, 3-chlorobenzyloxycarbonyl, 2-
chlorobenzyloxycarbonyl, 2,4-dichlorobenzyloxycarbonyl, 4-
bromobenzyloxycarbonyl, 3-bromobenzyloxycarbonyl, 4-
nitrobenzyloxycarbonyl, 4-cyanobenzyloxycarbonyl, t-
butoxycarbonyl, 2-(4-xenyl)isopropoxycarbonyl, 1,1-
diphenyleth-1-yloxycarbonyl, 1,1-diphenylprop-1-
yloxycarbonyl, 2-phenylprop-2-yloxycarbonyl, 2-(p-toluyl)-
- prop-2-yloxycarbonyl, cyclopentanyloxycarbonyl, 1-
methylcyclopentanvloxycarbonyl, cyclohexanyloxycarbonyl, 1-
- methylcyclohexanyloxycarbonyl, 2-
methylcyclohexanyloxycarbonyl, 2-~4-toluylsulfonyl)-
ethoxycarbonyl, 2-(methylsul~onyl)ethoxycarbonyl, 2-
(triphenylphosphino)-ethoxycarbonyl, fluorenylmethoxy-
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carbonyl ("FMOC"), 2-(trimethylsilyl)ethoxycarbonyl
allyloxycarbonyl, 1-(trimethylsilylmethyl)prop-1-
enyloxycarbonyl, 5-benzisoxalylmethoxycarbonyl, 4-
acetoxybenzyloxycarbonyl, 2,2,2-trichloroethoxycarbonyl, 2-
ethynyl-2-propoxycarbonyl, cyclopropylmethoxycarbonyl, ~-
(decyloxy)benzyloxycarbonyl, isobornyloxycarbonyl, ~-
piperidyloxycarbonyl and the likei benzoylmethylsu~fonyl,
2-nitrophenylsulfenyl, diphenylphosphine oxide and like
amino protecting groups. The species of amino protec~ing
group employed is not critical so long as the derivatized
amino group is stable to the condition of subse~uent
reaction(s) on other positions of the intermediate molecule
and can be selectively removed at the appropriate point
without disrupting the remainder of the molecule including
any other amino protecting group(s). Preferred amino
protecting groups are t-butoxycarbonyl (t-Boc),
allyloxycarbonyl and benzyloxycarbonyl (CbZ). Further
examples of groups referred to ~y the above terms are
described by J. W. Barton, "Protective Groups in Organic
Chemistry~, J. G. W. McOmie, Ed., Plenum Press, New York,
N.Y., 1973, Chapter 2, and T. W. Greene, "Protective Groups
in Organic Synthesis~, John Wiley and sons, New York, N.Y.,
1981, Chapter 7.
The term llinhibiting~, i.e. a method of
inhibiting parasitic or fungal activity, includes stopping,
retarding or prophylactically hindering or preventing the
growth or any attending characteristics and results from
the existence of a parasite or fungus.
The term "contacting~, i.e. contacting a
compound of the invention with a parasite or fungus,
includes a union or junction, or apparent touching or
mutual tangency of a compound of the invention with a
parasite or fungus. However, the term does not imply any
further limitations to the process, such as by mechanism of
inhibition, and the methods are defined to encompass the
spirit of the invention, which is to inhibit parasitic and
fungal activity by the action of the compounds and their
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inherent antiparasitic and antifungal prop~rties, or in
other words, the compounds, used in the claimed methods are
the causative agent for such inhi~ition.
The term l~pharmaceutically acceptable salt" as
used herein, refers ~o salts of the compounds of the above
formula which are substantially non-toxic to living
organisms. Typical pharmaceutically acceptable salts
include those salts prepared by reaction of the compounds
of the present invention with a mineral or organic acid or
an inorganic base. Such salts are known as acid addition
and base addition salts.
Acids commonly employed to form acid addition
salts are mineral acids such as hydrochloric acid,
hydrobromic acid, hydroiodic acid, sulfuric acid,
phosphoric acid and the like, and organic acids such as
p-toluenesulfonic, methanesulfonic acid, oxalic acid,
p-bromophenylsulfonic acid, carbonic acid, succinic acid,
citric acid, benzoic acid, acetic acid, and the like.
Examples of such pharmaceutically acceptable salts are the
sulfate, pyrosulfate, bisulfate, sulfite, bisulfite,
phosphate, monohydrogenphosphate, dihydrogenphosphate,
metaphosphate, pyrophosphate, chloride, bromide, iodide,
acetate, propionate, decanoate, caprylate, acrylate,
formate, isobutyrate, caproate, heptanoate, propiolate,
oxalate, malonate, succinate, suberate, sebacate, fumarate,
maleate, butyne-1,4-dioate, hexyne-1,6-dioate, benzoate,
chlorobenzoate, methylbenzoate, dinitrobenzoate,
hydroxybenzoate, methoxybenzoate, phthalate, sulfonate,
xylenesulfonate, phenylacetate, phenylpropionate,
phenylbutyrate, citrate, lactate, ~-hydroxybutyrate,
glycollate, tartrate, methanesulfonate, propanesulfonate,
naphthalene-1-sulfonate, napththalene-2-su~onate,
mandelate and the like. Preferred pharmaceutically
acceptable acid addition salts are those formed with
mineral acids such as hydrochloric acid and hydrobromic
acid, and those formed with organic acids such as maleic
acid and methanesulfonic acid.
_
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Base addition salts include ~hose derived from
inorganic bases, such as ammonium or alkali or alkaline
earth metal hydroxides, carbonates, bicarbonates, and the
like. Such bases useful in preparing the salts of this
invention thus include sodium hydroxide, potassium
hydroxide, ammonium hydroxide, potassium carbonate, sodium
carbonate, sodium bicarbonate, potassium bicarbonate,
calcium hydro~ide, calcium carbonate, and the like. trhe
potassium and sodium salt forms are particularly preferred.
It should be recognized that the particular
counterion forming a part of any salt of this invention is
not of a critical nature, so long as the salt as a whole is
pharmacologically acceptable and as long as the counterion
does not contribute undesired qualities to the salt as a
whole.
Preferred compounds of this invention are those
compounds of formula I where:
R', R" and R"' are each methyl;
RYl, RY2, RY3 and RY4 are each hydroxy;
~0 RXl is hydrogen, hydroxy or -O-R;
R is methyl, benzyl, -CH2CHOHCH20H, -(CH2)bNRZlRZ2
or -(CH2)2PORZ 3 RZ4;
b is 2, 3, 4, 5 or 6;
RZl and Rz2 are independently hydrogen or
Cl-C4 alkyl;
RZ3 and RZ4 are independently hydroxy or methoxy;
RX2 is hydrogen or hydroxy;
R0 is hydroxy, -OP(O)(OH)2 or a group of the
formula:
11o
O~ Rl orO-P- oRl ;
OH OH
Rl is me~hyl;
or a pharmaceutically acceptable salt thereof.
of these compounds, more pre~erred are those
compounds of formula I where:
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_
RXl is hydrogen or hydroxy;
Rx2 is hydrogen or hydroxy;
R~ is hydroxy;
R3 is Cl-C12 alkoxy or -O-(CH2)2-O-(Cl-C12 alkyl);
or a pharmaceutically acceptable salt thereof.
.
Of these compounds, further preferred are those
compounds of formula I where:
RXl is hydroxy;
RX2 is hydroxy;
x and Y are a bond;
R3 is Cl-Cg alkoxy;
or a pharmaceutically acceptable salt thereo~.
Of these compounds even more preferred are those
compounds of formula 1 wherein:
A is ~ , ~ ~ or
B is ~ , ~ ~ ~ or
C is ~ , ~ ~ ~ or ~
or a pharmaceutically acceptable salt thereo~.
The compounds of formula I may be prepared
according to Reaction Scheme I, as follows.
_
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12 PCT/US97/01607
Reaction Scheme
RYl ~
R'~ ~ N--~ na
R'~C ~>
0~ NH OH
N ~ (IA)
~RY3 RY4
R~
A. deacylate
RYl ~ RXl
R' ~ N ~ N- H
~ N H R~2 ~
R' ~ O ~ \ R"
OH NH OH
O ~ ~ (IB)
Rx ~ RY3o RY4
~o
B. ~e-acylate
-
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R~ ~ H ~ N- R2
~N
' ~ O ~ R"
OH NX OH (I)
, ~ ~ RY4
Rx~ ~RY3
R~
wherein:
Rnat is a naturally occurring cyclic peptide
sidechain; and
R , R , R , Rx~, Rx~, Ry1, RY~, RY3, RY~, R0 and
R2 are as defined above.
Reaction scheme I, above, is accomplished by
carrying out reactions A and B, above. Once a reaction is
complete, the intermediate compound may be isolated by
procedures well-known in the art, for example, the compound
may be crystallized or precipitated and then collected by
filtra~ion, or the reaction solvent may be removed by
extraction, evaporation or decantation. The intermediate
compound may be further purified, if desired, by common
techni~ues such as crystallization or precipitation or
chromatography over solid supports such as silica gel,
alumina and the like, before carrying out the next step of
the reaction scheme.
In reaction IA, a naturally occurring cyclic
peptide of the formula IA is deacylated using procedures
known in the art to provide an amino nucleus of formula IB.
This reaction is typically carried out using enzymatic
deacylation by exposing the naturally occurring cyclic
peptide to a deacylase enzyme. The deacylase enzyme may be
obtained ~rom the microorganism Actinoplanes utahensis and
,
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-
used substantially as described in U.S. Patent Nos.
4,293,482 and 4,304,716, herein incorporated by reference.
The deacylase enzyme may also be obtained from ~he
Pseudomonas species. Deacylation may be accompiished using
whole cells of Actino~lanes utahensis or Pseudomonas or the
crude or purified enzyme thereof or using an immobilized
form of the enzyme. $ee European Patent Application
No. 0 460 882 (December 11, lg91). Examples of naturally
occurring cyclic peptides which may be used as starting
materials include aculeacin (palmitoyl side chain),
tetrahydroechinocandin B (stearoyl side chain),
mulundocandin (branched C1s side chain), L-671,329
(C16 branched side chain), S 317~4/F1 (tetradecanoyl side
chain), sporiofungin (C1s branched side chain), -R~01379
(palmitoyl side chain) and the like. A preferred naturally
occurring cyclic peptide is echinocandin B ~a compound of
formula IA where R', R" and R"' are each methyl, RX1, RX2
RYl~ Ry2~ RY3, RY4 and Ro are each hydroxy and R2 is
linoleoyl).
In Reaction IB, the amino nucleus of ~ormula Is
is then re-acylated using procedures known in the art to
provide a compound of formula I where Ro is hydroxy; RXl is
hydroxy; and R2 is an acyl group as defined hereinabove.
For example, the amino nucleus may be acylated
by reaction with an appropriately substituted acyl halide,
preferably in the presence of an acid scavenger such as a
tertiary amine, such as triethylamine. The reaction is
typically carried out at a temperature of from about -20~C
to about 25~C. Typical solvents for this reaction include
polar aprotic solvents such as dioxane or
dimethylformamide. Solvent choice is not critical so long
as the solvent employed is inert to the ongoing reaction
and the reactants are sufficiently solubilized to e~fect
the desired reaction.
The amino nucleus may also be acylated by
reaction with an appropriately substituted carboxylic acid,
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in the presence of a coupling agent. Typical coupl_ng
agents include dicyclohexylcarbodiimide (DCC),
N,N'-carbonyldiimidazole, bis(2-oxo-3-
oxazolidinyl)phosphinic chloride (soP-cl),
N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ),
benzotriazol-1-yloxytripyrrolidinophosphonium
hexafluorophosphate (PyBOP) and the like.
In addition, the amino nucleus may be acylated
with an activated ester of a carboxylic acid such as an
ester of a carboxylic acid o~ the ~ormula R2-cOOH and
p-nitrophenyl, 2,4,5-trichlorophenyl, hydroxybenzotriazole
hydrate (HOBT H2O), penta~luorophenol, N-hydroxysuccinimide
and the like. Pre~erred acylating moieties are the active
esters of the carDoxylic acid R2-COOH such as a
benzotriazole ester. The reaction is typically carried out
for one to sixty five hours at a temperature ~rom about 0~C
to about 30~C in an aprotic solvent. The reaction is
generally complete after about twenty four to forty eight
hours when carried out a temperature of ~rom about 15~C to
about 30~C. Typical solvents ~or this reaction are
tetrahydrofuran and dimethyl~ormamide or a mixture o~ such
solvents. The amino nucleus is generally employed in
equimolar proportions relative ~o the activated ester or
with a slight excess of the amino nucleus.
The compounds o~ ~ormula I where RXl is hydroxy
may be reacted with an appropriately substituted alcohol in
the presence of an acid to provide a compound of formula I
where RXl is -O-R, where R is Cl-C6 alkyl, benzyl,
-(CH2)2Si(CH3)3, -CH2CH=CH2, -(CH2)aCOOH, -(CH2)bNRZlRZ2,
-(CH2)CPoRZ3RZ~ or -[(CH2) 2~ ] d- ( C 1 -C 6)alkyl. The reaction
is typically carried out in a polar aprotic solvent such as
dioxane or dimethyisulfoxide at a temperature o~ ~rom about
0~C to about 35~C, preferably at about room temperature.
Solvent choice is not critical so long as the solvent
employed is inert to the ongoing reaction and the reactants
are sufficiently solubilized to effect the desired
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16
reaction. Preferred acids include p-toluenesulfon acid,
hydrochloric acid and camphorsulfonic acid.
The compounds of formula I where RXl is
-~CH2)bNRZlRz2 where RZl and Rz2 are hydrogen may be
prepared via a protected compound wherein RXl is
-~CH2)bNHRa where Ra is an amino protecting group. The
resultant protected compound is then deprotected according
to procedures known in the art.
The compounds of formula I where RXl is
-CH2CHOHCH20H may be prepared by hydroxylating a compound
of formula I where RXl is -CH2CH=CH2 with osmium tetroxide
in the presence of a catalyst at a temperature in the range
of from about 0~C to about 40~C for about one to twen~y
four hours in a organic/a~ueous solvent mixture, for
example dioxane/water. Suitable catalysts include N-
methylmorpholine N-oxide (MMO) and the like. Typical
solvents suitable for use in this reaction include
dimethylformamide, tetrahydrofuran, acetone and dioxane.
Solvent choice is not critical so long as the solvent
employed is inert to the ongoing reaction and the reactants
are sufficiently solubilized to effect the desired
reaction. The reaction is preferably conducted at a
temperature in the range of from about 20~C to about 30~C
for about eighteen to twenty four hours.
The compounds of formula I where R0 is hydroxy
may be phosphorylated by reaction with an appropriately
substituted alkyl or phenyl phosphate to provide a compound
of formula I where Ro is -0-P(O)OH-R1 where R1 is C1-C6
alkoxy or phenoxy, or by reaction with an appropriately
substituted alkyl or phenyl phosphonic acid to provide a
compound o~ formula I where Ro is -0-P(O)OH-R1 where R1 is
C1-C6 alkyl, or an appropriately substituted phenyl or
benzyl moiety, ~o provide a compound of formula I where R~
is a group of the formula -OP(O)OH-R1. The phosphonic acid
is typically used in an activated form, for example as a
phosphonic halide, preferably a phosphonic chloride. The
reaction is carried out in the presence of a base such as
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17
lithium trimethylsilanolate ~LioTMs)t lithium
bis(trimethylsilyl)amide (LHMDS), pyridine and the like.
The reaction is typically carried out for up to one hour at
a temperature from about -30~C to about 0~C in an aprotic
solvent such as tetrahydrofuran and dimethylformamide. The
reaction is generally complete in about fifteen minutes
when carried out under these conditions. The phosphate or
phosphonate reactant is generally employed in e~uimolar
proportions to about a one mole excess relative to the
amino nucleus in the presence of an equimolar or slight
excess of the base. Phosphorylation of an amino nucleus
with unprotected aminal hydroxy groups is typically carried
out at lower temperatures, for exa~ple from about -30~C to
about -15~C.
Alternatively, the ~ml n~ 1 hydroxy moieties on
the compound of formula I are optionally protected with an
hydroxy protecting group using procedures known in the art.
For example, the reaction is typically carried out by
combining the compound of formula I with a suitable hydroxy
protecting group in the presence of a catalyst at a
temperature in the range of from about 0~C to about 40~C
for about one to five hours in a mutually inert solvent.
The hydroxy protecting group is generally employed in an
amount ranging ~rom about equimolar proportions to about a
100 molar excess relative to the compound of formula I,
preferably in a large molar excess. ~uitable catalysts
include strong acids such as p-toluenesulfonic acid,
camphorsulfonic acid tCSA), hydrochloric acid, sulfuric
acid, trifluoroacetic acid and the like. Typical solvents
suitable fcr use in this reaction include any organic
solvent such as dioxane. Solvent choice is not critical so
long as the solvent employed is inert to the ongoing
reaction ard the reactants are sufficiently solubilized to
effect the desired reaction. The reaction is preferably
conducted at a temperature in the range of from about 20~C
to about 3Q~C for about two to four hours. The protected
com~ound of formula I is then phosphorYlated as described
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18
above. The hydroxy protecting group(s) are then removed
according to procedures known in the art to provide a
phosphorylated compound of formula I. For example, the
protecting groups can be removed by reaction with a Lewis
acid in a mutual inert organic solvent such as methylene
chloride. Examples of Lewis acids include
trimethylsilylbromide, boron trifluoride etherate and the
like. The reaction is typically carried out at a
temperature of from about 0~C to about 40~C, preferably at
a temperature of from about 20~C to about 30~C. A
preferred Lewis acid is boron trifluoride etherate.
The dideoxy compounds of formula I are prepared
by removing the benzylic and aminal hydroxy groups (RX2 and
RXll respectively). The hydroxy groups may be removed by
subjecting a non-dideoxy compound of formula I (where R2 is
hydrogen or acyl) to a strong acid and a reducing agent at
a temperature of between -5~C and 70~C, in a suitable
solvent. Typical strong acids include trichloroacetic
acid, trifluoroacetic acid or boron trifluoride etherate.
2~ A preferred strong acid is trifluoroacetic acid. Typical
reducing agents include sodium cyanoborohydride or
triethylsilane. A preferred reducing agent is
triethylsilane. Suitable solvents include methylene
chloriàe, chloroform or acetic acid, preferab~y methylene
chloride. The strong acid should be present in an amount
of from 2 to 80 mol per mol of substrate, and the reducing
agent should be present in an amount of 2 to 80 mol per mol
of substrate. This process affords selective removal of
the aminal and benzylic hydroxy groups.
The cyclic peptides used to make the compounds
of the present invention may be prepared by fermentation of
known microorganisms. For example, the cyclic peptide of
formula IB where R', R" and R"' are methyl, RXl, RX2, Ryl
RY2, RY3, RY4 and R0 are each hydroxy (cyclic nucleus
corresponding to A-30912A) may be prepared using the
procedure detailed in ~hhott e~ al., U.S. Pat. Ser. No.
4,293,482, which is herein incorporated by reference. The
-
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cyclic peptide of formula IB where R', R and R"' are
methyl, Rxl is hydroxy, RX~ is hydrogen, RYl, RY2, RY3, RY4
and R~ are each hydroxy (cyclic nucleus corresponding to
A-30912B) may be prepared using the procedure detailed in
~hhott et al., U.S. Pat. Ser. No. 4,299,763, which is
herein incorporated by reference. Aculeacin may be
prepared using the procedure detailed in M; zllno et ~l.,
U.S. Pat. Ser. No. 3,978,210 which is herein incorporated
by reference. The cyclic peptide of formula IB where R' is
-CH2C(O)NH2, R" is methyl, R"' is hydrogen, RXl~ Rx2 Ry
RY2, RY3, RY4 and R0 are each hydroxy may be prepared by
deacylating the cyclic peptide prepared using the procedure
detailed in Chen et al., U.S. Pat. Ser. No. 5,lg8,421,
which is herein incorporated by reference.
The R2-COOH precursor acids may be obtained
commercially or prepared according to procedures known in
the art. For example, an appropriately substituted aryl
boronic acid or biaryl boronic acid reactant may be reacted
with a haloaryl carboxylic acid reactant in the presence of
a catalyst such as tetrakis(triphenylphosphine)palladium
and an inorganic base such as potassium carbonate in a
mutual inert organic solvent such as toluene at a
temperature of from about 20~C to the reflux temperature of
the reaction mixture to provide the corresponding biaryl
carboxylic acids and teraryl carboxylic acids used to
prepare the compounds of formula I. The reaction is
typically carried out with equimolar proportions of the
boronic acid reactant and the aryl carboxylic acid
reactant, or a slight molar excess of the aryl carboxylic
acid reactant relative to the boronic acid reactant, and a
1-2 molar excess of the inorganic base. The reaction is
generally complete after about four to about ten hours when
carried out at reflux temperature in toluene.
The boronic acid reactant may be prepared by
reacting an appropriately substituted haloaryl or
halobiaryl reactant with two e~uivalents of triisopropyl
borate in the presence of an alkyl lithium, for example
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sec-butyl lithium, in a mutual inert solvent such as
tetrahydrofuran. The alkyl lithium is typically employed
in a slight molar excess relative to the haloaryl or
halobiaryl reactant. The alkyl lithium is typically
combined with the solvent by dropwise addition at reduced
temperatures (<-70~C) and allowed to stir for approximately
thirty minutes before the addition of the triisopropyl
borate. The reaction is typically carried out initially at
a temperature of from about -100~C to about -50~C,
preferably from about -75~C to about -85~C for thirty
minutes to two hours and then warmed to room temperature
and reacted ~or an additional one to three hours. The
reaction is generally complete in several minutes to about
four hours. When the reaction is substantially complete,
the boronic acid moiety is formed by the addition of an
acid. A preferred acid is a 1~ hydrochloric acid solution.
The R2-COO~ precursor acids having an acetylene
-moiety may be prepared by reacting an appropriately
substituted acetylene reactant with an appropriately~0 substituted ar~l or biaryl reactant of the formula
~OOC ~ L or HOOC- ~ L
where ~ is a suitable leaving group such as bromo, iodo,
methanesulfonate, toluenesulfonate,
trifluoromethanesulfonate and the like, in the presence of
a catalyst and preferably in the presence of an acid
scavenger in a mutual inert solvent such as acetonitrile.
Examples of acid scavengers include triethylamine and
pyridine, preferably triethylamine. A preferred catalyst
is formed in si u from palladium ~II) chloride,
triphenylphosphine and copper (I) iodide. The reaction is
typically carried out for thirty minutes to twenty one
hours at a temperature from about room temperature to the
reflux temperature of reaction mixture. The reaction is
generally comple~e after about two to about six hours when
carried out at reflux temperature.
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21
-
Alternatively, a suitably substituted aryl
J reactant of the formula ~ may be reacted with an
appropriately substituted acetylene reactant as described
above to provide, for example, a compound of the formula
halo ~ C-- C~ 3c
which can be coupled with a~ aryl
boronic acid reactant as described above.
Compounds of the present invention wherein RX1 is
NHR or where R' is CH2C~I2NH2 can be prepared by procedures
and schemes disclosed herein in combination with procedures
10 well known in the art. For example, such procedures are
exemplifed by but not limited to the following publictions:
W094~25~48; W094/25050; WO 96/08266; and WO 96/08507.
The following Preparations and Examples further
describe how to synthesize the compounds of ~he present
1~ invention. The terms melting point, proton nuclear
magnetic resonance spectra, mass spectra, infrared spectra,
ultraviolet spectra, elemental analysis, high performance
liquid chromatography, and thin layer chromatography are
abbreviated ~m.p.~, "NMR", "MS", "IR", ~IWII, "Analysis~,
20 ~HPLC~ and "TLC", respectively. In addition, the
absorption maxima listed for the IR spectra are only those
of interest and not all of the maxima observed.
Pre~aration 1
6-Chloronicot;nic ~cid, methyl ester
Hydrochloric acid (gas) was bubbled through a
solution of 6.11 g (38.8 mmol) of 6-chloronicotinic acid in
approximately 350 ml o~ methanol. The resultant reaction
mixture was reacted for approximately two hours at reflux
temperature, then cooled to room temperature and
concentrated in vacuo ~o provide a white residue. This
residue was redissolved in methylene chloride and the
resultant solution was washed with a saturated sodium
bicarbonate solution. The organic portion was dried over
sodium sulfate, filtered and then reduced to dryness in
vacuo to provide 7.28 g of a tan solid. This solid was
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22
-
crystallized from pentane, followed by recrystallization
from hexane to provide the desired titled compound.
Pre~aration 2
A. l-Bromo-4-~entoxv-benzene
To a solution containing 25.017 g (0.14~ mol) of
4-bromo-phenol and 24.44 g (0.218 mol) of potassium
t-butoxide in 500 ml of tetrahydrofuran, was added 27 ml
(0.218 mol) of l-bromopentane, via syringe. The resultant
reaction mixture was reacted overnight at re~ux
temperature. When the reaction was substantially complete,
as indicated by TLC, the reaction mixture was filtered.
The filtrate was concentrated in vacuo to provide a residue
which was redissolved in diethyl ether and the resultant
solution was washed se~uentially with water and lM sodium
hydroxide. The organic portion was dried over sodium
sulfate, filtered and then concentrated in vacuo to provide
3.062 g of an oil that was used without further
purification.
Yield: 87~.
B. (HO)2~ ~ -O(CH2)4CH3
To a cold (-76~C) solution of 6.~ g (24.6 mmol)
of the subtitled compound of Preparation 2A in 500 ml of
tetrahydrofuran, was added 21.5 ml of a 1.6~ solution of
sec-butyllithium ( 3 4.4 mmol) in hexane, via syringe. After
approximately twenty minutes, 12 ml (52 mmol) of
triisopropyl borate was added, via syringe. The resultant
reaction mixture was warmed to room temperature followed by
the addition of 60 ml of a lN hydrochloric acid solution.
After approximately ten minutes, the reaction mixture was
concentrated in vacuo to provide a pale yellow solid. This
solid was recrystallized from diethyl ether, ~ollowed by
recrystallization from hexanes to provide a white solid.
Yield: 2.95 g (57%).
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23
C. Br ~ ~ - O(CH2)4CH3
To a solution of 1.44 g (6.92 mmol) of a
subtitled compound of Preparation 2B in 50 ml of toluene
and 20 ml of methanol under nitrogen, was added lS.S ml
t31.1 mmol) a 2M sol. of sodium carbonate which resulted in
the formation o~ a white precipitate. To the resultant
mixture, was added 800 mg (0.6g mmol) of palladium tetrakis
(triphenylphosphine), followed by 1.64 g (6.92 mmol) of
2,5-dibromopyridine. The resultant reaction mixture was
reacted for approximately three hours, forty-five minu~es
at reflux temperature. When the reaction was substantially
complete, as inaicated by TLC. the mixture was cooled to
room temperature and allowed to stir overnight. The
reaction mixture was placed in a separatory funnel and
combined with diethyl ether and water. The resultant
layers were separated and the organic layer was dried over
sodium sulfate, filtered and then concentrated in vacuo to
provide a residue. This residue was redissolved in pentane
and filtered to provide 0.95 g of the desired compound.
The filtrate was recrystallized to provide an additional
1.4 g of the desired compound. These solids were combined
and used without further purification.
D. ~HO)~B ~ ~ _O(CH2)4CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Preparation 2B, using 15.8 ml of a 1.3~ sec-butyllithium
solution (20.54 mmol), 7 ml (30.33 mmol) of triisopropyl
borate, 125 ml of a 1~ hydrochloric acid solution and
450 ml of anhydrous tetrahydrofuran.
Yield: 4.0 g of an orange solid.
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24
PreDara~ion 3
A. 2-He~toxv-S-bromo-pvridine
A solution of 20.4 ml (0.048 mol) of heptanol in
50 ml of toluene was slowly added to a warm (50~C) slurry
of 5.76 g of 60% sodium hydride ~0.144 mol) in 400 ml of
dimethylformamide, under nitrogen, which resulted in the
evolution of hydrogen gas. After stirring the resultant
mixture at 80~C for approximately two hours, 30 g
(0.126 mol) of 2,5-dibromopyridine was slowly added. The
resultant reaction mixture was refluxed overnight. The new
mixture was poured into water. The desired compound was
extracted from the resultant mixture using diethyl ether
and the organic portions were then washed with water, dried
over magnesium sulfate, filtered and concentrated n vacuo
to provide 35 g of an oil. Purification with column
chromatography using silica gel ~eluent of 10% ethyl
acetate in hexane) to provide a clear oil.
Yield: 20.7 g (60%).
HPLC: C}g reverse-phase column; eluent of 20% water in
acetonitrile; ~=254 nm; 2 ml/min; RT=6 03 min;
B. (HO)2B ~ O(CH2)6CH3
The desired subtitled compound was prepared
substantially ln accordance with the procedure detailed in
Preparation 2B, using 2.72 g (10 mmol) of the subtitled
compound of Preparation 3A, 10 ml (16 mmol) of a 1.6~
solution of sec-butyllithium in hexane, 5.5 ml (24 mmol) of
triisopropyl borate, 50 ml of a 1~ hydrochloric acid
solution and 60 ml of diethyl ether.
Yield: 1.9& g of a white solid (83%).
HPLC: Clg reverse-phase column; eluent of 20% water in
acetonitrile; ~=254 nm; 2 ml/min; RT=2.~3 min;
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Pre~tion 4
A. Br ~ ~ - O(CH2)6CH3
A solution of 230 mg (0.25 mmol) of
tris(dibenzylideneacetone)dipalladium and 520 mg (2 mmol)
of triphenylphosphine in 5 ml of toluene was added to a
solution of 1.11 g (4.7 mmol) of 2,5-dibromopyridine,
1.13 g (4.7 mmol) of the subtitled compound of
Preparation 3B and 1.15 g (10.85 mmol) of sodium carbonate
in 40 ml o~ toluene, 30 ml of methanol and 13 ml of water,
under nitrogen. The resultant reaction mixture was reacted
for approximately two hours at reflux temperature. When
the reaction was substantially complete, as indicated by
HPLC, the reaction mixture was concentrated in vacuo to
provide a residue. This residue was dissolved in diethyl
ether and then washed sequentially with a lN hydrochloric
acid solution and a 2N sodium hydroxide solution, dried
over sodium sulfate, filtered and then reduced to dryness
in v~cuo to provide a solid. This solid was redissolved in
pentane and the resultant mixture was filtered. The
filtrate was cooled resulting in the formation of a white
solid which was collected by filtration.
Yield: 0.4865 g (30%).
m.p. 47-49~C.
MS(FD): 348 (M-1).
B. (HO)2B ~ ~ O(CH2)6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Preparation 2B, using 0.48 g (1.38 mmol) o~ the subtitled
compound of Preparation 4A, 1.4 ml of a 1.6M solution of
sec-butyllithium in hexane (2.2 mmol), 0.76 ml (3.3 mmol)
o~ triisopropyl borate, an excess of a 1 hydrochloric acid
solution and 50 ml of diethyl ether.
Yield: 0.439 g.
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26
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Pre~ration 5
A. Br ~ O(CH2)4CH3
A solution containing 50 g (200 mmol) of 4-
bromophenol, 33.5 g (298 mmol) of potassium t-butoxide and
40 ml (298 mmol) of 1-iodopentane in 1000 ml of
tetrahydrofuran was reacted at reflux temperature for
approximately twenty four hours. When the reaction was
substantially complete, as indicated by TLC the reaction
was filtered. The resultant filtrate was concentrated in
vacuo to provide a purple solid. This solid was
redissolved in a water/diethyl ether mixture to provide a
yellow solution. This solution was washed sequentially
with 200 ml of water (twice~, 100 ml of 2N sodium hydroxide
(twice) and 200 ml of brine (twice), dried over sodium
sulfate and then concentrated in vacuo to provide a yellow
powder. This solid was recrystallized from hot hexanes to
provide a white powder.
Yield: 45.8 mg (72%).
B.(~O)~-B ~ O-(CH2)4CH3
To a cold (-78~C) solution of 10.0 mg
(42.9 mmol) of 29 g (90.8 mmol) of the compound of
Preparation 5A, was added 91 ml of sec-butyllithium in
1000 ml of tetrahydrofuran (118 mmol), dropwise. To the
resulting mixture was added 41.9 ml (181.7 mmol) of
triisopropyl borate, dropwise. The resultant reaction
mixture was stirred for approximately thirty minutes and
then warmed to room temperature and allowed to react for
approximately two hours. The reaction was then quenched by
the addition of 1~ hydrochloric acid. The resultant
mixture was concentrated in vacuo to provide a residue.
This residue was redissolved in diethyl ether, filtered and
reduced to dryness to provide the desired subtitled
compound.
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Pre~aration 6
A. Br ~ Cl
N
A mixture of 8.14 g (46.5 mmol) of 2-hydroxy-5-
bromopyrimidine in 25 ml of phosphorus oxychloride
(268 Irrnol) was refluxed for 1.5 hours. After allowing the
reaction mixture to cool to room temperature, the excess
phosphorus oxychloride was removed ~y distillation. Ice
water was added to the residue, ~ollowed by sodium
hydroxide until pH 7. The aqueous layer was extracted
three times with ethyl acetate. The organic layer was
dried over sodium sulfate, filtered and then concentrated
in vacuo to provide 7.16 g of the desired compound.
Yield: 80%.
N
B. Br ~ ~_ o(cH2)6cH3
To a warm (93~) suspension of 2.11 g (52.8 mmol)
of 60% sodium hydride in 100 ml of toluene and 100 ml of
dimethylformamide, was added 7.3 ml (51.6 mmol) of
heptanol, dropwise, which resulted in the evolution o~
hydrogen gas. After stirring the resultant mixture at
115~C for approximately two hours, a solution of 4.g4 g
(25.6 mmol) of the subtitled compound of Preparation 6A was
added. After reacting overnight at reflux temperature, the
reaction mixture was cooled to room temperature and poured
over ice water. The desired compound was extracted from
the resultant mixture using diethyl ether. The resultant
mixture was washed three times with water, dried over
sodium sulfate, filtered and then concentrated in vacuo to
provide a dark brown oil which was purified using column~ 30 chromatography (gradient eluent of 5-10% ethyl acetate in
hexane) to provide 2.65 g of the desired compound.
~ Yield: 38%.
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28
N
C. (HO)2B ~ ~-o(cH2)6cH3
N
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Preparation 2B, using 2.77 g (10.1 mmol) of the subtitled
compound of Preparation 6B, 12 ml of a 1.3~ solution of
sec-butyllithium in hexane, 400 g or 21.3 mmol (15.6 mmol)
of triisopropyl borate, 16 ml of a 1~ hydrochloric acid
solution. The resultant layers were separated and the
organic layer was concentrated in vaCuo to provide 2. 83 g
of a crude material which was used without further
purification.
PreDar~tion 7
A. sr ~ o(cH2)6cH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Preparation 2A, using 51 g (0.29 moles) of 4-bromo-phenol,
49.4 g (0.44 moles) of 1-bromoheptane in 800 ml of
tetrahydrofuran to provide 77 g of an oil which was used
without further purification.
B. (Ho)2s ~ O(cH?)6cH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Preparation 2B, using 2.72 g (10 mmol) of the subtitled
compound of Preparation 7A, 10 ml (16 mmol) of a
1.6~ solution of sec-butyllithium in hexane, 5.5 ml
(24 mmol) of triisopropyl borate, 50 ml of a lN
hydrochloric acid solution and 60 ml of diethyl ether.
Yield: 1.98 g of a white solid (83~).
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C. Cl_~3~0(CH2)6CEI~
The desired subtitled compound was prepared
substantiallY in accordance with the procedure detailed in
~ Preparation 2C, using 1.22 g (5.16 mmol) of the subtitled
compound of Preparation 7B, 1.0 g (5.16 mmol) of the
subtitled compound of Preparation 6A, 598.3 mg (O.516 mmol)
of palladium tetrakis (triphenylphosphine), 11.6 ml of a
2M sodium carbonate solution (23 mmol), 25 ml of toluene
and 10 ml of methanol.
Yield: 1.4520 g (92%).
D (CH~)3sn ~ ~ ~ o(CH2)6CH3
To a solution of 613.0 mg (2.01 mmol) o~ the
subtitled compound of Preparation 7C and 119.3 mg
(Q.1 mmol) of palladium tetrakis(triphenylphosphine) in
10 ml of dioxane, was added 760 mg (2.32 mmol) of
hexamethylditin under nitrogen. The resultant reaction
mixture was reacted overnight at reflux temperature. After
cooling to room temperature, the reaction mixture was
concentrated in vacuo to provide a residue. This residue
was redissolved in 20 ml of diethyl ether, combined with a
saturated potassium ~luoride solution and stirred for
approximately 3.5 hours. The resultant layers were
separated and the organic layer was concentrated in vacuo
to provide the desired subtitled compound which was used
without further purification.
Pre~ara~ion 8
5-Bromofuranoic acid, methvl ester
To a slurry of 15.00 g (0.0785 mol) of
5-bromofuranoic acid in 500 ml of methylene chloride, was
added 12.73 g (0.0785 mol) of N,N-carbonyldiimidazole.
After reacting at room temperature overnight, 6.4 ml
(0.~57 mol) of methanol was added and the resultant mixture
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W 097/27864 PCTrUS97/01607
was stirred for approximately one hour and then washed
sequentially with a 0.5~ sodium hydroxide solution (twice)
and a 1~ hydrochloric acid solution. The organic portion
was dried over sodium sulfate, filtered and then
concentrated in vacuo to provide a residue. This residue
was slurried in pentane to provide a white solid.
Yield: 13.15 g (82%).
H NMR: ~ 7.17 (d, lH, CH); 6.45 (d, lH, CH);
3.89 (5, 3H, CH3).
Pre~aration 9
N-methanesulfo~ate benzotriazole
To a cold (5~C) solution of 100 g (0. 653 mol) of
hydroxybenzotriazole (HOBT) in 750 ml o~ methylene
chloride, was added 82.59 g (0.816 mol) of triethylamine
while maintaining the temperature at 5-10~C followed by the
addition of 82.28 g (0.718 mol) of methanesulfonyl chloride
while maintaining the temperature at 4-10~C. The resultant
reaction mixture was reacted for approximately one hour at
4~C. When the reaction was substantially complete, as
indicated by TLC, the reaction mixture was transferred to a
separatory ~unnel and washed sequentially with water=(three
times) and a saturated sodium chloride solution, dried over
sodium sulfate, filtered and concentrated in vacuo to
provide a solid. This solid was combined with a small
amount of diethyl ether and the resultant mixture was
filtered and dried in vacuo to provide a white solid.
Yield: 126.2 g (g1%).
Pre~aration 10
A . CH3 O- C~ OH
A solution of 50 g (0.26 mol) 4-cyano~
hydroxybiphenyl and an excess of 50% sodium hydroxide in
2000 ml of ethanol was refluxed for three hours. A~ter
cooling to room temperature, the reaction mixture was
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_
acidified with concentrated hydrochloric acid whlch
resulted in the formation of a solid which was collected by
filtration. The solid was suspended in 20 ml of
concentrated hydrochloric acid in methanol and refluxed
overnight. After cooling to room temperature, water was
added to the solution which resulted in the formation of a
solid. This solid was collected by filtration and dried
in vacuo overnight at 60~C.
Yield: 52.6 g (81%).
B. CH3O-C ~ oso2cF3
To a cold (Q~C) solution of 16.2 g (0.07 mol) of
4-hydroxy-4'-car~oxymethyl biphenyl in pyridine, was added
50 g (0.177 mol) of trifluoromethylsulfonic acid anhydride,
dropwise. The resultant reaction mixture was stirred for
1 hour, and then was concentrated in vacuo to provide a
residue. This residue was redissolved in diethyl ether,
washed with a lN hydrochloric acid solution, dried over
sodium sulfate, filtered, and then concentrated in vacuo to
provide an oil. This oil solidified when slurried in
pentane and the solid was collected by filtration.
Yield: 40.1 g (80%).
m.p. 57-58~C.
MS(FD~: 360.
Elemental Analysis:
Calcd: C, 50.00; H, 3.05;
Found: C, 50.30; H, 3.11.
Preparation 11
A. CH3O-c ~ Sn(CH3)~
To a solution of 6.05 g (23.1 mmol) of methyl
4-iodobenzoate and 1.6 g (1.38 mmol) of palladium
tetrakis(triphenyl)phosphine in 250 ml of dioxane, was
added 8.81 g (~7 mmol) of hexamethylditin, under nitrogen.
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The resultant reaction mixture was reacted ~or ~c71r hours
at reflux temperature. After cooling to room temp~r5ture,
the reaction mixture was concentrated i~ vacuo to provide a
residue. This residue was redissolved in diethyl ether and
a saturated potassium fluoride solution and stirred
overnight at room temperature. The resultant layers were
separated and the organic layer was concentrated in vacuo
to provide 7.1 g of a solid which was used without ~urther
purification.
1H NMR: (CDC13) ~ 7.95 (d, 2H, ArH); 7 .59 (d, 2H, ArH);
3.95 (S, 3H, CH3); 0.36 (S, 6H, CH3) .
B . CH } O--C~ ~ C 1
A solution containing 4.0 g (13 . 3 mmol) of the
subtitled compound of Preparation 11A, 2.5g (13. 3 mmo') of
the subtitled compound of Preparation 6A, 1g2 . 7 mg
(1.34 mmol) of copper~II) bromide, and 769.7 mg (0.66 mmol)
of palladium tetrakis(~riphenylphosphine) in 80 ml of
dimethylformamide, under nitrogen, was refluxed for 2 . 5
hours. After cooling to room temperature, the reaction
mixture was concentrated in vacuo to provide a residue.
This residue was redissolved in a mixture of diethyl ether
and a saturated potassium fluoride solution and then
stirred at room temperatue for approximately forty eight
hours. The resultant layers were separated and the organic
layer was concentrated in vacuo to provide 1.88 g of crude
material which was purified using column chromatography
(eluent of 1% ethyl acetate in methylene chloride).
Yield: 0.47 g (14%).
MS~FAB) for C56H72~9~17 (M+1):
Calcd: 1142.5046
~ound: 1142.5085.
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33
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Pre~ar~tion 12
A. 2-~thoxvmethvl-1-~ro~enoic acid. ethvl ester
To a cold (0~C) gel-like mixture of 25 ml
(0.162 mmol) of ethoxypropanoic acid, ethyl ester and
40 ml (0.495 mol) of formic acid, ethyl ester, was
added 35 ml (0.37 mol) of dimethylsulfate, dropwise.
The resultant reaction mixture was warmed to room
temperature and then heated to 60~C and allowed to
react overnight. After adding more dimethylsulfate
(6 ml), the reaction mixture was hea~ed to ~0~C and
stirred for five hours. After cooling to room
temperature, an aqueous solution of 2~ sodium
carbonate was added to the reaction mixture to kring
the mixture to pH 12. The resultant layers were
separated and the organic layer was dried over sodium
sulfate, filtered and concentrated in vacuo to provide
the subtitled compound.
Yield: 8.56 g (28~).
B. 2-Oxo-5-ethoxvc~rhonvl-l~ 3,6-trihvdro~vrimi~ine
To a mixture of 1.0044 g (5.34 mmol) of the
subtitled compound of Preparation 12A in ethanol, was
added 321.2 mg (5.35 mmol) of urea, followed by
approximately 0.52 ~1 of concentrated hydrochloric
acid. The resultant reaction mixture was refluxed for
approximately 8.25 hours and then stirred overnight at
room temperature. The reaction mixture was
concentrated in vacuo to provide a white solid. This
solid was recystallized in ethanol to provide 321 mg
of the desired compound.
m.p. 167-172~C.
MS(FD): 171.
C. 2-oxo-5-ethoxvcarhonvl-3-monohvdro~yrimi~;ne
hvdrobro~ide
A solution of 3.31 g (20.7 mmol) of bromine
in 14 ml of glacial acetic acid was added to a mixture
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34
of 3.52 g (20.7 mmol) of the subtitled compound of
Preparation 12B in 71 ml of glacial acetic acid. The
resultant reaction mixture was reacted at re~lux
temperature. The resultant crude material was used
without ~urther puri~ication.
Yield: 3.9773 g (77%~.
m.p. 198-200~C (decomp.).
D. 2-Chloro-5-ethoxvcarbonvl-~vrimidine
To 3.659 g (14.7 mmol) of the subtitled
compound of Preparation 12C, was added 20.5 ml
(220 mmol) of phosphorus oxychloride, ~ollowed by 2 ml
(26.6 mmol) of dimethylphenylamine. The resultant
reaction mixture was slowly added to cold water. The
resultant layers were separated and the aqueous layer
was neutralized by the addition of a 5N sodium
hydroxide solution and then the desired compound was
extracted using ethyl acetate. The extracts were
combined and then concentrated in vacuo to provide a
residue. This residue was slurried in hot hexanes and
the resultant mixture was cooled to room temperature
and filtered. The filtrate was concentrated in vacuo
to provide a yellow solid.
Yield: 0.9~96 g (33%).
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x~mnle 1
A. CH30-C ~ o(cH2)4cH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Preparation 2C, using 1.99 g ~6.98 mmol) of the subtitled
compound of Preparation 2D, 1.25 g (7.28 mmol) of the
titled compound of Preparation 1, 0.811 g (0. 73 mmol) o~
palladium tetrakis (triphenylphosphine), 15.8 ml
(31.6 mmol) of a 2N sodium carbonate solution, 50 ml of
toluene and 20 ml of methanol.
Yield: 1. 3381 g t51%).
o
B. HO-C ~ O(CH~)4CH3
A mixture of 1.336 g (3. 55 mmol) of the
subtitled compound of Example lA and 9. 3 ml of a 2N sodium
hydroxide solution (18.6 mmol) in 250 ml of dioxane was
refluxed overnight. After cooling the reaction mixture to
room temperature, 18.6 ml of a 1~ hydrochloric acid
solution was added, which resulted in the formation of a
precipitate. This precipitate was isolated by filtration to
provide 1.044 g of material that was used without further
purification.
Yield: 81%.
N" 'N-O-C ~ O(CH2)4CH3
C.
To a mixture of 1.02 g t2.81 mmol) of the
subtitled compound of Example lB in 95 ml of
dimethylformamide, was added 630.6 mg (2.96 mmol) of the
titled compound of Preparation 9, followed by O.g22 ml
t3.035 mmol) of triethylamine. After reacting at room
temperature for approximately 2.5 hours, the reaction
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36
mixture was concentrated in vacuo to provide a black
residue. This residue was redissolved in methylene
chloride, washed with water, filtered, dried over sodium
sulfate, filtered and then concentrated in vacuo to provide
1.34 g of solid which was used without further
purification.
D. Pre~aration of the compound of formula I where R',
R" ~nd R"' are e~ch methvl and RX~ X2 RVl Rv2 ~v3
RV4 and RQ are each hvdroxv and R2 is
- C ~ O(CH~)4CH3
To a solution of 1. 34 g (2.79 mmol) of the
subtitled _ompound of Example lC in dimethylformamide, was
added 2.13 g (2.61 mmol) of the A-30912A nucleus (compound
of formula lB where R', R" and R"' are each methyl, R
Rx2, Ryl, Ry2, RY3, RY4 and Ro are each hydroxy). After
reacting at room temperature overnight, under nitrogen, the
reaction mixture was filtered. The resultant filtrate was
concentrated in vacuo to provide a residue which was
slurried in diethyl ether, filtered and then slurried in
methylene chloride and filtered to provide a gold powder.
This powder was redissolved in methanol and then puri~ied
using HPLC ieluent of 35% acetonitrile, 55% water and 10%
of a 1% a~ueous trifluoroacetic acid solution). The
fractions containing the desired compound were combined and
concentrated in vacuo to provide 1.20 g (39%) of the
desired compound.
MS(FAB) fo~ C56~72N9~l7 (M+H):
Calcd: 1142.5046;
round 1142.5085-
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37
_
~.x~mnl e 2
A. CH3O- C ~ O(CH2)4CH3
The desired subtitled compound was prepared
- substantially in accordance with the procedure detailed in
Preparation 2C, using 2.00 g (7.01 mmol) of the subtitled
compound of Preparation 2D, 2.02 g (7.71 mmol) of methyl-~-
iodobenzoate, 830 mg (0.70 mmol) of palladium tetrakis
(triphenylphosphine), 16 ml (31. 5 mmol) of 2M sodium
carbonate, 50 ml of toluene and 20 ml of methanol to
provide an off-white powder which was used without
purification.
Yield: 2.28 g (90%).
B. HO-C ~ O(CH2)4CH.
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lB, using 2.28 g ( 6.3 mmol) of the subtitled
compound of Example 2A, 16. 6 ml of a 21~[ sodium hydroxide
solution (33.2 mmol), 500 ml of dioxane and 33. 2 ml of a
lN hydrochloric acid solution.
Yield: 1.55 g (71%).
o
N N - O - C ~ O(CH2)4CH3
C.
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lC, using 1.55 g (4.46 mmol) of the subtitled
compound of Example 2B, 1.02 g (4.78 mmol) of the titled
compound of Preparation 9, 0.67 ml (4.82 mmol) of
triethvlamine and 125 ml of dimethylformamide to provide
0.320 g of a tan powder.
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38
-
D. Pre~aration of the com~ound of formula I where R~'. R"
~nd R"' are each methvl an~ RXl~ R~. RY~,_gY~ 3, ~v~_~n~
RQ are e~ch hvdroxv and R2 s
- C ~ O(CH2)4CH~
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lD, using 0.320 g (0.668 mmol) of the subtitled
compound of Example 2C, 486.4 mg (0.609 mmol) of the
A-30912A nucleus in dimethylformamide.
Yield: 0.2832 (41%)
MS (FAB) for Cs7H72N8ol7Li:
Calcd: 1147.5205;
Found: 1147.5175.
~m~le 3
A. CH30-C ~ o(cH2)6cH3
A solution of 230 mg (0.25 mmol) of
tris(dibenzylidene acetone) dipalladium (0) (Pd2dba3) and
520 mg (2 mmol) of triphenylphosphine in 10 ml of toluene
was added to a solution containing 1.4 g (3.9 mmol) of
1-methoxycarbonyl-4-(4'-trifluorosulfonate)biphenyl, 1.13 g
(4.7 mmol) of the subtitled compound of Preparation 3B,
1.15 g (10.85 mmol) of sodium carbonate in 30 ml of
toluene, 20 ml of methanol and 13 ml of water. The
resultant reaction mixture was refluxed overnight, cooled
and the resultant layers were separated The organic layer
was concentrated in vacuo to provide a residue. This
residue was slurried in methanol and then filtered to
provide 0.797 g of a solid which was determined to be 98%
pure using HPLC (eluent of 90~ acetonitrile in water;
~=254; 3 ml/min.i R~ = 3.76 min.).
Yield: 51~.
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.
O
B. HO-C ~ O(CH2)6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lB, using 0.797 g (1.9 mmol~ of the subtitled
compound of Example 3A, 5 ml of a 2N sodium hydroxide
solution (10 mmol), 200 ml of dioxane and 10 ml of a
l~ hydrochloric acid solution.
Yield: 518 mg (70%).
N~ N-O-C ~ O(CH~)6CH3
C. ~ .
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lC, using 0.512 g (1.33 mmol) of the subtitled
compound of Example 3B, 298 mg (1.4 mmol~ of the titled
compound of Preparation 9, 0.195 ml (1.4 mmol) of
triethylamine and 50 ml of dimethylformamide to provide
145 mg of solid which was determined to be 95~ pure using
HPLC (eluent of 90 acetonitrile in water, 3 ml/min.,
~=280 nm, R~=3.88 min.)
D. PreDaration of the com~ollnd of formula I where R',
R" and R"' are each methvl and R ~ R ~ , R~;L, R ~ RV3
RV4 an~ R0 are each hvdroxv and R~ ;s
--C~ O (CH2 ) 6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example 1 D, using 225 mg (0.445 mmol) of the subtitled
compound of Example 3C, 355 mg (0.445 mmol) of the A-30912A
nucleus in dimethylformamide to provide 314 mg of a white
solid which was determined to be 99.6% pure using HPLC
~eluent of 55~ acetonitrile in water containing
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- 40
O.5% monoammonium phosphoric acid, ~=230 nm, 2 ml/min.,
RT=4-35 min.).
MS tFAB) for C59H77N8~17:
Calcd: 1169.5407;
Found: 1169.5391.
~xam~]e 4
A. CH30-C ~ O(CH2)6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example 3A, using 47 mg (0.051 mmol) of Pd2dba3~ 106 mg
(0.41 mmol) of triphenylphosphine, 253 mg (0.97 mmol) of
methyl-4-iodobenzoate, 303 mg (0.97 mmol) of the subtitled
compound of Preparation 4B, 0.46 g (4.4 mmol) of sodium
carbonate in 8 ml of toluene, 6 ml of methanol and 2.6 ml of
water to provide 11 mg of a solid which was determined to be
90% pure using HPLC ~eluent of 90% acetonitrile in water,
2 ml/min., ~=280 nm, RT=5.15 min.).
B. HO-C ~ O(CH2)6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lB, using 0.458 g (1.13 mmol) of the subtitled
compound of Example 4A, 5 ml of a 2~ sodium hydroxide
solution (10 mmol), 60 ml of dioxane and 10 ml of a
1~ hydrochloric acid solution to provide 0.51 g of material
that was used without further purification.
N" 'N-O-C ~ O(CH2)6CH3
C.
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
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41
Example lC, using 0.51 g of the subtitled compound of
Example 4B, 298 mg (1.4 mmol) of the titled compound of
Preparation 9, 0.195 ml (1.4 mmol) of triethylamine and
50 ml of dimethylformamide to provide 234 mg o~ a white
solid which was determined to be 98% pure using HPLC
~eluent of 90 acetonitrile in water; 3 ml/min.; ~280 nm;
RT=3.43 min.).
D. Pre~aration of the com~ound of formul~ I where
R', R" and R"'are each methvl and R~1, RX2~ RVl~ RY2 RV3
R~ and RQ are each hvdroxv and R~
--C~ o ( CEI2 ) 6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lD, using 230 mg (0.45 mmol) of the subtitled
compound of Example 4C, 359 mg (0.45 mmol) of the A-30912A
nucleus in 15 ml of dimethylformamide to provide 371 mg of
a solid which was determined to be 94% pure using HPLC
(eluent of 50% acetonitrile in water containing
0.5% monoammonium phosphoric acid; ~230 nm; 2 ml/min.;
RT=3.91 min.).
MS(FAB) for C58H75N9~17Li:
Calcd: 1176.5441;
Found: 1176.5476.
2~
~mI-le 5
u ~N fiN ~=N
~. CH30--C~ O(CH~)6CH3
The desired su3~titled compound was prepared
substantially in accordance with the procedure detailed in
Example 3A, using 131 mg (0.14 mmol) of Pd2dba3, 299 mg
(1.14 mmol) of triphenylphosphine, 463 mg (2.7 mmol) of the
titled compound of Preparation l, 1 g (3.18 mmol) of the
subtitled compound of Preparation 4B, 1.3 g (12.2 mmol) of
sodium carbonate in 22 ml of toluene, 16 ml of methanol and
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42
-
7 ml of water to provide 206.9 mg of a solid which was
determined to be 96% pure using HPLC (eluent of
90% acetonitrile in water, 3 ml/min., ;~=280 nm,
RT=3.13 min.).
Il N N N
B. HO-C ~ O(CH~)6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lB, using 200 mg (0.49 mmol) of the subtitled
compound of Example 5A, 3 ml o~ a 2~ sodium hydroxide
solution (6 mmol), 15 ml of dioxane and 6 ml o~ a lN
hydrochloric acid solu~ion to provide 153 mg of material
which was used without further purification.
Yield: 80%.
N" 'N-O-C ~ O(CH2)6CH3
C.
A solution containing 153 mg (0. 39 mmol) of the
subtitled compound of Example 5B, 54 mg (0.4 mmol) of the
titled compound of Preparation 9, and 83 mg (0.4 mmol) o~
dicyclohexvlcarbodiimide (DCC) in 25 ml of methylene
chloride was stirred overnight. The resultant reaction
mixture was filtered and then concentrated in vacuo to
provide a solid. This solid was slurried in diethyl ether
and then filtered to provide 161.3 mg of a solid.
D. Pre~aration of the com~ound of formula I where R',
R' ~nd R"' are each methvl and R~;L F?~2 R~;L, R~. RY~
RV4 and RQ are each hvdroxy and R2_
- C ~ ~ -o(CH~)6cH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lD, using 161 mg (0.24 mmol) of the subtitled
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g3
compound of Example 5C, 190 mg (0.24 mmol) of the A-30912A
nucleus in dimethylformamide to provide 102 mg of a solid
which was determined to be 96~ pure using HPLC (eluent o~
50% acetonitrile in water containing 0. 5% monoammonium
phosphoric acid; ~ 230 nm; 2 ml/min.; RT=3. 03 min.).
MS(FAB) ~or C57H74N10~17Li:
Calcd: 1177. 5393;
Found: 1177.5350.
Exam~le 6
A. CH30-C ~ 0(CH2)4CH3
The desired subtitled compound was prepared
su~stantially in accordance with the procedure detailed in
Example 2C, using 2.00 g (7.03 mmol) o~ the subtitled
compound o~ Preparation 5B, 1.51 g (8.80 mmol) of the titled
compound of Preparation 1, 0.81 g (0.7 mmol) of palladium
tetrakis (triphenylphosphine), 15.8 ml of a 2~ solution of
sodium carbonate ~31.6 mmol), 50 ml of toluene and 20 ml of
methanol to provide 0.8856 g of a solid.
Yield: 30%.
B. Ho-C ~ o(CH2)4CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example 1 s, using 0.8586 g (2.286 mmol) of the subtitled
compound of Example 6A, 6 ml of a 2~ sodium hydroxide
solution (12 mmol), 228 ml of dioxane and 12 ml of a lN
hydrochloric acid solution to provide 0.65 g of material
that was used without further purification.
Yield: 78%.
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44
~N~N 11 ~ 0tCH2)4CH3
The desired subtitled compound was prepared
substantially in accQrdance with the procedure detailed in
Example lC, using 0.65 g (1.8 mmol) of the subtitled
compound of Example 6B, 405 mg (l.gO mmol) of the titled
compound of Preparation 9, 0.27 ml (1.94 mmol) of
triethylamine and 60 ml of dimethylformamide.
Yield: 624.4 mg (7396).
D. Pre~aration of the com~ound of formula ~ here
R', R" and R"' are e~ch methYl and RXl R~. R'", ~-~2
R '-, RV4 and RQ are e~ch hydro~ ~nd R~ s
--C ~ O ( CH2 ) q CH~
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lD, using 587.7 mg (1.229 mmol) o~ the subtitled
compound of Example 6C, 893 mg (1.119 mmol) of the A-30912A
nucleus in 30 ml of dimethylformamide.
Yield: 542. a mg (38%).
MS(FAB~ ~or C57H71N8016~ H2O)
Calcd: 1123.4988:
Found: 1123.5024.
F.~am7~1e 7
~. C~3O- C ~ f>-- O(CH2)6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example 3A, using 1.415 g (5.95 mmol) o~ the sul~titled
compound of ?reparation 6C, 2.75 g (7.63 mmol) of
1-methoxycar2Onyl-4-(4'-trifluorosulfonate)phenyl, 863.3 mg
(0.75 mmol) _ palladium tetrakis(triphenylphosphine), 13 ml
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_ 45
of a 2~ solution of sodium carbonate (26 mmol), 50 ml of
toluene and 20 ml of methanol to provide 0 .54~;3 g o:E a solid
which was 94% pure by HPLC (20% water in acetonitrile;
~=280 nm; 3 ml/min.; RT=5 . 61 min).
Yield: 13%.
g HO--C ~ N~-O(CH2)6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lB, using 0.518 g (1.28 mmol) of the subtitled
compound of Example 7A, 3.2 ml of a 2~ sodium hydroxide
solution (6.4 mmol), dioxane and 6.4 ml of a 1~ hydrochloric
acid solution to provide 458.9 mg of material which was used
without further purification.
~ ' O C~ ~>--O (CH2 ) 6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lC, using 439.0 mg (1.12 mmol) of the subtitled
compound of Example 7B, 251.5 mg (1.18 mmol) of the titled
compound of Preparation 9, 0.165 ml (1.18 mmol) of
triethylamine and 30 ml of dimethylformamide to provide
309 mg of a solid.
MS(~D): 507.2 (M).
D. Pre~aration of the com~ound of formula I where R',
R" and R"' ~re each methvl and R ~ R2~2. R~;L, RV;2-, ~ 3, R
and R~ are each hvdroxY and R-- iS
~ O (CH2) 6CH3
~he desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
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46
-
Example lD, using 300 mg (0.591 mmol) of the s~lbtitled
compound of Example 7C, 430.1 mg (0.539 mmol) of the
A-30912A nucleus in 15 ml of dimethylformamide to provide
0.3263 g of a white powder which was 98% pure by HPLC (50%
acetonitrile in water; ~=230 nm; 2 ml/min; ~T=3.22 min).
MS(FAB) for C58H74N9~16:
Calcd: 1152.5254:
Found: 1152.5247.
F~,~mnle 8
A. CH30-C ~ N ~ O(CH2)6CH3
To a solution containing 4.43 g (10.23 mmol) of
the subtitled compound of Preparation 7D and 3.22 g
(12.28 mmol) of methyl-4-iodobenzoate in 55 ml of
1,2-dichloroethane, was added 201 mg (0.31 mmol) of
bis(triphenylphosphine)palladium (II) chloride, under
nitrogen. The resultant reaction mixture was reacted at
reflux temperature for approximately forty eight hours.
After cooling to-room temperature, the reaction mixture was
concentrated i~ vacuo to provide a residue which was
slurried in acetonitrile and ~iltered to provide 1.7135 g
of an orangebrown solid.
Yield: 41%.
MS(FD): 40~ (M).
B. HO-C ~ N ~ O(CH~)6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lB, using 1.58 g (3.91 mmol) of the subtitled
compound of Example 8A, 10.4 ml of a 2N sodium hydroxide
solution (20.8 mmol), 250 ml of dioxane and 20.8 ml of a
lN hydrochloric acid solution to provide 1.4133 g of
material which was used without further purification.
MS(FD). 390.2.
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--
'N' O C ~ N ~ O(CH2)~CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lC, using 352.1 mg (0.902 mmol) of the subtitled
compound of Example 3B, 202.2 mg (0.948 mmol) of the ti~led
compound of Preparation 9, 0.13 ml (0.937 mmol) of
triethylamine and 30 ml of dimethylformamide to provide
0.294 g of a solid which was used without further
purification.
D. PreDar~ion of the com~ound of formula I where R',
R" ~nd R" are each methvl and ~1, R~2, R~l, RY2 ~Y_ Rv4
and R0 are e~ch hvdroxv and R2
-C ~ ~ ~ O(CH2)6CH3
The desired subtitled compound was prepared
su~stantially in accordance with the procedure detailed in
Example lD, using 0.2881 g (0.5675 mmol) of the subtitled
compound of Example 8C, 0.4116 g (0.516 mmol) of the
A-30912~ nucleus in 14 ml of dimethylformamide.
Yield: 338.7 mg (51~).
MS(FAB) for Csg~74NgO16:
Calcd: 1152.5254:
Found: 1152.5247.
~x~m~le 9
A. CH30-C ~ \ ~ - O(CH2)4CH3
The desired su~titled compound was prepared
substantially in accordance with the procedure detailed in
Example 2C, using 2.01 g (7.03 mmol~ of the subtitled
compound of Preparation 5B, 1.51 g (7.3 mmol) of the titled
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48
compound of Preparation 8, 816 mg of palladium tetrakis
(triphenylphosphine), 2N sodium carbonate, toluene and
methanol.
Yield: 1.9961 g (78%).
HP~C: eluent of 80% acetonitrile in water, 2 ml/min;
~=280 nm; RT=4.64 min.
o
B. HO~ O(cH2)4cHl
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lB, using 1.996 g (5. 47 mmol) of the subtitied
compound of Example 9A, 13. 7 ml of a 2N sodium hydroxide
solution (27.4 mmol), 200 ml of dioxane and 13.7 ml of a
lN hydrochloric acid solution to provide 2.14 g of a solid.
MS(FD): 351.
N N--O--C ~ 6 ~ O (CH2)4CH3
C.
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lC, using 0.514 mg (1. 47 mmol) of the subtitled
compound of Example 9B, 0.393 g (1.84 mmol) of the titled
compound of Preparation 9, 0.224 ml (1.61 mmol) of
triethylamine in dimethylformamide.
Yield: 0.377 g (55%)-
MS(FD): ~67.
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--
D. PreDaration of the compound of formula I where R',
R" and R"'are e~ch methyl and RXl, RX2~ RV~ Rv3 Rv4
and RQ are each hvdroxv and R2 is
-C ~ ~ _ ~ -O(CH2)4CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lD, using 355 mg (0.76 mmol) of the subtitled
compound of Example 9C, 550.4 mg (0.69 mmol) of the
A-30912A nucleus in 20 ml of dimethylformamide to provide
223.1 mg of a solid.
MS (FAB) for C56H70N7~17:
Calcd: 1112.4828;
Found: 1112.~847.
F.7~mnle 10
A. CH3CH2O -C~ O(CH2)4CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Preparation 2C, using 1.50 g (5.28 mmol) of the subtitled
compound of Preparation 5B, 1.06 g (5.56 mmol) of
2-ethoxycarbonyl-5-chloro-thiophene, 0.62 g (0.536 mmoi) of
palladium tetrakis(triphenylphosphine), 48 ml of a 2~
solution of sodium carbonate (96 mmol), 39 ml of toluene
and 16 ml of methanol to provide 1.6088 g of a solid which
was used without further purification.
o
B. HO-C~ o(c~2)4cH3
The desired subtitled compound was prepared
sukstantially in accordance with the procedure detailed in
Example lB, using 1.609 g (4.08 mmol) of the subtitled
compound of Example lOA, 10.5 ml of a 2N sodium hydroxide
solution (21 mmol), 100 ml of dioxane and 21 ml of a lN
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hydrochloric acid solution to provide 1.4935 ~ of a white
solid which was used without purification.
MS(FD): 366.
o
N' N-O-C ~ ~ - ~ - O(CH2)4CH3
C. ~
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lC, using 1.47 g (4.02 mmol) of the subtitled
compound of Example lOB, 907 mg (4.25 mmol) of the titled
compound of Preparation 9, 0.6 ml (4.37 mmol) of
triethylamine in dimethylformamide to provide 1.0498 g of a
yellow powder which was used without puri~ication.
D. PreDaration of the comnollnd of formula I where R',
R" ~nd R"' ~re e~ch methvl ~nd RXl R~,B ~l BV2 RY3 RY4
and R0 are each hvdroxv and R2 is
- C ~ ~ _ ~ -O(CH2)4CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lD, using 1.043 g (2.15 mmol) of the subtitled
compound of Example lOC, 1.56 g (1.96 mmol) of the A-30912A
nucleus in dimethylformamide.
Yield: 2.4611 g.
MS(FAB) for Cs6H7oN7ol6s:
Calcd: 1128.4600;
Found: 1128.4626.
~xam-~le 11
A. CH-~O ~ ~ N ~ - O(CH2)6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
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Preparation 2C, using 150.6 mg (0.64 mmol) of the subtitled
compound of Preparation 7B, 194.6 mg (0.78 mmol) of the
subitled compound of Preparation llB, 73.4 mg (O.06 mmol)
of palladium tetrakis(triphenylphosphine), 1.5 ml of a 2DI
solution of sodium carbonate (3 mmol), 5 ml of toluene, and
2 ml of methanol to provide 190 mg of a light-brown solid
which was used without further purification.
MS(FD: ~04.1
o
B. HO C ~ ~ N ~ - O(CH2)6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lB, using 190 mg (0.47 mmol) of the subtitled
compound of Example 11A, 2.5 ml of a lN sodium hydroxide
solution (2.5 mmol), 33 ml of dioxane, and 2.5 ml ol a lN
hydrochloride acid solution to provide 165.3 mg of crude
material which was used without further purification.
MS(FD): 390.
N N-0-C ~ ~ ,>- ~ -O(CH~)6CH~
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lC, using 150.4 mg (0.305 mmol) of the subtitled
compound of Example llB, 86.7 mg (0.407 mmol) of the titled
compound of Preparation 9, 57~1 (0.415 mmol) of
triethylamine in dimethylformamide to provide 73.3 mg of a
light-brown solid which was used without further
purification.
-
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D. PreDaration of the compound of formula I where R',
R" and R"' ~re each methvl and RX1, R~ RYl, Rv2 ~ 3 Rv4
and R0 ~re e~ch hvdroxv and R~ is
D. - C- ~ _ C ~>_ ~ -O(CH2)6CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lD, using 60.1 mg (0.118 mmol) of the subtitled
compound of Example llC, 86.5 mg (0.108 mmol) of the
A-30912A nucleus in 3.5 ml of dimethylformamide to provide
54.1 mg of a white powder.
MS(FAB) for CsgH74N9ol6:
Calcd: 1152.5254;
Found: 1152.5236.
~ r~le 12
A. CH3CH2~ -C ~ N ~ O(CH2)4CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example 3A, using 1.04 g (3.66 mmol) of the subtitled
compound of Preparation 5B, 0.75 g (4.02 mmol) of the
subtitled compound of Preparation 12D, 0.375 g (0.32 mmol)
of palladium tetrakis(triphenylphosphine), 8 ml of a 2M
solution of sodium carbonate (16 mmol), 26 ml of toluene and
10 ml of methanol.
Yield: 0. 6678 g (47%).
B. HO-C ~ ~ ~ O(CH2)4CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lB, using 559.3 mg (1.432 mmol) of the subtitled
compound of Example 12A, 7. 2 ml of 2~ sodium hydroxide
(7.2 mmol), 90 ml of dioxane and lII hydrochloric acid.
Yield: 167.1 mg (92%).
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C. N ~ N-O-C ~ ~ O(CH2)4CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lC, using 154.5 mg (0.426 mol) of the subtitled
compound of Example 12B, 97.6 mg (0.458 ITunol) of~ the titled
compound of Preparation 9, 0.06 ml (0. 431 mmol) of
triethylamine and 14 ml of dimethylformamide.
~ield: 100 mg (49%).
D. Pre~aration of the com~ound of formula I where R',
R" and R"' are each methvl and R~1 R~2 Rvl Rv2 RV3 Rvg
and RQ are each hvdroxv and R2 is
- C ~ ~ O(CH2)4CH3
The desired subtitled compound was prepared
substantially in accordance with the procedure detailed in
Example lD, using 100 mg (0.208 mmol) of the subtitled
compound of Example 12C, 158 mg (0.198 mmol) of the
A-30912A nucleus in dimethylformamide to provide 11.8 mg of
the desired compound.
MS ~FAB) for Cs6H70N9ol6 ~M-H2O):
Calcd: 1124.4941:
Found: 1124.4919.
The compounds of formula I exhibit antifungal
and antiparasitic activity. For example, the compounds of
formula I inhibit the growth of various infectious fungi
including Candida spp. such as C. albicans,
C. parapsilosis, C. krusei, C. glabrata, or C. tropicalis,
C. lusita~iaei Torulopus spp. such as T. glabrata;
Aspergillus spp. such as A. fumigatus; Histoplasma spp.
such as H. capsulatum; Cryptococcus spp. such as
C. neoformans; Blastomyces spp. such as B. dermatitidis;
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Fusarium spp., Trichophyton spp., Pseudallescheria boydii,
Coccidioides immitis, Sporothrix schenckii and the like.
Antifungal activity of a test compound was
determined in vitro by obtaining the m;n;ml~m inhibitory
concentration (MIC) of the compound using a standard agar
dilution test or a disc-diffusion test. The compound was
then tested in vivo ( in mice) to determine the e~fective
dose of the test compound for controlling a systemic fungal
infection.
Accordingly, the following compounds were tested
for antifungal activity against C. albicans.
Table 1
~inimal inhibitorv concentration a~ainst C. albicans
~ m~le No. MIC (~ml)
lD 0.78
2D 0.039
3D 0.02
4D 0.002
5D 0.01
6D 0.02
7D 0.01
8D 0 0O
9D 0.312
lOD 0.078
llD 0.039
12D 0.02
In addition, the effective dose of the following
compounds for controlling a systemic fungal infection
(C. albicans) was tested in vivo (mice).
,
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T~hle 2
-~250 !mouse, i.~.)
~xamDl e No . Fns o ( ma / kçr )
lD 1.57
2D l.25
3D 0.31
4D 0.4l
5D l.13
6D 0.31
7D 0.47
8D 0.31
9D >2.5
l0D 5.52
llD 0.38
The compounds of the invention also inhibit the
growth of certain organisms primarily responsible for
opportunistic infections in immunosuppressed individuals.
For example the compounds of the invention inhibit the
growth of Pneumocystis carinii the causative organism of
pneumocystis pneumonia (PCP) in AIDS and other
;mm~nocompromised patients. Other protozoans that are
inhibited by compounds of formula I include Plasmodium
spp ., Lei .c n m~ n i a spp ., T~panos orna spp ., Cryp ~ ospori d i um
spp., Isospora spp. , Cycl ospora spp., Trichomonas spp .,
Microsporidiosis spp. and the like.
The compounds of formula I are active in vitro
and in vivo and are useful in combating either systemic
fungal infections or fungal skin infections. Accordingly,
the present invention provides a method of inhibiting
fungal activity comprising contacting a compound of formula
I, or a pharmaceu~ically acceptable salt thereof, with a
fungus. A preferred method includes inhibiting Candi da
al~icans or ~sper~illus fumigatis activity. The present
invention further provides a method of treating a fungal
infection ~.-hich comprises administering an effective amount
of a compound of formula I, or a pharmaceutically
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acceptable salt thereof, to a host in need of such
treatment. A preferred method includes treating a C~ndida
albicans or Aspergillus fumigatis infection.
With respect to antifungal activity, the term
~effective amount,~ means an amount of a compound of the
present invention which is capable of inhibiting fungal
activity. The dose administered will vary depending on
such factors as the nature and severity of the infection,
the age and general health of the host and the tolerance of
the host to the antifungal agent. The particular dose
regimen likewise may vary according to such factors and may
be given in a sinyle daily dose or in multiple doses during
the day. The regimen may last from about ~-3 days to about
2-3 weeks or longer. A typical daily dose (administered in
single or divided doses) will contain a dosage level of
from about 0.01 mg/kg to about lO0 mg/kg of body weight of
an active compound of this invention. Preferred daily
doses generally will be from about 0.1 mg~kg to about
60mg/kg and ideally from about 2.5 mg/kg to about 40 mg/kg.
The present invention also provides
pharmaceutical formulations useful for administering the
antifungal compounds of the invention. Accordingly, the
presen~ invention also provides a pharmaceutical
formulation comprising one or more pharmaceutically
acceptable carriers, diluents or excipients and a compound
of claim 1. The active ingredient in such formulations
comprises from 0.1% to 99.9% by weight of the ~ormulation,
more generally from about 1~% to about 30% by weight. B~
~pharmaceutically acceptable~ it is meant that the carrier,
diluent or excipient is compatible with the other
ingreaients of the formulation and not deleterious to the
recipient thereof.
A compound of formula I may be administered
paren-erally, for example using intramuscular, sub-
cutaneous, or intra-peritoneal injec~ion, nasal, or oral
means. In addition to these methods of administration, a
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compound of formula I may be applied topically for skin
infections.
For parenteral administration the formulation
comprises a compound of formula I and a p~ysiologically
acceptable diluent such as deionized water, physiological
saline, 5% dextrose and other commonly used diluents. The
~ormulation may contain a solubilizing agent such as a
polyethylene glycol or polypropylene glycol or other known
solubilizing agent. Such ~ormulations may be made up in
sterile vials containing the antifungal and excipient in a
dry powder or lyophilized powder form. Prior to use, a
physiologically acceptable diluent is added and the
so~ution withdrawn via syringe for administration to the
patient.
The present pharmaceutical f ormulations are
prepared by known procedures using known and readily
available ingredients. In making the compositions o~ the
present invention, the active ingredient will generally be
admixed with a carrier, or diluted by a carrier, or
enclosed within a carrier which may be in the ~orm of a
capsule, sachet, paper or other container. When the
carrier serves as a diluent, it may be a solid, semi-solid
or liquid material which acts as a vehicle, excipient or
medium for the active ingredient. Thus, the compositions
can be in the form of tablets, pills, powders, lozenges,
sachets, cachets, elixirs, suspensions, emulsions,
solutions, syrups, aerosols, (as a solid or in a li~uid
medium), ointments containing, for example, up to 10% by
weight of the active compound, soft and hard gelatin
capsules, suppositories, sterile injectable solutions,
sterile packaged powders and the like.
For oral administration, the antifungal compound
is fllled into gelatin capsules or formed into tablets.
Such tablets may also contain a binding agent, a dispersant
or other suitable excipients suitable ~or preparing a
proper size tablet for the dosage and particular antifungal
compound of the formula I. For pediatric or geriatric use
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the antifungal compound may be formulated into a flavored
liquid suspension, solution or emulsion. A preferred oral
formulation is linoleic acid, cremophor RH-60 and water and
preferably in the amount (by volume) of 8% linoleic acid,
5% cremophor RH-60, 87~ sterile water and a compound of
formula I in an amount of from about 2.5 to about 40 mg/ml.
For topical use the antifungal compound may be
formulated with a dry powder for application to the skin
surface or it may be formulated in a liquid formulation
comprising a solubilizing a~ueous liquid or non-aqueous
liquid, e.g., an alcohol or glycol.
The following formulation examples are
illustrative only and are not intended to limit the scope
of the invention in any way. The term "active ingredient~
means a compound according to formula I or a
pharmaceutically acceptable salt thereof.
- Formulation 1
Hard gelatin capsules are prepared using the
following ingredients:
Quantity
(m~/ca~sule)
Active ingredient250
Starch, dried 200
Magnesium stearate10
Total 460 mg
The solution of the above ingredients generally
is administered intravenously to a subject at a rate of 1
ml per minute.
The present invention further provides a method
for treating or preventing the onset of Pneumocystis
pneumonia in a host susceptible to Pneumocystis pneumonia
which comprises ~m;nistering an e~~ective amount of a
compound of formula I, or a pharmaceutically acceptable
salt thereof, to a host in need of such treatment. The
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_
compounds of formula I can be used prophylactically to
prevent the onset of the infection which is caused by the
organism Pneumocystis carinii, or alternatively they can be
used to treat a host that has been infected with P.
carinii. A compound of ~ormula I may be administered
parenterally, for example using intramuscular, intravenous
or intra-peritoneal injection, orally or by inhaling
directly into the airways of the lungs. A preferred mode
of administration is inhalation of an aerosol spray
formulation of a compound of formula I.
With respect to antiparasitic activity, the term
~effective amount," means an amount of a compound of the
present invention which is capable of inhibiting parasitic
activity. An effective amount of the compound of formula I
is from about 3 mg/kg of patient body weight to about
100 mg/kg. The amount administered may be in a single
daily dose or multiple doses of, for example, two, three or
four times daily throughout the treatment regimen. The
amount of the individual doses, the route of delivery, the
fre~uency of dosing and the term of therapy will vary
according to such factors as the intensity and extent of
infection, the age and general health of the patient, the
response of the patient to therapy and how well the patient
tolerates the drug. It is known that Pneumocystis
pneumonia infections in AIDS patients are highly refractory
owing to the nature of the infection. For example, in
severe, advanced infections the lumenal surface of the air
passages ~ecomes clogged with infectious matter and
extensive parasite development occurs in lung tissue. A
patient with an advanced infection will accordingly require
higher doses for longer periods of time. In contrast,
immune deficient patients who are not severely infected and
who are susceptible to Pneumocystis pneumonia can be
treated with lower and less fre~uent prophylactic doses.
