Note : Les descriptions sont présentées dans la langue officielle dans laquelle elles ont été soumises.
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, TITLE OF THE INVENTION
WET GRANULATION FORMULATION OF A GROVVTH
HORMONE SECRETAGOGUE
5 FIELD OF THE INVENTION
The present invention relates to solid dosage formulations
comprising a growth hormone secretagogue and process for their
manufacture. More specifically, the invention relates to a wet
gr~n~ tion formulation of a growth hormone secretagogue compound.
10 The present invention further relates to an amorphous form of a growth
hormone secretagogue, processes for its preparation and uses thereof.
BACKGROUND OF THE INVENTION
Growth hormone, which is secreted from the pituitary,
15 stim~ tes growth of all tissues of the body that are capable of growing.
In addition, growth hormone is known to have the following basic
effects on the metabolic processes of the body: (1) Increased rate of
protein synthesis in all cells of the body; (2) Decreased rate of
carbohydrate lltili7~tion in cells of the body; (3) Increased mobilization
20 of free fatty acids and use of fatty acids for energy. A deficiency in
growth hormone secretion can result in various medical disorders, such
as dwarfism.
Various ways are known to release growth hormone. For
example, chemicals such as arginine, L-3,4-dihydroxyphenyl~l~nine
25 (L-DOPA), glucagon, vasopressin, and insulin inrl~lced hypoglycemia, as
well as activities such as sleep and exercise, indirectly cause growth
hormone to be released from the pituitary by acting in some fashion on
the hypoth~l~mns perhaps either to decrease somatostatin secretion or to
increase the secretion of the known secretagogue growth hormone
30 releasing factor (GRF) or an unknown endogenous growth hormone-
releasing hormone or all of these.
In cases where increased levels of growth hormone were
desired, the problem was generally solved by providing exogenous
grow~ hormone or by ~1mini~tering GRF or a peptidal compound
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which stimulated growth hormone production and/or release. In either
case the peptidyl nature of the compound necessitated that it be
~tlmini~stered by injection. Tniti~lly the source of growth hormone was
the extraction of the pituitary glands of cadavers. This resulted in a
5 very expensive product and carried with it the risk that a disease
associated with ~e source of the pituitary gland could be tr~ncmi~te~l to
the recipient of the growth hormone. Recombinant growth hormone
has become available which, while no longer carrying any risk of
disease tr~n~mi~.~ion, is still a very expensive product which must be
10 given by injection or by a nasal spray.
Other compounds have been developed which stimulate the
release of endogenous growth hormone such as analogous peptidyl
compounds related to GRF or the peptides of U.S. Patent 4,411,890.
These peptides, while considerably smaller than growth hormones are
15 still susceptible to various proteases. As with most peptides, their
potential for oral bioavailability is low. Non peptidal growth hormone
secretagogues with a benzolactam structure are disclosed e.g., in U.S.
Patents 5,206,235, 5,283,241, 5,284,841, 5,310,737, 5,317,017,
5,374,721, 5,430,144, 5,434,261, 5,438,136 and PCT Publications WO
20 95/03289, WO 95/03290, WO 95/09633. Other growth hormone
secretagogues are disclosed in PCT Patent Publications WO 94/11012,
WO 94/13696, WO 94/19367, WO 95/13069 and WO 95/14666.
In particular, Examples 18, 19 and 55 of U.S. Patent No.
5,536,716 (PCT Patent Publication WO 94/13696) and Proc. Natl.
25 Acad. Sci. USA. 2~, 7001-7005 (July 1995) disclose the compound N-
[1 (R)- t(1,2-dihydro- 1 -methanesulfonyl-spiro [3H-indole-3,4'-piperdin] -
1 '-yl)carbonyl]-2-(phenylmethyl-oxy)ethyl]-2-amino-2-rnethyl-
prop~n~mide, and salts thereof, especially the methanesulfonate salt,
which has the structure:
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,~ H H CH~3 CH3
--O--C~N~NH2 ~ CH3S~2H
so2CH3
This compound is a growth hormone secretagogue which
stim~ tes the release of growth hormone in humans and ~nim~l~. This
property can be utilized to promote the growth of food ~nim~l~ to
5 render the production of edible meat products more efficient, and in
humans, to treat physiological or medical conditions characterized by a
deficiency in growth hormone secretion, and to treat medical conditions
which are improved by the anabolic effects of growth hormone.
U.S. Patent No. 5,536,716 and PCT Patent Publication WO
10 94/13696 disclose methods for preparing this compound (see Examples
18, 19 and 55). In particular, Example 55 states that the compound
prepared by recryst~lli7~tion from ethyl acetate-ethanol-water had a
melting point of 166-168~C. Proc. Natl. Acad. Sci. USA~ 92, 7001-
7005 (July 1995) notes that this compound isolated as a monohydrate
had a melting point of 168-170~C.
Standard methods for tablet formulation of the active
ingredient such as direct compression suffer from problems. In
particular, this compound is relatively unstable in standard
ph~ eutical formulations. In addition, this compound as a buL~ drug
20 ~urrels from poor flow properties, nevertheless, wet granulation was
discovered to overcome these difficulties preparing tablet formulations.
Tablets prepared by the wet gr~n~ tion method produced excellent
content uniforII~ity, coupled with suitable tablet dissolution and stability.
The tablets of the present invention, prepared by wet gr~nnl~tion~
25 possessed good hardness at normal machine pressures.
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The present invention provides a wet granulated
form~ tion of the compound N-[l(R)-[(1,2-dihydro-1-methane-
sulfonyl-spiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenyl-
methyl-oxy)ethyl]-2-amino-2-methyl-propanamide methanesulfonate and
5 process therefore wherein the tablet formulation is stable and robust.
The present invention further provides an amorphous forrn of the
compound, processes for its preparation and uses thereof.
~UMMARY QF THE INVENTION
The present invention relates to a process for the
preparation of a tablet cont~inin~ a growth hormone secretagogue as the
active ingredient. The tablet is prepared by forming a powder blend of
the active ingredient N-[l(R)-[(1,2-dihydro-1-methanesulfonyl-
spiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyl-
oxy)ethyl]-2-amino-2-methylprop~n~mide, or a pharmaceutically
acceptable salt thereof, in particular the meth~n~sulfonate salt, with a
binder/diluent, a first diluent, a second diluent, a first portion of a
integrant, and a lubricant; wet gr~ tin~ the powder blend with a
solution of ethanol/water to form granules; drying the granules to
remove the ethanol/water; ~1t1in~ a second portion of a ~ integrant;
lubricating the granules; and compressing the dried granules into the
desired tablet form.
The present invention fur~er relates to a novel amorphous
form of the compound N-[l(R)-[(1,2-dihydro-1-methanes-ulfonyl-
spiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)-
ethyl]-2-amino-2-methylpropanamide meth~nesulfonate. This
amorphous form is produced directly for example as a result of the
process of tablet form~ tion and has advantages over the other known
forms of N-[ 1 (R)-[( 1 ,2-dihydro- 1 -methanesulfonylspiro[3H-indole-3,4'-
piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-~mino-2-
methylpropanamide methanesulfonate in tenns of chemical and physical
stability.
-
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S
DETAILED DESCRIPTION OF THE INVENTION
The present invention is concerned with a process for the
preparation of a tablet cont~ining an active ingredient of the compound:
N-[l(R)-[(1,2-dihydro-1-methanesulfonylspiro[3H-indole-3,4'-piperdin]-
5 1 '-yl)carbonyl~-2-(phenylmethyloxy)ethyl]-2-amino-2-methyl-
propanamide, or a pharmaceutically acceptable salt thereof, in particular
the methanesulfonate salt, comprising the steps of:
(1) forming a powder blend of the active ingredient with a
binder/diluent, a first diluent, a second diluent, and a first
portion of a r~ integrant,
(2) wet granulating the powder blend with a solution of
ethanol/water to form granules,
(3) drying the granules to remove the ethanol/water,
(4) adding a second portion of a ~ ntegrant;
(5) lubricating the granules; and
(6) compressing the dried granules into a desired tablet form.
In a class is the process wherein the active ingredient is N-
[l(R)-[(1,2-dihydro-1-methanesulfonylspiro[3H-indole-3,4'-piperdin]-
20 1'-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-methyl-
prop~n~micle methanesulfonate.
In one embodiment is the process wherein the tablet
contains about 0.1 to 50% by weight of the active ingredient.
In a subclass is the process further comprising the step of
25 coating the tablet. Illustrative of the invention is the process wherein
coating the tablet is accomplished by:
(1) dry blending titanium dioxide (optionally mixed with talc)
with hydroxypropyl methylcellulose and hydroxypropyl
cellulose to form a dry powdered blend;
(2) ~ lin~ the dry powdered blend to water to form a slurry;
3) ~ in~ water to the slurry with stirring to form a
suspension; and
(4) applying the suspension to the tablets.
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The present invention is also concerned with
pharmaceutical formlll7.tions prepared by the subject process and their
use in the treatment of certain disorders and diseases.
Exemplifying the present invention is ~e process
comprising the steps of:
(1) forming a powder blend of the active ingredient with a
binder/diluent, a first ~ ent a second diluent, and a
intf-grant, from 2 to 25 minutes using a mixer;
(2) wet gr~n~ tin~ the powder blend by ~ri(lin~ a solution of
ethanol/water to the powder blend while rnixing over a 1 to
30 minute period to form granules;
(3) drying the granules to remove the ethanollwater with
heated air in a fluid bed dryer or tray dryer for 10 minlltes
to 24 hours;
(4) millin~ the dried granules to a uniform size;
(S) adding and blending a ~ integrant with the dried rnilled
particles for 2 to 30 minutes;
(6) ~ 1in~ and blending a lubricant to the mixture cont~inin~
the di.cintegrant for 30 seconds to 20 minutes; and
(7) compressing the lubricated granules mixture into a desired
tablet form.
Further illustrating the invention is the process wherein the
25 active ingredient is N-[l(R)-[(1,2-dihydro-1-methanesulfonylspiro[3H-
indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-
amino-2-methylpropanamide methanesulfonate.
Additional illustrations of the invention include the process
wherein: the binder/diluent is pregel~tini7:efl starch; the first diluent is
30 microcrystalline cellulose; the second diluent is calcium phosphate
dibasic; the disintegrant is croscarmellose sodium; and the lubricant is
magnesium stearate. Preferably, the solution of ethanol/water is in the
range of 0% to 80 % ethanol in water (w/w), more preferably in the
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range of 5% to 75 % ethanol/water (w/w) and even more preferably
approxim~tely 25% ethanol/75% water (w/w).
A fur~er illustration of the invention is the foregoing
process further comprising the step of applying a coating to the tablet.
5 More particularly illustrating the invention is the process wherein
coating the tablet is accomplished by:
( 1 ) dry blending titanium dioxide (optionally mixed with talc)
with hydroxypropyl methylcellulose and hydroxypropyl
cellulose to form a dry powdered blend;
(2) ~ ling the dry powdered blend to water to form a slurry;
(3) ~cltling water to the slurry with stirring to form a
suspension; and
(4) applying the suspension to the tablets.
More specifically exemplifying the invention is the process
comprising the steps of:
(1) forming a powder blend of the active ingredient with
pregel~tini7e-1 starch, microcrystalline cellulose, calcium
phosphate dibasic, and croscarmellose sodium, in a mixer
for about 3 to 25 rninutes;
(2) wet gr~n~ ting the powder blend by ~ 1ing a solution of
25% ethanol/75% water (w/w) to the powder blend while
mixing over a 1 to 30 minute period to form granules;
(3) drying the granules on a tray dryer or a fluid bed dryer for
about 1 to 12 hours to remove the ethanol/water;
(4) millin~ the dried granules to a uniform size using a Quadro
Comill or Fitz type mill;
(5) ~ ling and blending croscarmellose sodium with the dried
milled particles for about S to 30 minutes;
.. 30 (6) ~ ling and blending magnesium stearate to the mixture
cont~inin~ the croscarmellose sodium with a V blender for
about 1 to 5 minutes; and
(7) compressing the lubricated granules mixture into a desired
tablet forrn.
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Another example of the invention is the process wherein ~e
active ingredient is N-[ 1 (R)-[( 1 ,2-dihydro- 1-methanesulfonyl-spiro[3H-
indole-3,4'-piperdin~- 1 '-yl)carbonyl]-2-(phenylmethyloxy)-ethyl]-2-
amino-2-methylpropanarnide methanesulfonate.
S In a subclass is the foregoing process further comprising
the step of coating the tablet. Further exemplifying the invention is the
foregoing process further comprising the steps of:
( 1 ) dry blending titanium dioxide (optionally mixed with talc)
with hydroxypropyl methylcellulose and hydroxypropyl
cellulose to form a dry powdered blend;
(2) adding the dry powdered blend to water to form a slur~y;
(3) adding water to the slurry with stirrin~ to form a
suspension; and
(4) applying the suspension to the tablets.~5
An additional illustration of the present invention is a solid
dosage form cont~ining an active ingredient of N-[l(R)-[(1,2-dihydro-1-
methanesulfonylspiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-
(phenylmethyloxy)ethyl]-2-amino-2-methylprop~n~mide, or a
20 rh~ ceutically acceptable salt thereof, in particular the
methanesulfonate salt, wherein the dosage form is prepared by the
process.
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The present invention further relates to a novel amorphous
form of the compound N-[l(R)-[(1,2-dihydro-1-methane-sulfonyl-
spiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenyl-
methyloxy)ethyl]-2-amino-2-methylprop~n~mide methanesulfonate.
This amorphous form is produced directly as a result of the instant
process of tablet formulation.
The amorphous form of N-[l(R)-[(1,2-dihydro-1-
methanesulfonyl-spiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-
(phenylmethyl-oxy)ethyl]-2-amino-2-methylprop~n~mide
m~tll~nesulfonate exhibits a lack of crystallinity. The lack of
crystallinity was confirmed by X-ray analysis wherein he ~-ray
diffraction pattern showed an amorphous halo.
The X-ray powder diffraction (XRPD) pattern was
collected using a Phiulips APD3720 Automated Powder Diffraction
lS instrument with copper Ka radiation. Measurements were made from
2~ to 40~ (2 theta) with the s~mple m~int~ined at ambient room
temperature.
In addition, e~min~tion of the amorphous form under
microscopy showed no biorefringence.
The ~morphous form may be prepared by evaporating a
concentrated solution of N-[ 1 (R)-[( 1 ,2-dihydro- 1 -methanesulfonyl-
spiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)-
ethyl]-2-amino-2-methylprop~qn~midP methanesulfonate in 25% aqueous
ethanol (980 mglml) at 40~C to give a solid.
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Granulation is ~e process of ~ lin~ a solvent, such as
water or water/ethanol, to a powder mixture until granules are formed.
The gr~nul~tion step may be varied from 2 to 35 minlltes, preferably 3
to 10 minutes, most preferably 4 to 8 minutes. Preferably, the granules
5 are dried using a fluid bed dryer or tray dryer. Milling of ~e dried
granules is accomplished using a Quadro Comill or Fitz rnill. The
lubrication step is the process of ~l(iin~ lubricant to the mixture. The
lubrication step may be varied from 30 seconds to 20 minlltes,
preferably about 1 mirlute.
The disclosed process may be used to prepare solid dosage
forms, particularly tablets or granules, for medicinal ~-imini~tration.
The term "tablet," as used herein, is intended to encompass
compressed ph~rm~ceutical dosage form~ tions of all shapes and sizes,
whether coated or uncoated. Substances which may be used for coating
15 include hydroxypropyl methylcellulose (HPMC), hydroxypropyl
cellulose (HPC), titanium dioxide, talc, sweeteners and colorants.
The term "active ingredient," as used herein includes both
the free base N-[1(R)-[(1,2-dihydro-1-methane-sulfonyl-spiro[3H-
indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenyl-methyl-oxy)ethyl]-2-
20 amino-2-methyl-prop~n~rnide, as well as the ph~rm~ceutically
acceptable salts thereof, in particular, N-[l(R)-[(1,2-dihydro-1-methane-
sulfonyl-spiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenyl-
methyl-oxy)ethyl]-2-amino-2-methyl-propanamide methanesulfonate and
crystal forms thereof. A preferred crystal form for use in the present
25 invention is that designated Form I.
Preferred diluents include: lactose, microcryst~llin
cellulose, calcium phosphate(s), mannitol, powdered cellulose,
pregel~tini7e-1 starch and other suitable diluents (see, e.g., Remington's
Pharmaceutical Sciences, 18th Edition, 1990, p. 1635). Microcrystalline
30 cellulose and calcium phosphate dibasic, are particularly preferred.
Specifically, microcrystalline cellulose NF, especially Avicel PH101, ~e
tr~tlem~rk~d name for microcrystalline cellulose NF manufactured by
FMC Corp. is preferred.
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Preferred binders include pregel~tini7e.d starch, hydroxy-
propyl methylcellulose, hydroxypropyl cellulose, polyvinylpyrrolidone
(PVP) and other known binders (see, e.g., Remington's Ph~ eutical
Sciences, 18th Edition, 1990, pp. 1635-1636) and mixtures thereof.
S Most preferably, pregel~tini7ed starch as employed as a binder.
Specifically, starch pregel~tini~e~l NF 1500 m~nnf~ctured by Colorcon
Corporation is most preferred.
The disintegrant may be one or more of several starches,
clays, celluloses, algins, gums or cro.~.elink~l polymers known to those
10 slcilled in the art (See, e.g., Remington's Ph~rm~c eutical Sciences, 18th
Edition, 1990, p. 1637) and mixtures thereof. Preferably, one or more
of several modified starches or modified cellulose polymers, such as
microcrystalline cellulose and croscarmellose sodium, are used.
Croscarmellose sodium Type A, commercially available under the trade
15 name "Ac-di-sol," is particularly preferred.
Preferred lubricants include m~,~nesium stearate, zinc
stearate, calcium stearate, stearic acid, s~ ce active agents such as
sodium lauryl sulfate, m~nesium lauryl sulfate, propylene glycol,
sodium do~lec~ne sulfonate, sodium oleate sulfonate and sodium laurate
20 mixed with stearates and talc, sodium stearyl fumarate, hydrogenated
vegetable oils, glyceryl palmitostearate, glyceryl behenate, sodium
benzoate, mineral oil, talc and other known lubricants (see, e.g.,
Remington's Ph~rm~ceutical Sciences, 18th Edition, 1990, pp. 1636-
1637), and mixtures thereof. An especially preferred lubricant is
25 m~nesium stearate.
The active ingredient, N-[1(R)-[(1,2-di-hydro-1-methane-
sulfonylspiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenyl-
methyloxy)ethyl]-2-amino-2-methylpropanamide, may be prepared
according to the methods disclosed in U.S. Patent No. 5,536,716, PCT
30 Patent Publication WO 94/13696 and the methods disclosed herein.
The pharmaceutically acceptable salts of N-[1(R)-[(1,2-
dihydro- 1 -methane-sulfonyl-spiro[3H-indole-3,4'-piperdin]- 1 '-
yl)carbonyl] -2-(phenyl-methyl-oxy)ethyl] -2-amino-2-methyl-
propanamide may be employed in the instant invention. Examples of
_
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ph~rm~eeutically acceptable salts include the pharm~relltically
acceptable acid addition salts, such as the salts derived from using
inorganic and organic acids. Examples of such acids are hydrochloric,
nitric, sulfuric, phosphoric, formic, acetic, trifluoroacetic, propionic,
maleic~ succinic, malonic, methane sulfonic and the like.
The pharmaceutical compositions of the present invention
comprise 0.1 to 50 % by weight of an active ingredient, N-[l(R)-[(1,2-
dihydro- 1 -methanesulfonylspiro[3H-indole-3,4'-piperdin]- 1'-
yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-methylpropanamide,
or a pharmaceutically acceptable salt thereof, preferably N-[l(R)-[(1,2-
dihydro- 1 -methanesulfonylspiro[3H-indole-3,4'-piperdin]- 1'-
yl)carbonyl] -2-(phenylmethyloxy)ethyl] -2-amino-2-methylpropanamide
methanesulfonate; 0 to 77 % by weight of a binder/diluent; 0 to 77 % by
weight of a first diluent; 0 to 77 % by weight of a second diluent;
0 to 6 % by weight of a ~ integrant; and 0 to 5 % by weight of a
lubricant. It will be appreciated by one skilled in the art that ~e sum of
the proportions of the active ingredient, the binder/diluent, the first
diluent, the second diluent, the di~integrant, and the lubricant are not
greater than 100% by weight.
More speci~lcally, the binder/diluent is selected from
hydroxy-propyl methylcellulose, hydroxypropyl cellulose,
pregel~tini7e-1 starch or polyvinylpyrrolidone; the first and second
diluents are independently selected from lactose, microcrystalline
cellulose, calcium phosphate dibasic, mannitol, powdered cellulose or
pregel~tini7ed starch; the ~ integrant is selected from microcrystalline
or crosc~rmellose sodium; and the lubricant is selected from m~nesium
stearate, calcium stearate, steric acid or a s~ ce active agent.
In a specific embodiment, the binder/diluent is
pregel~tini7ed starch; the first diluent is microcrystalline cellulose; the
second diluent is calcium phosphate dibasic; the ~ integrant is
crosc~ llose sodium; and the lubricant is m~gnesium stearate.
,
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The ph~rm~ceutical compositions of the present invention
are preferably in the form of tablets. The tablets may be, for example,
from 50 mg to 1.0 g in net weight, preferably 100 to 800 mg net
weight, more preferably 100 to 400 mg net weight.
Plefel-~,d ph~ ceutical compositions comprise about
1 to 30% by weight of the active ingredient; about 20 to 40% by weight
of pregel~tini7ed starch; about 10 to 20% by weight of microcrystalline
cellulose; about 20 to 50% by weight of calcium phosphate dibasic;
about 5 to 15% by weight of crosc~rmellose sodium; and about 0.05 to
5% by weight of m~nesium stearate.
It will be appreciated by one skilled in the art that the sum
of the above proportions of the active ingredient, pregel~tini7e~1 starch,
microcrystalline cellulose, calcium phosphate dibasic, croscarmellose
sodium, and m~gn~si-lm stearate are not greater than 100% by weight.
More preferred ph~ ceutical compositions in accordance
with the present invention include those comprising the noted
ingredients:
(1) about 1 to 2% by weight of the active ingredient;
about 25 to 35% by weight of pregel~tini7ed starch; about 10 to 20%
by weight of microcrystalline cellulose; about 45-55% by weight of
calcium phosphate dibasic; about 4 to 8% by weight of croscarmellose
sodium; and about 0.1 to 1% by weight of m~gn~sium stearate.
(2) about 5 to 10% by weight of the active ingredient,
about 25 to 35% by weight of pregel~tini7ed starch; about 10 to 20% by
weight of microcrystalline cellulose; about 40 to 50% by weight of
calcium phosphate dibasic; about 4 to 8% by weight of croscarmellose
sodium; and about 0.1 to 1% by weight of m~gnesium stearate.
(3) about 25 to 35% by weight of the active ingredient,
about 15 to 25% by weight of pregel~tini7ed starch; about 10 to 20% by
weight of microcrystalline cellulose; about 15-25% by weight of
calcium phosphate dibasic; about 10 to 20% by weight of croscarmellose
sodium; and about 0.1 to l~o by weight of magnesium stearate.
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It will be appreciated by one skilled in the art that the sum
of the above proportions of the active ingredient, pregel~tini7ed starch,
microcrystalline cellulose, calcium phosphate dibasic, croscarmellose
sodium, and m~gnesium stearate are not greater than 100% by weight.
Especially lefell~,d ph~ ceutical compositions as
envisioned for commercial development are as follows:
Tablets of 1.0 mg potency free base:
about 1.18% by weight of active ingredient as the
10 methanesulfonate salt; about 30.0% by weight of pregel~tini7e.d starch;
about 15.0% by weight of microcrystalline cellulose; about 47.3% by
weight of calcium phosphate dibasic; about 6.0% by weight of
crosc~rmellose sodium; and about 0.5% by weight of magnesium
stearate. This composition comprises about 1.2 mg of active ingredient
15 as the mt-th~nesulfonate salt; about 30 mg of pregel~tini7e-1 starch; about
15 mg of microcrystalline cellulose; about 47.3 mg of calcium
phosphate dibasic; about 6.0 mg of croscarmellose sodium; and about
0.5 mg of m~gnesium stearate per dosage unit.
Optionally, the 1.0 mg potency tablet may be coated with a
20 coating comprising about 0.8% by weight of hydroxypropyl
methylcellulose; about 0.8% by weight of hydroxypropyl cellulose;
about 0.32% by weight of titanium dioxide; and about 0.08% by weight
of talc (as a percentage of the core tablet weight).
Tablets of 5.0 mg potency free base:
about 1.48% by weight of active ingredient as the
methanesulfonate salt; about 30.0% by weight of pregel~tini7e-1 starch;
about 15.0% by weight of microcrystalline cellulose; about 47.0% by
weight of calcium phosphate dibasic; about 6.0% by weight of
croscarmellose sodium; and about 0.5% by weight of m~gnesium
stearate. This composition comprises about 6 mg of active ingredient as
the methanesulfonate salt; about 120 mg of pregeliqtini7e-1 starch; about
60 mg of microcrystalline cellulose; about 188 mg of calcium phosphate
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- 15 -
dibasic; about 24 mg of croscarmellose sodium; and about 2 mg of
m~gnesium stearate per dosage unit.
Optionally, the 5.0 mg potency tablet may be coated with a
coating comprising about 0.8% by weight of hydroxypropyl
5 methylcellulose; about 0.8% by weight of hydroxypropyl cellulose;
about 0.32% by weight of titanium dioxide; and about 0.08% by weight
of talc (as a percentage of the core tablet weight).
Tablets of 25 m~ potency free base:
about 7.39% by weight of active ingredient as the
methanesulfonate salt; about 28.2% by weight of pregel~tini7e~1 starch;
about 14.2% by weight of microcrystalline cellulose; about 43.6% by
weight of calcium phosphate dibasic; about 6.0% by weight of
croscarmellose sodium; and about 0.5~o by weight of m~gn~-.sium
lS stearate. This composition comprises about 30 mg of active ingredient
as the methanesulfonate salt; about 113 mg of pregel~tini7e~1 starch;
about 57 mg of microcrystalline cellulose; about 174 mg of calcium
phosphate dibasic; about 24 mg of croscarmellose sodium; and about 2
mg of magnesium stearate per dosage unit.
Optionally, the 25 mg potency tablet may be coated with a
coating comprising about 0.8% by weight of hydroxypropyl
methylcellulose; about 0.8% by weight of hydroxyprop~l cellulose;
about 0.32% by weight of ~ llliUlll dioxide; and about 0.08% by weight
of talc (as a percentage of the core tablet weight).
Tablets of 100 mg potency free base:
about 29.5% by weight of active ingredient as the
meth~nesulfonate salt; about 19.5% by weight of pregel~tini7e,d starch;
about 15.0% by weight of microcrystalline cellulose; about 20.4% by
weight of calcium phosphate dibasic; about 15.0% by weight of
croscarmellose sodium; and about 0.5% by weight of magnesium
- stearate. This composition comprises about 1 18 mg of active ingredient
as the methanesulfonate salt; about 78 mg of pregel~tini7ed starch; about
60 mg of microcrystalline cellulose; about 82 mg of calcium phosphate
_
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dibasic; about 60 mg of croscarmellose sodium; and about 2 mg of
m~gn~sium stearate per dosage unit.
Optionally, the 100 mg potency tablet may be coated with a
coating comprising about 0.8% by weight of hydroxypropyl
5 methylcellulose; about 0.8% by weight of hydroxypropyl cellulose;
about 0.32% by weight of titanium dioxide; and about 0.08% by weight
of talc (as a percentage of the core tablet weight).
The tablets of the 1.0 mg potency are preferably
forrnulated in an 100 mg tablet by using 30 ~11 of a solution of 25%
10 ethanol/75% water per tablet. The tablets of the 5.0 mg potency are
preferably formulated in an 400 mg tablet by using 120 ,ul of a solution
of 25~o ethanol/75% water per tablet. The tablets of the 25 mg potency
are preferably formulated in an 400 mg tablet by using 120 ,ul of a
solution of 25% ethanol/75% water per tablet. The tablets of the 100
15 mg potency are preferably ~orm~ e~l in an 400 mg tablet by using 120
,ul of a solution of 25~o ethanol/75% water per tablet.
In a particularly preferred embodiment, the tablet
formulations of the instant invention are coated. In the pharmaceutical
compositions envisioned for commercial development described above,
20 the tablets of 1.0 mg, 5.0 mg, 25 mg and 100 mg potency free base are
coated with about 0.8 % by weight of hydroxypropyl methylcellulose;
about 0.8 % by weight of hydroxypropyl cellulose; about 0.32 % by
weight titanium dioxide; and about 0.08 % by weight of purified talc.
In ~e most preferred embodiment, the active ingredient in
25 the above-described pharmaceutical compositions is N-[1(~)-[(1,2-
dihydro- 1 -methanesulfonylspiro[3H-indole-3,4'-piperdin]- 1 '-
yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-methylprop~n~mic~
methanesulfonate .
The compositions of the present invention are in a form for
30 oral ~rlmini.ctration and may take the forrn of tablets, capsules, granules,
powders, tablets or granules for buccal ~(1mini~tration, or liquid
preparations such as suspensions. Granules and powders may be
ingested directly, or dispersed in water or other suitable vehicle prior to
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1ministration. Capsules may be of the hard or soft gelatin type,
including soft gelatin capsules.
The ph~rm~ceutical compositions of the present invention
may also contain other excepients conventional in the art such as
5 flavorings, sweeteners, and the like. Suitable flavorings include for
example fruit flavors or natural or synthetic mint or peppermint
fiavors. Suitable sweeteners include for example sugar, saccharin or
aspartame.
The utility of the active ingredient of the form~ tion of the
10 present invention as growth hormone secretagogues may be
demonstrated by methodology known in the art, such as an assay
described by Smith, et al., Science~ ~Q, 1640-1643 (1993) (see text of
Figure 2 therein). In particular, the active ingredient used in the
form~ tion the present invention had activity as a growth hormone
15 secretagogue in the aforementioned assay. Such a result is indicative of
the activity of the formulation of the present invention as a growth
hormone secretagogue.
The formlll~tions of the present invention may be
z~1mini~tered to ~nim~l~, including man, to release growth hormone in
20 vivo. For example, the forrnulations can be ~-1ministered to
commercially important ~nim~l~ such as swine, cattle, sheep and the like
to accelerate and increase their rate and extent of growth, to improve
feed efficiency and to increase milk production in such ~nim~l~. In
addition, these formulations can be ~-lmini~tered to humans in vivo as a
25 diagnostic tool to directly determine whether the pituitary is capable of
releasing growth hormone. For example, the formulation of the present
invention can be ~mini~tered in vil~o to children. Serum samples taken
before and after such ~(lrnini~tration can be assayed for growth
horrnone. Comp~ on of the arnounts of growth hormone in each of
30 these samples would be a means for directly determining the ability of
the patient's pituitary to release growth hormone.
Accordingly, the present invention includes within its scope
ph~ celltical compositions comprising, as an active ingredient, the
compound N- [ 1 (R)- [( 1 ,2-dihydro- 1 -methane-sulfonyl-spiro[3H-indole-
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3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenyl-methyl-oxy)ethyl]-2-amino-2-
methyl-propanamide in association with a pharn~ceutical carrier or
diluent. Optionally, the active ingredient of the pharmaceutical
compositions may comprise an anabolic agent in addition to the
5 compound N-[ 1 (R)-[( 1 ,2-dihydro- 1 -methane-sulfonyl-spiro[3H-indole-
3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenyl-methyl-oxy)ethyl]-2-amino-2-
methyl-propanamide or another composition which exhibits a different
activity, e.g., an antibiotic growth permittant or an agent to treat
osteoporosis or in combination with a corticosteroid to minimi7e the
10 catabolic side effects or with other pharmaceutically active materials
wherein the combination enhances efficacy and minimi7.es side effects.
Growth promoting and anabolic agents include, but are not
limited to TRH, diethylstilbesterol, amino acids, estrogens, 13-agonists,
theophylline, anabolic steroids, enkeph~lin~, E series prostaglandins,
15 compounds disclosed in U.S. Patent No. 3,239,345, e.g., zeranol, and
compounds disclosed in U.S. Patent No. 4,036,979, e.g., sulbenox or
peptides disclosed in U.S. Patent No. 4,411,890.
A still further use of the formulations of this invention is in
combination with other growth hormone secretagogues such as the
20 growth hormone releasing peptides GHRP-6, GHRP-l as described in
U.S. Patent Nos. 4,411,890 and publications WO 89/07110, WO
89/071 11 and B-HT920 as well as hexarelin and GHRP-2 as described in
WO 93/04081 or growth hormone releasing hormone (GHRH, also
design~te~l GRF) and its analogs or growth hormone and its analogs or
25 somatomedins including IGF-1 and IGF-2 or a-adrenergic agonists such
as clonidine or serotonin SH~ID agonists such as sumilli~lall or agents
which inhibit somatostatin or its release such as physostigmine and
pyridostigmine. In particular, the formulations of this invention may be
used in combination with growth hormone releasing factor, an analog of
30 growth hormone releasing factor, IGF-1, or IGF-2. For example, a
forrn~ tion of the present invention may be used in combination with
IGF-l for the treatment or prevention of obesity. In addition, a
formulation of this invention may be employed in conjunction with
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retinoic acid to improve the condition of musculature and skin that
results frorn intrinsic aging.
As is well known to those skilled in the art, the known and
potential uses of growth hormone are varied and mllltitl~tlinous. The
5 ~lmini~tration of the forrn~ tions of this invention for purposes of
stim~ tin~ the release of endogenous ~lowLh hormone can have the
same effects or uses as growth hormone itself. These varied uses of the
present formlllAtions thus may be sllrnm~rized as follows: stim~ ting
growth hormone release in elderly humans; treating growth hormone
10 deficient adults; prevention of catabolic side effects of glucocorticoids;
treatment of osteoporosis; stimulation of the immune system,
acceleration of wound healing; accelerating bone fracture repair;
treatment of growth retardation; treating acute or chronic renal failure
or insufficiency; treatment of physiological short stature, including
15 growth hormone deficient children; treating short stature associated
with chronic illness; treatment of obesity and growth retardation
associated with obesity; treating growth retardation associated with
Prader-Willi syndrome and Turner's syndrome; accelerating the
recovery and reducing hospit~li7~tion of burn patients or following
20 major ~,ur~ e~y such as gastrointestinal ~ ,e~y; treatment of intrauterine
growth retardation, and skeletal dysplasia, treatment of peripheral
neuropathies; replacement of growth hormone in stressed patients;
tre~tmerlt of osteochondrody-splasias, Noonans syndrome,
schizophrenia, depression, Alzheimer's ~ e~e, delayed wound h~lin~,
25 and psychosocial deprivation; treatment of pulmonary dysfunction and
ventilator dependency; attenuation of protein catabolic response after a
major operation; treating malabsorption syndromes; reducing cachexia
and protein loss due to chronic illness such as cancer or AIDS;
accelerating weight gain and protein accretion in patients on TPN (total
30 parenteral nutrition); tre~tTnent of hyperinsulinemia including
nesidioblastosis; adjuvant treatment for ovulation induction and to
prevent and treat gastric and duodenal ulcers; to stimulate thymic
development and prevent the age-related decline of thymic function;
adjunctive t~herapy for patients on chronic hemodialysis; treatment of
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- 20 -
immllno~u~lessed patients and to enhance antibody response following
vaccination; increasing the total lymphocyte count of a human, in
particular, increasing the T4/Tg-cell ratio in a human with a depressed
T4/Tg-cell ratio resulting, for example, from physical trauma, such as
5 closed head injury, or from infection, such as bacterial or viral
infection, especially infection with the hllm~n imm~lnode~lciency virus;
improvement in muscle strength, mobility, m~intrnance of skin
thickness, metabolic homeostasis, renal hemeostasis in the frail elderly;
stimulation of osteoblasts, bone remodelling, and cartilage growth;
10 stimlll~tion of the immnne system in companion ~nim~ and treatment
of disorders of aging in companion ~nim~ ; growth promotant in
livestock; and stimulation of wool growth in sheep. Further, the instant
compounds are useful for increasing feed efficiency, promoting growth,
increasing milk production and improving the carcass quality of
15 livestock. Likewise, the instant formulations are useful in a method of
treatment of diseases or conditions which are benefited by the anabolic
effects of enhanced growth hormone levels.
In particular, the instant formulations are useful in the
prevention or treatment of a condition selected from the group
20 consisting of: osteoporosis; catabolic illness; immlmP deficiency,
including ~at in individuals with a depressed T4/Tg cell ratio; hip
fracture; musculoskeletal impairment in the elderly; growth hormone
deficiency in adults or in children; obesity; cachexia and protein loss due
to chronic illness such as AIDS or cancer; and treating patients
25 recovering from major ~ul~,ely, wounds or burns, in a patient in need
thereof.
In addition, the instant form~ tions may be useful in the
treatment of illnesses induced or facilitated by corticotropin releasing
factor or stress- and anxiety-related disorders, including stress-induced
30 depression and he~ rhr, abdominal bowel syndrome, immlmP:
suppression, HIV infections, Alzheimer's disease, gastrointestinal
disease, anorexia nervosa, hemorrhagic stress, drug and alcohol
withdrawal symptoms, drug addiction, and fertility problems.
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It will be known to those skilled in the art that there are
numerous compounds now being used in an effort to treat the diseases
or therapeutic indications enumerated above. Combinations of these
therapeutic agents some of which have also been mentioned above in and
5 with the formulations of this invention will bring additional,
complementary, and often synergistic properties to enhance the growth
proInotant, anabolic and desirable properties of these various
therapeutic agents. In these combinations, the therapeutic agents and the
active ingredient in the formulations of this invention may be
10 independently present in dose ranges from one one-hundredth to one
times the dose levels which are effective when these compounds and
active ingredients are used singly.
Combined therapy to inhibit bone resorption, prevent
osteoporosis and enhance the he.~ling of bone fractures can be illustrated
15 by combinations of bisphosphonates and the formulations of this
invention. The use of bisphosphonates for these utilities has been
reviewed, for example, by Hamdy, N.A.T., Role of Bisphosphonates in
Metabolic Bone Diseases, Trends in Endocrinol. ~etab., 4, 19-25
(1993). Bisphosphonates with these utilities include alendronate,
20 tiludronate, dimethyl-APD, risedronate, etidronate, YM-175,
clodronate, pamidronate, and BM-210995. According to their potency,
oral daily dosage levels of the bisphosphonate of between 0.1 mg and 5
g and daily dosage levels of the active ingredient of the formulation of
this invention of between 0.01 mg/kg to 20 mg/kg of body weight are
25 ~lmini~tered to patients to obtain effective treatment of osteoporosis.
In the case of alendronate daily oral dosage levels of 0.1 mg
to 50 mg are combined for effective osteoporosis therapy with 0.01
mg/kg to 20 mg/kg of the active ingredient employed in the formnl~tion
of this invention. Osteoporosis and other bone disorders may also be
30 treated with the formulations of this invention in combination with
calcitonin, estrogens, raloxifene and calcium supplements such as
calcium citrate.
Anabolic effects especially in the treatment of geriatric
male patients are obtained with formulations of this invention in
-
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combination with anabolic steroids such as oxymetholone,
methyltesterone, fluoxymesterone and stanozolol.
The ph~ eutical tablet compositions of the present
invention may also contain one or more additional formulation
5 ingredients selected from a wide variety of excipients (also referred to
as "additives") known in the ph~ ceutical form~ qtion art. According
to the desired properties of the tablet, any number of additives may be
selected, alone or in combination, based upon their known uses in
preparing tablet compositions. Such additives include, but are not
10 limite-l to, diluents, binders, compression aids, ~lisintegrants, lubricants, flavors, flavor enhancers, sweeteners and preservatives. Due to the
bitter taste of the active ingredient, the inclusion of a sweetener may be
desired.
The dosage of active ingredient in ~e compositions of this
15 invention may be varied; however, it is necessary that the amount of the
active ingredient be such that a suitable dosage form is obtained. The
selected dosage depends upon the desired therapeutic effect, on the route
of ~rlmini~tration~ and on the duration of the tre~tm~nt Generally,
dosage levels of between 0.0001 to 10 mg/kg. of body weight daily are
20 ~iminictered to patients and ~nim~l~, e.g., m~mm~l~, to obtain effective
release of growth hormone. Preferably, the dosage level will be about
0.001 to about 25 mg/kg per day; more preferably about 0.01 to about
10 mg/lcg per day.
Methods for preparing the formulations of the present
25 invention, as well as the active ingredient are illustrated in the following
Examples. The following examples are given for the purpose of
illustrating the present invention and shall not be construed as being
limit~tions on the scope or spirit of the in~t~nt invention.
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FXAMPLE 1
N-[ 1 (R)-[( 1 ,2-Dihydro- 1 -methanesulfonylspiro~3H-indole-3,4'-
piperdin~- 1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-
methylpropanamide
~tep A: 1,2-Dihydro-1-methanesulfonylspiro[3H-indole-3,4'-
piperdinelhydrochloride
To a solution of 1.20 g (5.8mmol) of 1'-methyl-1,2-
dihydro-spiro[3H-indole-3,4'-piperdine] (prepared as described by H.
Ong, et al., J. Med. Chem., ~, 981-986 (1983)) in 20 mL of dry
dichloromethane at 0~C was added triethylamine (0.90 mL; 6.4 mmol)
and methanesulfonyl chloride (0.49 mT.; 6.35 mmol) and stirred for 30
min. The reaction mixture was poured into 15 mT of saturated aqueous
sodium bicarbonate solution and extracted with dichloromethane (2X10
mL). The combined organics were washed with brine (20 mL), dried
over anhydrous potassium carbonate, filtered and the solvent removed
under reduced pressure to yield 1.44 g of the methanesulfon~mi~l~
derivative as pale yellow oil which was used without purification.
To a solution of above crude product in 20 mL of dry 1,2-
dichloroethane at 0~C was added 1.0 mL (9.30 mmol) of 1-chloroethyl
chloroformate, and then stirred at RT for 30 min and finally at reflux
for lh. The reaction mixture was concentrated to approximately one
third of the volume and then diluted with 20 mL of dry methanol and
refluxed for l.Sh. The reaction was cooled to RT and concentrated to
approximately one half of the volume. The precipitate was filtered and
washed with a small volume of cold methanol. This yielded 1.0 g of the
piperidine HCl salt as a white solid. The filtrate was concentrated and a
small volume of methanol was added followed by ether. The
precipitated material was once again filtered, washed with cold
methanol, and dried. This gave an additional 0.49 g of the desired
product. Total yield 1.49 g (70%).
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lH NMR (CDC13, 200MHz) o 7.43-7.20 (m, 3H), 7.10 (dd, lH), 3.98
(bs, 2H), 3.55-3.40 (bd, 2H), 3.35-3.10 (m, 2H), 2.99 (s, 3H), 2.15 (t,
2H), 2.00 (t, 2H).
5 Step B: N-[l(R)-[(1,2-Dihydro-l-methanesulfonylspiro[3H-indole-
3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-
2-[(1,1 -dimethylethoxy)carbonyl]arnino-2-rnethyl-
prop~n~mide
To 0.35g (l.lS rnmol) of (2R)-2-[(1,1-dimethylethoxy)-
carbonyl]amino-3-[2-(phenylmethyloxy)ethyl]-1-propanoic acid in 13
ml of dichloromethane was added 1,2-dihydro-1-methanesulfonylspiro-
[3H-indole-3,4'-piperdine] hydrochloride (0.325 g; 1.07 rnmol), 0.18
mL (1.63 rnrnol) of N-methylmorpholine, O.lS9 g (1.18 mmol) of 1-
hydroxyben7tri~7.01e(HOBT) and stirred for 15 min. EDC (0.31 g; 1.62
mol) was added and stirring was continllç~l for lh. An additional 60 ~L
of N-methylmorpholine was added and stirred for 45 min. The reaction
mixtllre was poured into S mL of water and the organic layer was
separated. The organic layer was washed with 5 mL of 0.5N aqueous
hydrochloric acid and 5 mL of saturated aqueous sodium bicarbonate
solution. The combined organics were dried over anhydrous
m~gn~.sium sulfate, and concentrated to yield 0.627 g of the product as a
yellow foarn which was used without puri~lcation.
To a 0.627 g (1.07 mmol) of the above product in 5 mL of
dichloromethane was added 1.0 mL of trifluoroacetic acid and stirred at
RT for 75 rnin. An additional 1.00 mL of trifluoroacetic acid was
added and stirred for 10 min. The reaction mixture was concentrated,
diluted with 5.0 mL of dichloromethane and carefully basified by
pouring into 10 mL of 10% aqueous sodium carbonate solution. The
organic layer was separated and the aqueous layer was further extracted
with 2X15 mL of dichloromethane. The combined organics were
washed with S mL of water, dried over potassium carbonate, ~lltered
and concentrated to give the 0.486 g of the amine as a light yellow foam
which was used without purification.
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- 25 -
To 0.486 g (1.01 mmol) of the amine and 10 mL of
dichloromethane was added 0.26g (1.28 mmol) of 2-[(1,1-dimethyl-
ethoxy)carbonyl]amino-2-methyl-propanoic acid, 0.173 g (1.28 mmol)
of l-hydroxybenztriazole (HOBT) and EDC (0.245 g; 1.28 mol) and
stirried at RT overnight. The reaction mixture was poured into 5.0 mL
of water and the organic layer was separated. The aqueous layer was
back extracted with S mL of dichloromethane. The combined organics
were washed with 5.0 mT. of 0.5N aqueous hydrochloric acid, 5 mL of
saturated aqueous sodium bicarbonate solution dried over anhydrous
magnesium sulfate, and concentrated to yield 0.751 g of the crude
product as a yellow foam A solution of this crude product in
dichloromethane was chromatographed on 25 g of silica gel and eluted
first with hexanes/acetone/dichloromethane (70/25/5) and then with
hexanes/acetone/dichloromethane (65/30/S). This gave 0.63 g of the
title compound as a white solid.
lH NMR (CDC13, 400MHz) Compound exists as a 3:2 mixture of
rotamers o 7.40-7.10 (m, 6H), 7.06 (d, 1/3H), 7.02 (t, 1/3H), 6.90 (t,
1/3H), 6.55 (d, 1/3H), S.lS (m, lH), 4.95 (bs, lH), 4.63 (bd, 1/3H),
4.57-4.40 (m, 2 2/3 H), 4.10 (bd, 1/3H), 4.00 (bd, 1/3H), 3.82 (t, lH),
3.78-3.62 (m, 2H), 3.60-3.50 (m, lH), 3.04 (q, lH), 2.87 (s, lH), 2.86
(s, 2H), 2.80-2.60 (m, lH), 1.90 (bs, lH), 2.85-2.75 (m, lH), 1.82-1.60
(m, 3H), 1.55-1.45 (m, lH), 1.45 (s, 4H), 1.42 (s, 2H), 1.39 (s, 9H).
~tep C: N-[l(R)-[(1,2-Dihydro-l-methanesulfonylspiro[3H-indole-
3,4'-piperidiIl]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-
2-amino-2-methylpropanamide hydrochloride
To 0.637 g (0.101 mmol) of the intermediate from Step B
in S mL of dichloromethane was added 2.5 mL of trifluoroacetic acid
and stirred at RT for 30 min. The reaction mixture was concentrated to
an oil, taken up in 10 mL of ethyl acetate and washed with 8 mL of 10%
aqueous sodium carbonate solution. The aqueous layer was further
extracted with S mL of ethyl acetate. The combined organics were
washed with 10 mL of water, dried over m~n~.sium sulfate, filtered and
concentrated to give the 0.512 g of the free base as a white foam.
-
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- 26 -
To 0.512 g of the free base in 5 mL of ethyl acetate at 0~C
was added 0.2 mL of saturated hydrochloric acid in ethyl acetate and
stirred for l.S h. The white precipitate was filtered under nitrogen,
washed with ether, and dried to give O.S0 g of the title compound as a
white solid
lH NMR (400MHz, CD30D) Compound exists as 3:2 mixture of
rotamers. ~ 7.40-7.28 (m, 4H), 7.25-7.17 (m, 2H), 7.08 (t, 1/3H), 7.00
(t, 1/3H), 6.80 (d, 1/3H), 5.16 (ddd, lH), 4.60-4.42 (m, 3H), 4.05 (t,
lH), 3.90 (bs, 2H), 3.83-3.70 (m, 2H), 3.30-3.15 (m, lH0, 2.97 (s, lH),
2.95 (s, 2H), 2.90-2.78 (m, lH), 1.96 (t, 1/3H), 1.85-1.65 (m, 4H), 1.63
(s, 2H), 1.60 (s, 4H).
FXAMPLE 2
lS N-[l(R)-[(1,2-Dihydro-l-methanesulfonylspiro[3H-indole-3,4'-
piperdin]-l'-yl) carbonyl]-2-(phenyln~ethyloxy)ethyl]-2-amino-2-
methylpropanamide
~tep A: (2R)-[[[-2-(1,1-dimethylethoxy)carbonyl]amino]-2,2-
dimethyl- 1 -oxoethyl]amino-2-(phenylmethoxy)ethyl]- 1 -
propanoic acid allyl ester
Prepared from (2R)-2-[(1,1-dimethylethoxy)carbonyl]-
amino-3-(phenylmethyloxy)ethyl-propanoic acid and allyl alcohol by
carrying out the coupling reaction in CH2C12 in the presence of EDC
and DMAP.
lH NMR (400MHz, CDC13) ~ 7.25 (s, 5H), 5.8 (m, lH), 5.2 (dd, 2H),
5.0 (bs, lH), 4.7 (m, lH), 4.6 (m, 2H), 4.4 (dd, 2H), 3.9 (dd, lH), 3.6
(dd, lH), 1.45 (d, 6H), 1.39 (s, 9H).
Step B: (2R)-[[[-2-(1,1-dimethylethoxy)carbonyl]amino]-2,2-
dimethyl- l-oxoethyl]amino-2-(phenylmethyloxy)ethyl)- 1-
propanolc acid
To a stirred solution of the crude intermediate obtained in
Step A (6.7 g, lS.9 mmol), tetrakis (triphenylphosphine)-palladium (1.8
g, 0.1 eq) and, triphenyl phosphine (1.25 g, 0.3 eq) was added a solution
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- 27 -
of potassium-2-ethyl hexanoate (35 mL, 0.5M solution in EtOAc). The
reaction mixture was stirred at room temperature under nitrogen
atmosphere for lh and then diluted with ether (100 mL) and poured into
ice-water. The organic layer was seperated and the aqueous fraction
5 was acidified with citric acid (20%), then extracted with EtOAc. The
EtOAc extracts were washed with brine, dried over m~nesium sulfate,
filtered and evaporated to give the title compound as a solid.
lH NMR (400Hz, CD30D) ~ 7.3 (s, 5H), 4.7 (m, lH), 4.5 (s, 2H), 4.0
(m, lH), 3.6 (m, lH), 1.4 (d, 6H), 1.3 (s, 9H).
Step C: N-[1(R)-[(1,2-Dihydro-l-methanesulfonylspiro[3H-indole-
3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-
2-[(1,1 -dimethyl-ethoxy)carbonyl]amino-2-methyl-
prop~n~mide
To a solution of 1.0 g (3.44 mrnol) of l-meth~nPsulfonyl-
spiro[indoline-3,4'-piperidine] hydrochloride, 1.44 g (3.78 mmol) of
(2R)-[[-2-(1,1 -dimethylethoxy)carbonyl)amino]-2,2-dimethyl- 1 -oxo-
ethyl]-amino-2-(phenylmethyloxy)ethyl)-1-propanoic acid, N-methyl
morpholine (0.58 mL; 5.20 mmol), and l-hydroxybenztriazole (HOBT)
20 (0.58 g; 3.78 mmol), in 50 mL of dichloromethane was added EDC
(1.03 g; 5.20 mmol) and stirred at RT for 16h. The reaction mixture
was diluted with an additional 50 mL of dichloromethane and washed
with aqueous sodium bicarbonate solution (50 mL), dried over
anhydrous m~nesium sulfate, filtered, and concentrated. Flash
25 chromatography (50 g silica gel) of the crude oily residue gave 2.148 g
(90%) of the desired material as a colorless foam.
lH NMR (CDCl3, 400MHz) Compound exists as a 3:2 mixture of
rotamers ~ 7.40-7.10 (m, 6H), 7.06 (d, 1/3H), 7.02 (t, 1/3H), 6.90 (t,
1/3H), 6.55 (d, 1/3H), 5.15 (m, lH), 4.95 (bs, lH), 4.63 (bd, 1/3H),
30 4.57-4.40 (m, 2 2/3 H), 4.10 (bd, 1/3H), 4.00 (bd, 1/3H), 3.82 (t, lH),
3.78-3.62 (m, 2H), 3.60-3.50 (m, lH), 3.04 (q, lH), 2.87 (s, lH), 2.86
(s, 2H), 2.80-2.60 (m, lH), 1.90 (bs, lH), 2.85-2.75 (m, lH), 1.82-1.60
(m, 3H), 1.55-1.45 (m, lH), 1.45 (s, 4H), 1.42 (s, 2H), 1.39 (s, 9H).
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~tep D: N-[1 (R)-[(1,2-Dihydro- 1 -methanesulfonylspiro[3H-indole-
3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-
~-amino-2-methylprop~n~n-ide hydrochloride
To a solution of 2.148 g (3.41 mmol) of the intermediate
S from Step C in 10 mL of dichloromethane was added S mL of
trifluoroacetic acid and stirred for lh. The reaction mixture was
concentrated and basified with 100 rnL of 5% aqueous sodium carbonate
solution and extracted with dichloromethane (3XS0 mL). The combined
organics were washed with brine (S0 mL), dried over anhydrous
10 potassium carbonate, filtered, and concentrated to yield a colorless
foam. To a solution of the foam in 25 mL of ethyl acetate at 0~C was
added 4 mL of lM solution of hydrochloric acid in ethyl acetate. The
precipitate was filtered and washed first with ethyl ~cet~t~- and then with
ethyl acetate-ether (1:1), dried to yield 1.79 g (93%) of the title
15 compound as a colorless solid.
lH NMR (400MHz, CD30D) Compound exists as 3:2 mixture of
rotamers. ~ 7.40-7.28 (m, 4H), 7.25-7.17 (m, 2H), 7.08 (t, 1/3H), 7.00
(t, 1/3H), 6.80 (d, 1/3H), 5.16 (ddd, lH), 4.60-4.42 (m, 3H), 4.05 (t,
lH), 3.90 (bs, 2H), 3.83-3.70 (m, 2H), 3.30-3.15 (m, lH0, 2.97 (s, lH),
20 2.95 (s, 2H), 2.90-2.78 (m, lH), 1.96 (t, 1/3H), 1.85-1.65 (m, 4H), 1.63
(s, 2H), 1.60 (s, 4H).
EXAMPLE 3
25 N-[l(R)-[(1,2-Dihydro-1-methanesulfonylspiro[3H-indole-3,4'-
piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-methyl-
propan~mide mesylate
This compound was prepared by the treating the free base
obtained in Example 5, Step D, with methane sulfonic acid. The title
30 compound was obtained by recryst~lli7ing it from ethyl acetate-ethanol-
water. m.p. = 166~-168~C.
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~XAMPLE 4
O~,OBn
CO2H
Isonipecotic acid-N-benzyl carbamate (3)
Materials:
Isonipecotic acid (2) T.C.I. 4.02 kg (31.1 mol)
Benzyl chloroformate (Schweitzerhall) 6.91 kg (40.5 mol)
K2CO3 10.1 kg (72.9 mol)
Water 40.2 L
Isonipecotic acid (2) and K2CO3 were dissolved in 40.2 L
of water in a 100 L 4 neck flask with mechanical stirring under N2 and
the solution was cooled to 10~C. Benzyl chloroformate was ~ e~l~
m~int7inin~ the temperature between 9 and 14~C, and the mixt~lre was
15 warmed up to 22~C after the addition was complete and aged for 58 h.
The addition was completed in 4 h at which point the pH was 9Ø After
aging for 58 h there was no change in the pH.
The reaction mixture was transferred to a 200 L extractor
and washed with 3 x 13 kg (15 L) of IPAC and 1 x 12 L of EtOAc.
20 The aqueous layer was extracted with 8 L of toluene. After the washes
the benzyl alcohol content was reduced from 3.8% to 1.4% by HPLC
analysis. HPLC analytical: Dupont Zorbax 25 cm RXC8 column with
1.5 mL/min flow and detection at 254 nm; isocratic mixture with 35%
MeCN, 65% of 0.1% aqueous H3PO4; retention times: 3 = 6.9 min,
25 benzyl alcohol = 3.3 min, toluene = 17.3 min.
The aqueous phase was acidified with 37% aqueous HCl to
pH 1.8. Carbon dioxide was evolved during the addition of HCl, but gas
evolution was easily controlled. The addition of HCl took <1 h and
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required 10 L of conc. HCl. The aqueous phase was extracted with 3 x
6.6 L of toluene. The toluene extracts were dried with 2 kg of sodium
sulfate and filtered through a pad of SoLka-flocTM. The combined
filtrates weighed 17.8 kg. The crude yield of carbamate 3 was 7.89 kg
5 (97%) (as obtained by evaporation of weighed aliquots of the filtrates to
dryness). The filtrates were transferred through a 10 ,u inline ~llter to a
100 L flask. The extracts were concentrated at 10 mbar at <25~C to a
volume of 18 L. The final concentration of carb~m~te 3 was 440 g/L.
The concentration of the toluene filtrate served to azeotropically remove
10 final traces of water (final KF = 170mg/L). The product was 99.1 area
% pure with 0.9 area % benzyl alcohol as the only impurity.
EXAMPLE 5
o;~,OBn
~ 4
COCI
15 T~onipecotic acid chloride-lV-benzyl carbamate (4)
Materials:
Isonipecotic acid N-benzyl carbamate (3) 7.89 kg (30.0 mol) in
in toluene. (MW = 263.30) 17.9 L
Oxalyl chloride (MW = 126.93) 3.94 kg (31.0 mol)
DMF (MW = 73.10) 10 mL
Toluene 12 L
To the toluene solution of benzyl carbamate 3 from the
25 prece~lin~ step was added 5 rnL of DMF and 10 L of toluene. The
oxalyl chloride was added over a period of 20 min. The reaction
rnixture was aged for 16 h at 18~C under a slow stream of nitrogen.
HPLC analysis of the reaction mixture showed that 1.3% of the
carboxylic acid 3 still remained unreacted. The reaction mixture was
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warmed to 26~C, and S mL of DM~ were ~cle-1 The mixture was aged
for 2.5 h. A 1.0 mL aliquot of the reaction mixture was quenched with
5.0 mL of tert-butyl~mine and analyzed after evaporation by HPLC: 25
cm Dupont Zorbax RXC8 column at 50~C with 1 mL/min flow and
detection at 220 nm; isocratic 42% MeCN, 58% of 0.1% aqueous
H3PO4. This method showed that <0.05% of the acid 3 remained (as
judged by A) and showed >3 area % B (>1 mol% (COC1)2).
Oq,OBn
0~ N H-t-Bu
O N H-t-Bu
CONH-t-Bu
A B
The mixture was concentrated at 10 mbar and a
10 temperalurc~ of 20-25~C until S L of solvent had been removed.
The typical HPLC profile of concentrated toluene solution
after t-BuNH2 quench described above is as follows:
Retention time (min) Area % Identity
2.1 <0.5% carboxylic acid 3
7.8 <0.5% benzyl chloride
11.0 >99% Cbz-t-butylcarboxamide A
12.1 NA toluene
12.7 <0.5% ditert-butyloxamide B
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EXAMPLE 6
O~,OBn
CHO
Piperidine-4-carboxaldehyde-1-benzyl carbamate (5)
S ~tçri~l.c:
Isonipecotic acid chloride N-benzyl carbamate (4) 3.38 kg (12.0
mol)
in toluene (MW = 281.74) in 5.54 kg
DIEA (KF = 18 mg/L) 1.55 kg (15.0 rnol)
10% Pd/C (KF < 20 mg/g) 101 g
thioanisole (MW = 124.21, d = 1.058) 0.56 g
The DIEA and thioanisole were added to the solution of (4)
in toluene from the previous step and the catalyst was suspended in this
15 mixture. The mixture was immediately placed into the 5 gal autoclave
and hydrogenated at 20~C and 40 psi of H2. After 18 h the reaction had
taken up 70% the theoretical amount of hydrogen and HPLC analysis of
an aliquot that was quenched with tert-butylarnine indicated that 14.2
area % of acid chloride 2 remained. HPLC conditions same as above.
20 Retention time: 5 = 8.1 rnin.
A second charge of catalyst (101 g) and thioanisole (0.54 g)
were added as a slurry in 1375 mT toluene to the hydrogenator. After
23 h HPLC analysis of an aliquot that was quenched with tert-
butylamine indicated that 1.8 area % of acid chloride 2 remained. The
25 mixture was purged with nitrogen and the catalyst and precipitated
DIEA-HCl were removed by filtration through SoLka-flocTM. The filter
cake was washed with 10 L of toluene. The filtrates were trans~erred
through a 10 ~ inline filter to a 50 L extractor and washed with 2 x 7.2
L of 1 M aqueous HCl and 2 x 7.2 L of water. The mixture was
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concentrated at 10 mbar and a temperature of 25-30~C until 5 L of
residue remained.
Retention time (min) Area % Identity
2.1 <2 carboxylic acid 3
6.6 <1 dimer 21
8.1 >95 aldehyde 5
The assay yield of aldehyde 3 was 94% by HPLC analysis.
EXAMPLE 7
O~,OBn
~N~
~J 9
~N~H
CBZ-Spiroindoline (9)
Materials:
Piperidine-4-carboxaldehyde-1-benzyl 1.71 kg (6.89 mol)
carbamate (5) in toluene solution in 21.4 kg
Phenylhydrazine 900 mL, 981 g (9.15 mol)
Trifluoroacetic acid (TFA) 2.20 L, 3.26 kg (28.6 mol)
NaBH4 300 g, (7.93 mol)
Toluene 34.4 kg
MeCN 7.0 L
MeOH 7.0 L
The crude aldehyde 5 solution from the previous step was
transferred through a 10 ~ inline filter to a 100 L reactor equipped with
Teflon coated copper coils for cooling or heating and a mechanical
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- 34-
stirrer. Toluene (34.4 kg) and MeCN (7 L) were ~l(le~l, and the
resulting solution was cooled to 0~C. Phenylhydrazine was added in
portions and the temperature was m~int~ined at -1 to 3~C while nitrogen
was continuously bubbled through the reaction mixture.
S The phenylhydrazine was added until TLC and HPLC
analysis indicated complete con~u~ ion of the aldehyde 5 and the
appearance of a slight excess (cS%) of phenylhydrazine. TLC
conditions: Silica, E. Merck Kieselgel G60 F254 0.25 mm; diethyl
ether/pentane (4/1); and developing agent 0.5% ceric sulfate, 14%
ammonium molybdate in 10% aqueous sulfuric acid then heat; Rf:
aldehyde 5 = 0.52, phenylhydrazone 7 = 0.61, phenylhydrazine 6 =
0.21.
HPLC conditions: 25 cm Dupont Zorbax RXC8 colurnn at 30~C with 1.0
mL/min flow and detection at 254 nm; gradient schedule:
Time (min) acetonitrile:water
0 57:43
65:35
75:25
18 75:25
retention times: phenylhydrazine 6 = 4.5 min, toluene = 7.2 rnin,
phenylhydrazone 7 = 11.4 min.
The reaction mixture was aged for 30 min at 0-2~C, and
TFA was added m~int~ining the temperature between 2 and 7~C. The
25 reaction mixture was warmed to 50~C over 30 min, and m~int~inç-l for
17 h. The nitrogen sparge through the reaction mixture was stopped
and a slow stream of nitrogen was rnaintained over the reaction mixtllre.
During the first hour at 5~C the color gradually darkened to a deep
green, and a relatively small amount of a white crystalline precipitate
30 (ammonium trifluoroacetate) formed. After 17 h HPLC analysis (same
conditions as above) indicated that the reaction mixhlre contained 91.6
area % indolenine 8 and 1.5% of unreacted phenylhydra~one remained.
Aging the mixture for longer periods of tirne did not increase the assay
yield of indolenine 8.
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The reaction mixture was cooled to 12~C, and 7.0 L of
MeOH was added. NaBH4 was added in small (<20 g) portions
m~int~ining the temperature below 15~C. The addition took 30 min.
Moderate hydrogen evolution was observed ~hlrinf~ the addition, but it
S was easily controlled and there was virtually no frothing. Near the end
of the addition the color rapidly changed from green to brown and then
bright orange. A small amount (<200 mL) of a heavier phase had
separated (presumably aqueous salts). HPLC analysis (conditions as
before) indicated that all of the indolenine 8 had been consumed (90.4
10 area % CBZ-indoline 9); retention times: indolenine 8 = 7.5 min,
indoline 9 = 8.2 min. TLC: ethyl ether as solvent, ceric sulfate-
ammonium molybdate stain or 1% anisaldehyde stain; retention factors:
indolenine 8 = 0.18, CBZ-indoline 9 = 0.33.
The color change from green to orange corresponds very
15 closely to reaction end point. The quantity of NaBH4 required to
complete the reaction is heavily dependent on the tempe,aL~lle and rate
of addition of NaBH4, but the yield and quality of the product is
virtually unaffected provided that the reaction is complete. The reaction
mixtllre was cooled to 5~C over a period of 30 min. Then 8 L of 3%
20 aqueous NH40H (8 L) were added to bring the pH of the aqueous phase
to 7.4, the mixhlre was agitated, and allowed to settle. The tempela~u~e
rose to 15~C. The cloudy yellow lower aqueous phase was separated.
The organic phase was washed with 4 L of 3% aqueous NH40H, 2 x 4 L
of water, and 2 x 4 L of brine. The weight of the organic phase after
25 the w~.~hingc was 53.5 kg, and the assay yield was 94%.
The washed toluene solution was combined with the washed
organic phases of two other simil~rly processed reactions. The total
aldehyde used in the three reactions was 5.06 kg, (20.5 mol). The total
weight of CBZ-indoline 9 assayed in the combined organic phases was
30 5.91 kg, (18.3 mol, 90% assay yield). The combined organic phases
were dried with 5 kg of sodium sulfate, treated with 250 g of Darco
G60 carbon for 30 min, and filtered through Solka-flocTM. The filtrates
were vacuum concentrated at 10 mbar at <25~C until the residue was
near dryness. The solvent switch was completed by slowly bleeflin~ in
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30 L of IPAC and reconcentrating to 14 L at 200 mbar at 50-60~C. The
mixt,ure was heated to reflux in order to obtain a clear homogeneous
deep orange solution. lH NMR analysis indicated that the solut,ion
contained ca. 6 mol% of residual toluene after solvent switch.
S The solution was cooled to 68~C and seeded with 4 g of
crystalline CBZ-indoline 9. The solution was allowed to gradually cool
to 26~C over 6 h and aged for 9 h at 20-26~C. The slurry was cooled to
2~C over 1 h and aged at 2~C for lh. The product was isolated by
filtration, and the filter cake was washed with 2 x 2 L of 5~C IPAC and
2 x 2 L of 5~C MTBE. The product was dried in the vacuum oven at
30~C under a nitrogen bleed to give 4.37 kg (74%) of the title
compound 9 as a light tan crystalline powder. HPLC analysis of the
product indicated 99.5 area ~o purity. The mother liquor (11 L) and the
washes contained 1.15 kg (19%) of additional product 9 and ca 3% of
Cbz-isonipecotic acid phenylhydrazide (retent,ion time = 4.8 min).
EXAMPLE 8
O~,OBn
~ ) --
~,
SO2Me
CB7.-Spiroindoline-methanesulfonamide (1)
Materials:
CBZ-Spiroindoline (9) 1.69 kg (5.23 mol)
Meth~n~,sulfonyl chloride 599 g (5.23 mol)
Et3N (KF = 151) 635 g (6.27 mol )
THF (KF = 41) 12 L
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A 22 L flask was charged with the solid CBZ-spiloil-doline
9 and then 11.5 L of THF and the Et3N were transferred into the flask
through a 10 ~ inline filter. The resulting homogenous solution was
cooled to 0~C. A 1 L dlo~pillg funnel was charged with the
5 methanesulfonyl chloride and 500 mL of THF. The solution of the
MsCl in THF was added to the reaction mixture m~int~ining the
temperature between 0 and 4~C. The addition took S h and was
exothermic. ~ white precipitate, presumably triethylammonium
hydrochloride formed during the addition. HPLC analysis indicated that
10 the reaction was complete at the end of the addition (9 was
undetectable).
HPLC conditions: 25 cm Dupont Zorbax RXC8 column
with 1.5 mL/min flow and detection at 254 nm. Gradient Schedule:
Time (min) 0.1 % aq. H~4:MeCN
0 70:30
3 70:30
12 20:80
25 20:80
Retention times: 9 = 7.6 min, 1 = 13.6 min.
After the addition was complete the reaction mixture was
warmed to 18~C and aged for 16 h. There was no change in the
appearance of the reaction mixture, and HPLC profile between the end
25 of the addition and after the 16 h age. The reaction mixture was slowly
transferred over lh into a vigorously stirred solution of 30 L of water
and 200 mL of 37% aqueous HCl in a 50 L flask. The temperature in
the 50 L flask rose from 22 to 28~C. The product separated as a pale
tan gummy solid which changed to a granular solid. The aqueous
30 suspension was cooled to 22~C and aged for 1 h. The suspension was
filtered, and the filter cake was washed with 2 x 4 L of MeOH/water
(50/50). HPLC analysis indicated that <0.1% of the CBZ-Spiroindoline-
methanesulfonamidel was in the mother liquors.
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The filter cake was washed with 4 L of MeOH/water
(50/50) to which 50 mL of 28% aqueous NH40H had been ~ ecl The
filter cake was washed with 2 x 4 L of MeOH/water (50/50), and the
solid was dried in the vacuum oven at 50~C under a nitrogen bleed to
5 give 2.03 kg (97%) of the title product 1 as an of~-white powder.
HPLC analysis of the solids indicated 93.7 area % 1.
EXAMPLE 9
O~,OBn
~N
10 Optional Procedure for Isolation of Intermediate
CBZ-Spiroindolenine (8)
Materials:
Piperidine-4-carboxaldehyde-1-benzyl 12.37 g (0.050 mol)
carbamate (5)
Phenylhydrazine 5.41 g (0.050 mol)
Trifluoroacetic acid (TFA) 11.56 mL,17.10 g
(0.150 mol)
Methylene chloride 500 mL
The CBZ-aldehyde 5 was dissolved in dichloromethane in a
1 L flask equipped with Teflon coated magnetic stirring bar. The
resulting solution was cooled to 0~C. Phenylhydrazine was added via a
weighed syringe over 5 min and the temperature was m~int~ined at -1 to
25 3~C while nitrogen was continuously bubbled through the reaction
mixtllre. TLC and HPLC analysis indicated complete con~um~lion of
the CBZ-aldehyde 5 and the appearance of a slight excess (<2%) of
phenylhydrazine. TLC conditions: Silica, E. Merck Kieselgel G60 F254
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0.25 mm; diethyl ether/pentane (4/1); and developing agent 0.5% ceric
sulfate, 14~o ammonium molybdate in lO~o aqueous sulfuric acid then
heat; Rf: aldehyde 5 = 0.52, phenylhydrazone 7 = 0.61,
phenylhydrazine 6 = 0.21. HPLC conditions: 25 cm Dupont Zorbax
S RXC8 column at 30~C with 1.0 mLlmin flow and detection at 254 nm,
gradient schedule:
Time (min) acetonitrile:water
0 57:43
65:35
lS 75:25
18 75:25
retention times: phenylhydrazine 6 = 4.5 min, toluene = 7.2 min,
phenylhydrazone 7 = 11.4 min.
The reaction mixture was aged for 10 min at 0-2~C, and
lS TFA was added by syringe m~int~ining the temperature between 2 and
7~C. The reaction mixhlre was warmed to 35~C over 30 min, and
m~int~ined for 17 h. The nitrogen sparge through the reaction mixture
was stopped and a slow stream of nitrogen was m~int~ined over the
reaction mixture. During the first hour at 35~C the color gradually
20 darkened to a rosy pink then to a deep green, and a relatively small
amount of a white crystalline precipitate (ammonium trifluoroacetate)
formed. After aging for 17 h HPLC analysis (same conditions as above)
indicated that the reaction mixture contained 93 area % indolenine 8 and
cO.5% of unreacted phenylhydrazone remained. Aging the mixture for
25 longer periods of time did not increase the assay yield of indolenine 8.
The reaction mix~re was cooled to 10~C, and a mixture cont~ining 60
mL 28-30% ammonium hydroxide, 90 mL water and lS0 g crushed ice
was added with good stirring. The color of the mixture changed to a
salmon color. The organic phase was separated and washed twice with
30 400 mL water then 100 mL saturated aqueous NaCl. The organic phase
was dried over magnesium sulfate and filtered through a plug of 5 g of
silica. The filtrate was evaporated to give 15.84 g (99%) of indolenine
8 as a pale orange oil.
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EXAMPLE 10
O~,OBn
~ ~ 1
~,
SO2Me
Procedure for the Preparation of CBZ-Spiroindoline-
methanesulfonamide (1) without Isolation of Intermediate CBZ-
Spiroindoline (9)
Step 1: CBZ-Spiroindoline (9)
~te.ri~ls:
Piperidine-4-carboxaldehyde-1-benzyl 49.5 g (0.20 mol)
carbamate (5)
Phenylhydrazine (Aldrich) 23.7 g (0.22 mol)
Trifluoroacetic acid (TFA) 75.4 g (0.66 mol)
Toluene (KF < 250 mg/L) 654 mL
MeCN (KF < 250 mg/L) 13.3 mL
NaBH4 11.3 g, (0.30 mol)
Toluene 20 mL
MeOH 50 m1
A 2% (by volume) solution of MeCN in toluene was made
up using 654 mT of toluene and 13.3 mL of MeCN. In a 2 L 3 neck
flask equipped with a mechanical stirrer 617 m1 of the above solution
were degassed by passing a fine stream of nitrogen through the solution
for S min. Phenylhydrazine and TFA were added to the mixture while
still degassing.
The CBZ-aldehyde 5 was dissolved in the rest of the
solution prepared above (50 mL) and degassed by bubbling nitrogen
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through the solution while in the addition funnel. The solution in the
flask was heated to 35~C, and the aldehyde solution was slowly added to
the phenylhydrazine-TFA over 2 h. The mixture was aged at 35~C for
16h.
HPLC conditions: 25 cm Dupont Zorbax R~C8 column at
50~C with 1 mL/min flow and detection at 220 nm; isocratic 55%
MeCN, 45% of 0.1% aqueous H3PO4. Typical HPLC profile after 16 h
age:
Retention time (min) Area % Identity
1.6 0.1-0.5 phenylhydrazine 6
4.1 <0.1 dimer 21
4.7 c0.1 aldehyde 5
5.0 NA spiroindoline 9
6.3 NA toluene
6.9 97 spiroindolenine 8
10.3 <0.2 phenylhydrazone 7
2-3 tot. other impurities ~0.2% ea.
The rnixture was cooled to -10~C and MeOH was added. A
20 suspension of sodium borohydride in 20 mL toluene was added in small
portions (1 mL) over 30 min taking care that the temperature did not
exceed -2~C.
Area % Identity
0.1-1 phenylhydrazine 6
85-90 CBZ-spiroindoline 9
<0.1 CBZ-spiroindolenine 8
10-15 tot. other impurities (<3% ea.)
The temperature was raised to 10~C over lh, and 6%
30 aqueous ammonia (200 mL) was ~ lerl. The mixture was ~it~ted for
10 min, allowed to settle for another 10 min, and the lower aqueous
phase was drawn off. Acetonitrile (20 mL) and MeOH (20 mL) were
added to the organic phase and it was washed with 150 mL of 15%
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brine. The organic phase was found to contain a 92% assay yield of
CBZ-spiroindoline 9.
Step 2: CBZ-Spiroindoline-methanesulfonamide (1)
s
Materials:
CBZ-Spiroindoline (9) (MW = 322.51) (0.184 mol)
Methanesulfonyl chloride 21.1 g (0.184 mol)
DIEA (KF = 150 mg/L) 29.7 g, 40.1 mL
(0.230 mol)
THF (KF = 41 mg/L) 150 mL
The crude solution of CBZ-spiroindoline 9 solution from
Step 1 above was concentrated in a lL 3 neck flask (60-70~C, 150-200
15 Torr) until 250 g of residue remained. The THF and DIEA were
added, and the resulting homogenous solution was cooled to 0~C. A 125
mT~ dropping funnel was charged with the methanesulfonyl chloride and
50 mL of THF. The solution of MsCl in THF was added over 2 h to the
reaction mixture m~int~inin~ the temperature between 0 and 4~C and the
20 mixture was aged for 2 h at 5-8~C. The addition was slightly
exothermic. A white precipitate, presumably DIEA-hydrochloride,
formed during the addition. HPLC conditions were the same as above.
HPLC analysis indicated that the reaction was complete 1 h after the end
of the addition (9 was undetectable) and the assay yield was 94% from
25 9. Retention time: 1 = 7.8 min. Typical HPLC profile of reaction
mixtllre after 2 h age:
Area % Identity
<0.1 CBZ-spiroindoline 9
90-92 CBZ-sulfonamide 1
8-10 tot. other impurities (<2% ea.)
The mixture was warmed to 20~C, and 200 mL of lM
aqueous HCl was added. The mixture was warmed to 50~C, and the
aqueous phase was separated. The organic phase was washed sequentialy
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with 100 mL water, 100 mT 5% aqueous sodium bicarbonate, and 100
mL water. The organic phase was transferred to a 1 L 3 neck flask
equipped ~or mechanical stirring and distillation. The mixt~lre (ca 400
mL) was distilled at atmospheric pressure until 150 mL of distillate had
5 been collected. The head temperature reached 107~C; the pot
tell~eldLulc was 110~C. The distillation was continued with continuous
addition of n-propanol at such a rate as to m~int~in a constant volume
(ca 350 mL) in the pot. The distillation was stopped when a total of 525
mL of n-PrOH had been added and a total of 800 mL of distillate had
10 been collected.
The temperature of both the head and pot rose from 94~C
to 98~C during the solvent switch. Toluene and n-PrOH form an
azeotrope boiling at 97.2~C composed of 47.5% toluene and 52.5% n-
PrOH. The mixture was allowed to cool gradually to 20~C over 3h and
15 aged for 12 h. The mother liquor was found to contain 2% toluene and
4 mg/mL of sulfonamide. The solubility of the sulfon~mitle in various
mixtllres of toluene and n-PrOH has been determined by HPLC assay:
%toluene in n-PrOH solubility of 1 in m~/mL
0 2.36
3.02
4.23
7.51
10.3
The crystalline slurry was filtered and washed with 3 ~ 100
mL of n-PrOH. The product was dried in a vacuum oven at 50~C with a
nitrogen bleed for 16 h to furnish 65.5g (82 % from aldehyde 5) of 6 as
a tan solid with 93.5 wt% purity.
Typical HPLC profile of solid:
Area % Identity
<0.1 CBZ-spiroindoline 9
>99 CBZ-sulfonamide 1
~1 tot. other impurities (<0.2% ea.)
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For additional purification, a 40.0 g sample of the n-PrOH
crystallized sulfonamide was dissolved in 134 mL of EtOAc at 60~C and
treated with 8.0 g of Darco G-60 carbon for 1 h at 60~C. After the
5 addition of 2.0 g SolkaflocTM, the slurry was filtered through a pad of
4.0 g SoLkafloc~M, and the pad was washed with 90 mLof EtOAc at
60~C. Prior to the addition of the carbon the solution was a brown
color. The filtration proceeded well without plugging to give a golden
yellow filtrate. The filtrate was distilled at atmospheric pressure in a
500 mL flask (pot temperature 80-85~C) until 100 g (100 mL) of
residue remained. This solution was allowed to cool to 35~C over 3 h.
Over a lh period, 116 mL of cyclohexane was added with good
agitation at 35~C. The mixtllre was cooled to 20~C over 1 h and aged at
20~C for 12 h. At 35~C much of the sulfon~ 1e has cry~t~11i7e-1 out
15 and the mixture was thick. Addition of cyclohexane at 20~C makes
agitation difffcult. After the aging period, the supern~t~nt was found to
contain 2.5 mg l/g. The crystalline slurry was ~lltered and the cake was
washed with 77 mL of 2:1 cyclohexane-EtOAc and 2 x 77 mL of
cyclohexane. The product was dried in a vacuum oven at 50~C with a
20 nitrogen bleed for 16 h to furnish 34.2 g of 1 (MW = 400.3) as a white
crystalline solid (85 ~o recovery from crude 1, 70 % from 5 with >99.9
wt % purity).
EXAMPLE 1 1
H
~N~ HCI
1a
~C N~
SO2Me
H~l Salt of Spiroindoline-methanesulfonamide (la)
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Materials:
CBZ-spiroindoline-methanesulfonamide (1) 941 g (2.35 mol)
Pearlman's catalyst 20% Pd(oH)2/c 188 g
THF 8 L
MeOH 7 L
The catalyst was suspended in 7 L of MeOH and transferred
into the 5 gal autoclave followed by the solution of 1 in 8 L of THF.
The mixture was hydrogenolyzed at 25~C at 80 psi of H2. After 2.5 h
the temperature was raised to 35~C over 30 min.
HPLC analysis indicated complete con~u~ ion of Cbz-
~iroindoline-methanesulfonamide. HPLC conditions: 25 cm Dupont
Zorbax RXC8 column with 1.5 mL/min flow and detection at 254 nm.
Gradient Schedule:
Time (min) 0.1% aq. H~PO4:MeCN
0 70:30
3 70:30
12 20:80
25 20:80
retention times: Spiroindoline = 7.6 min,
Cbz-spiroindoline-methanesulfonamide = 13.6 min.
The mixture was purged with nitrogen and the catalyst was
removed by filtration through SoL~a-flocTM while still warm. The
catalyst was washed with 4 L of THF and 2 L of MeOH. The pale
yellow filtrates were concentrated to a thick oil at 10 mbar and <25~C.
The solvent switch was completed by slowly bleeding in 15 L of EtOAc
and reconcentrating to dryness. The residue solidified to a hard off-
white mass. MeOH (1.5 L) was added and the mixture was heated to
70~C to give a homogenous solution. While the solution was at 70~C,
10.5 L of EtOAc at 20~C was added. The temperature fell to 40~C, and
the mixture remained homogenous.
Subsequent experiments suggested that it is more convenient
to solvent switch the MeOH-THF filtrates to MeOH, concentrate to the
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desired volume, and then add the EtOAc. This avoids the solidi~lcation
of the residue upon concentration of the EtOAc solution.
Hydrogen chloride diluted with about an equal volume of
nitrogen was passed into the solution. The temperature rose to 60~C
5 over the course of 15 min, and a white precipitate of the hydrochloride
salt formed. Diluting the HCl with nitrogen only avoids the reaction
mixture sucking back and may not be necessary.
The mixture was cooled in an ice bath, and the hydrogen
chloride addition was continued for lh. The temperature gradually fell
10 to 20~C. The suspension was aged for 2 h while the temperature was
lowered to 10~C. The crystalline product was isolated by filtration, and
the filter cake was washed with 3 L of EtOAc. It was dried in the
vacuum oven at 35~C to give 1.18 kg (86%) of the title product la as an
off-white crystalline solid of >99.5 area % purity by HPLC analysis.
15 HPLC conditions: 25 cm Dupont Zorbax RXC8 column with 1.5
mlJmin flow and detection at 230 nm; isocratic 35% MeCN, 65% of
0.1% aqueous ammonium acetate. Retention time: la - 5.4 min.
FXAMPLE 12
H
I
~N~
~ J 1b
N'SO2Me
~piroindoline-methanesulfonamide (Free base form) (lb)
A 250 mL aliquot of the filtrate from the Cbz-
hydrogenolysis cont~inin~ 4.67 g of lb (free base) was concentrated to
ca 10 mL. The residue was dissolved in 20 rnL of EtOAc and the
25 solution was reconcentrated to ca 10 mL. This was repeated once more,
and 10 mL of EtOAc was added to the residue. A crystalline precipitate
began to form. MTBE (20 mL) was added in one portion. Additional
crystalline solid precipitated. but the supernatent still contained a
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subst~nti~l quantity of dissolved product which did not precipitate on
st~n~1ing. Hexanes (70 rnL) were added ~kv~wise over 2 h to the
mixhlre with vigorous stirrin~:. The slow addition of the hexanes is
neccessary to avoid the oiling out of the amine.
The ~git~ted mixture was aged for lh and filtered. The
filter cake was washed with 20 mL of 1: 1 MTBE-hexanes and then with
20 mL of hexanes. The product was dried under a stream of nitrogen to
give 3.86 g (82%) of the free amine of lb as an off white cryst~llin~
solid of >99.5 area % purity. HPLC conditions: 25 cm Dupont Zorbax
RXC8 column with 1.5 rnL/min flow and detection at 230 nm; isocratic
35% MeCN, 65% of 0.1% aqueous ammonium acetate. Retention time:
lb = 5.4 min.
EXAMPLE 13A
H
I
N'S~2~ke
Spiroindoline-methanesulfonamide (Free base form) (lb)
Materials:
CBZ-Spiroindoline-sulfonamide (1)833.5 gr (2.08 mol)
Pd(OH)2/C (20% weight of Pd(OH)2) 124.5 (15%)
THF 6.5 L
MeOH 19.5 L
NH4OH (conc) 60 mL
The hydrogenation was run three (3) times due to
equipment limitations; this procedure refers to a single run. The CBZ
spiloilldoline sulfonamidel was dissolved in THF (6.5 L, KF = 53
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,ug/,uL) and then MeOH (KF=18 ~Lg/mL, 4L) was added followed by
addition of the catalyst and the slurry was transferred to a 5 gal
autoclave. The rem~incler of the MeOH (2.5 L) was used for rinsing.
The mixture was heated to 40~C at 50 psi for 24 hours. The catalyst
5 loading and reaction time are a function of the purity of starting
material 1. This m~teti~l was unique requiring 2 15% catalyst and long
reaction time. Purer batches of spiroindoline required only 5% of
catalyst and 4-6 hrs reaction time.
Upon completion (<0.1 A% 1 by LC) the mixture was
10 hltered thru SoLka FlocTM and the carbon cake washed with MeOH (13
L) cont~inin~ NH40H (0.5%, 60 mL). The combined filtrates (assay
shows 1587 g of spiroindoline amine lb) were concentrated in vacuo
and the resulting solids were partitioned between 40 L (of toluene:THF
(3:1) and O.5N NaOH (18 L). Although the layers separated easily a
15 heavy precipitate could be seen in the aqueous layer. The aqueous
suspension was thus extracted with CH2C12 (15 L). The aqueous and
organic layer separated slowly. Prior to CH2Cl2 addition THF was
added to the aqueous layer along with enough NaCl to saturate the layer.
However dissolution of the product was not achieved which necessitated
20 the use of CH2C12.
The combined toluene, THF and CH2C12 layers were
combined and concentrated in the batch concentrator The residue was
flushed with 7 L of CH3CN. Finally 10 L of CH3CN were added and
the solution stood overnight under N2 atmosphere.
FXAMPLE 13B
H
I
~N~
lb
~ SO2Me
Spiroindoline-methanesulfonamide (Free base form~ (lb)
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- 49 -
Materials:
CBZ-Spiroindoline-sulfonamide (1) 3 kg (7.49 mol)
Darco G-60 600 g
Ethyl Acetate 36 L
Absolute Ethanol 189 L
10% Pd/C 450 g
Ammonia Solution 500 rnl
SoL~ca FlocTM 2.5 kg
Isopropyl Acetate 65 L
A mixture of CBZ-spiroindoline (1) (1 kg) and Darco G-60
(200 g) in ethyl acetate (9 L) was stirred and heated at 60-65~C under a
nitrogen atmosphere for 8 hours. The Darco was removed by filtration
at 60-65~C, the solid washed with hot ethyl acetate (3 L) and the filtrate
and w~hin.~s combined. LC wt/wt assay con~ l negligible loss to
the Darco. The ethyl acetate solution was evaporated to dryness in
vacuo using a 20 L Buchi apparatus and then flushed with absolute
ethanol (2 x 5 L). This material was then slurried in absolute ethanol (8
L) warmed to 65-70~C and placed in the 20 L autoclave. The batch was
rinsed in with absolute ethanol (1 L). A slurry of 10% Palladium on
charcoal (75 g, 7.5% by weight) in absolute ethanol (750 ml) was then
added to the autoclave and rinsed in with a further portion of absolute
ethanol (250 ml).
The batch was hydrogenated at 65~C with vigorous stirring
under 40 psi hydrogen pressure for 3 hours, a second portion of 10%
palladium on charcoal (75 g) was ~ e~l~ the batch was hydrogenated for
a further 2 hours and then sealed overnight. The batch was transferred
(still hot, 60-65~C) to a 20 L Buchi apparatus and degassed in vacuo to
remove formic acid by "feeding and bleeding" absolute ethanol (18 L
total).
This procedure was repeated twice more and the three
batches were combined in a 10 gallon glass-lined vessel and the
combined batch was degassed again by the addition and distillation (in
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vacuo) of absolute ethanol (2 x 10 L). SoLka flocTM (0.5 kg) was added
to the batch and rinsed in with ethanol (10 L). An Estrella filter was
loaded with SolkaflocTM (2 kg) as a slurry in ethanol (20 L). The
resulting mixture was warmed to 60-65~C and then transferred at this
S temperature via heated filter using pump to two tared stainless-steel
bins. The initial vessel, the filter, the pump and the lines were rinsed
with a hot (60-65~C) mixture of aqueous ammonia (500 ml) in absolute
ethanol (25 L). The filtrate and w~.~hin~;.s were combined in the two
stainless-steel bins.
The batch was then transferred to a vessel using an in-line
~llter cont~ining a 10 micron cartridge, and then concentrated in vacuo
to low buLk (~15 L). The ethanol was replaced by isopropyl acetate by
the "feeding and blee-lin~" of 3x batch volumes of isopropyl acetate (45
L total), while m~int~ining a batch volume of ~15 L. The solvent
switch, when complete, contained <1% residual ethanol by GC. The
batch was then diluted to ~33 L by the addition of isopropyl acetate (20
L), and this solution of spiroindoline-amine lb (1.855 kg by LC
analysis) in isopropyl acetate was used for the next stage of the process.
FXAMPLE 14A
H
i~
~N'SO2Me
Boc-O-Benzylserine Spiroindoline (11)
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Materials:
Spiroindoline-amine (lb) 1587 g (5.966 moles)
Amino acid (10) 1938 g (6.563 moles)
Ph~O CO2H
NHBOC
DCC 1334.5 g (6.563 moles)
HOBT 884 g (6.563 moles)
CH3CN 25 L
0.5N NaOH 18 L
0.5N HCl 18 L
NaHCO3 sat. 18 L
iPrOAc 28 L
The spiroindoline-aminelb in CH3CN or iPrOAc:H2O (25
L) at ambient temperature under N2 was treated in sequence with HOBT
(884 g; 1.1 eq) as a solid, DCC (1334.5 g, 1.1 eq) as the melt (heating in
hot water at 60~C for ca. 1 hr) and ~mally the amino acid 10 (1938 g) as
the solid. The mixture was stirred for 3 hr upon which time heavy
precipitation of DCU occurred and LC analysis showed ca. 0.5 A% of
amine lb rem~inin~. IPAc (9 L) was ~-lcle-l, the slurry was filtered
20 through SoLka FlocTM and the cake was washed with IPAc (19 L). The
combined organic solution was washed in sequence with 0.5N NaOH (18
L), 0.5N HCl (18 L) and saturated NaHCO3 (18 L). A final water wash
at this point resulted in an emulsion and was thus elimin~ted.
The organic layer was concentrated in vacuo and the
25 residue was dissolved in MeOH or EtOH (10 L final volume). Assay
yield 3026 gr (89%).
The use of alternative peptide coupling agents such as
carbonyldiimidazole or formation of mixed anhydrides, such as sec-
butyl carbonate, gave inferior yields of 11 and/or 14 with a high degree
30 of epimerization in the case of the former compound. Other peptide
coupling reagents were prohibitively expensive.
CA 02234817 1998-04-08
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l~XAMPLE 14B
H
1~ 11
~N~
SO2Me
B~c-Q-E~enzylserine Spiroindoline (11)
S Materials:
Spiroindoline-amine (lb) 1.855 kg (6.96 mol)
Isopropyl acetate 29 L
Dicyclohexylcarbodiimide (DCC) 1.58 kg (7.65 mol)
1-Hydroxybenzotriazole (HOBt) 1.03 kg (7.62 mol)
N-Boc-O-benzyl-D-Serine 2.26 kg (7.65 mol)
lM Aqueous sodium hydroxide 26 L
0.5M Aqueous hydrochloric acid 26 L
Satd. Aqueous sodium hydrogen carbonate 26 L
Absolute Ethanol 50 L
Water (20 L) was added to a stirred solution of the
spiroindoline-aminelb (1.855 kg) in isopropyl acetate (33 L) in a
reaction vessel. The following chemicals were then added sequentially
at room tempelalule under a nitrogen atmosphere: DCC (1.58 kg, 1.1
20 equivs.), HOBt (1.03 kg, 1.1. equivs.) and finally N-Boc-O-benzyl-D-
Serine (2.26 kg, 1.1 equivs.). The reagents were rinsed in with
isopropyl acetate (7 L). The batch was stirred at room temperature
under nitrogen atmosphere for 5 hours when LC showed the ratio of
product/starting material to be 99.4/0.6. The mixture was then filtered
25 through an Estrella filter using cloth and cardboard only and lltili7.in~ a
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- 53 -
pump into another vessel. The sending vessel was rinsed with isopropyl
acetate (22 L) and this was used to rinse the filter, the pump and the
lines into the receiving vessel. The 2-phase mixture in the vessel was
stirred for 10 minutes and then allowed to settle for 15 minlltes. The
5 lower aqueous layer was separated off and the organic solution was left
to stand at room temperature overnight.
The next day, the organic solution was washed with lM
aqueous sodium hydroxide solution (26 L) then O.5M aqueous
hydrochloric acid (26 L) and finally saturated aqueous sodium hydrogen
10 carbonate (26 L). LC analysis gave an assay yield of 3.787 kg, 93%
overall yield from 7.49 moles (3 kg) of starting CBZ-spiroindoline (1).
The batch was concentrated in vacuo (internal temperature = 13-15~C.
jacket temperature = 40~C, Vacuum = 29") to low bulk (~15 L) and
solvent switched to ethanol by "feeding and bleeding" ethanol (50 L)
15 whilst m~int~inin~ the volume at ~15 L. GC showed <1% iso~lo~yl
acetate rem~ining. This solution was used for the next stage of the
process.
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EXAMPLE 15A
~ NH2
~ ~0
12
~'
SO2Me
O-Benzylserine Spiroindoline (free base form) (12)
Materials:
Boc-O-Benzylserine Spiroindoline (11) 3026 g (5.57 moles)
Methane sulfonic acid (MsOH) 1.16 L (17.9 moles)
MeOH 10 L
iPrOAc 24 L
0.5NNaOH 35L
The Boc-O-benzylserine spiroindoline 11 in 10 L of
MeOH (or EtOH) was treated with neat MsOH (1.16 L~ added over ca.
30-40 min, (initial temperature 16~C, final temperature 28~C). The
15 dark red solution was aged overnight under N2. The mixture was then
pumped into a 100 L extractor contslinin~ 24 L iPrOAc and 35 L 0.5 N
NaOH. The pH of the aqueous layer was 7. NaOH (6M) was added
until pH > 10.5. As the pH increased the color changed from red to
yellow. The layers were separated and the organic layer (24 L) was
20 shown by NMR to contain 13 mole % of MeOH in iPrOAc [5 volume
%]. LC assay 2.48 kg.
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PCT~US96/17196
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- 55 -
FXAMPLE 15B
H
~o ~c~ NH2
~ ~0
~C'
SO2Me
O-Benzylserine Spiroindoline (free base form) (12)
Materials:
Boc-O-Benzylserine Spiroindoline (11) 3.787 kg (6.96 mol)
Meth~nt-.sulphonic acid 2.006 kg (20.87mol)
Isopropyl acetate 38 L
~lM Aqueous sodium hydroxide 16 L
50% Aqueous sodium hydroxide 1.6 L
Methanesulphonic acid (2.006 kg, 1.355 L, ~3 equivs.) was
added to the stirred solution of Boc-O-benzylserine spiroindoline (11)
(3.787 kg) in ethanol (total volume ~15 L) in a reaction vessel. The
15 batch was warmed to 35-40~C. After 7 hours, LC showed the absence
of starting material and the reaction was allowed to cool to room
temperature overnight. The next day, water (44 L) was added to the
batch with stirring. The batch was cooled to ~5~, stirred for 30 minlltes
and then filtered through an in-line filter (loaded with a 10~ cartridge)
20 into a bin. The batch was then sucked back into the vessel. A water
rinse (10 L) was used to rinse the vessel and lines into the bin and this
was used to then rinse back into the vessel. Isopropyl acetate (38 L) was
added followed by a lM aqueous sodium hydroxide (16 L). The batch
was cooled to 10-15~C, the pH of the lower aqueous layer was
25 confirmed as ~7 and 50% aqueous sodium hydroxide solution was added
(1.6 L) (pH >10). The batch was stirred at 10-15~C for 25 minutes and
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- 56 -
then allowed to settle for 10-15 minlltes. The lower aqueous layer was
separated (78.1 kg). LC assay indicated 28.4 g of 12 (0.85% of theory)
contained in the aqueous liquors. Volume of the organic solution = 51
L. LC assay indicated 3.057 kg, 92% overall yield from 3 kg,
5 7.49 moles of CBZ-spiroindoline sulfon~mi(l~ (1). This solution was
used for the next stage.
FXAMPLE 16A
H N ~ BOC
14
SO2Me
10 Roc-Aminoisobutyryl O-Benzylserine Spiroindoline (14)
Materials:
Spiroindoline amine (12) 2481 g (5.57 moles)
amino acid peptide (13) 1247.1 g (6.16 moles)
CH~<CH3
15 HO2C NH-BOC
DCC 1266.7 g (6.16 moles)
HOBT 827 g (6.16 moles)
IPAc 52 L
H2O 37 L
0.5NNaOH 36 L
0.5N HCl 36 L
Sat. NaHCO3 36 L
CA 02234817 1998-04-08
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The solution of the amine 12 in IPAc was tlil~lte-l to a total
volume of 39 L with IPAc and 37 L of H2O was added. The biphasic
mixtllre was then treated in sequence with HOBT (827 g) as a solid,
DCC (1266.7 g) as a melt, and amino acid 13 at ambient temperature
5 under nitrogen. The reaction mixture was stirred for 2 h upon which
time LC analysis indicated dissappearance of the starting material 12
(<0.3 A%). The mixture was filtered through SoL~a FlocTM and the
solids were washed with 13 L of IPAc. The material may be stored at
this point as a biphasic mixture overni~ht
The mixture was transferred to a 100 L extractor, the
aqueous layer was separated and the organic layer was washed
successively with 36 L of 0.5N NaOH, 0.5N HCl and saturated NaHCO3.
Assay yield 3160 g (81% from spiroindoline + 5% for volume
measurement error). The solution was concentrated to a small volume
15 and was flushed with ethanol (2 x 4 L). If desired, the inermediate
compound 14 may be isolated by ~ 1in~ water to crystalize it out.
I~e use of alternative peptide coupling agents such as
carbonyldiimidazole or formation of mixed anhydrides, such as sec-
butyl carbonate, gave inferior yields of 14 with a high degree of
20 epimenzation. Otherpeptide coupling reagents were prohibitively
expenslve.
EXAMPLE 16B
H H CH3 CH3
~~ i=o O H
14
~,
N~
SO2Me
Boc-Aminoisobutyryl O-Benzylserine Spiroindoline (14)
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Materials:
Spiroindoline arnine (12) 3.057 kg (6.89 mol)
Dicyclohexylcarbodiimide (DCC) 1.56 kg (7.56 mol)
l-Hydroxybenzotriazole (HOBt) 1.02 kg (7.55 mol)
Boc-2-Aminoisobutyric acid (13) 1.54 kg (7.58 mol)
Isopropyl acetate 32 L
lM Aqueous sodium hydroxide 38 L
0.5M Aqueous hydrochloric acid 38 L
Satd. aqueous sodium hydrogen carbonate 38 L
10 Absoluteethanol 45 L
Water (49 L) was added to the stirred solution of the
~hoilldoline an~ine 12 (3.057 kg) in isopropyl acetate (total volume
~51 L) in a reaction vessel at room temperature under a nitrogen
15 atmosphere. The following chemicals were then added sequentially:
DCC (1.56 kg, ~1.1 equivs.), HOBt (1.02 kg, ~1.1 equivs.) and finally,
N-Boc-2-aminoisobutyric acid 13 (1.54 kg, ~1.1 equivs.). The mixtllre
was stirred vigorously at room tempeLaLule for 2 hours when LC
showed the reaction to be complete. The rnixture was filtered to to
20 another vessel via an Estrella filter using a pump. Isopropyl acetate (22
L) was used to rinse vessel, the filter, the pump and the lines into the
receiving vessel. The 2-phase mixt~lre was then stirred for 5 minutes
and the layers were allowed to separate. The lower aqueous layer was
separated without incident (weight of aqueous liquors = 51.1 kg). The
25 organic solution was then washed sequentially wi~ lM aqueous sodium
hydroxide (38 L), 0.5M aqueous hydrochloric acid (38 ~) and finally,
saturated aqueous sodium hydrogen carbonate (38 L) without incident.
The organic solution was then transferred using a pump via
an in-line filter (cont~ininp a 10,u cartridge) to another vessel for the
30 solvent switch to ethanol. The vessel was rinsed with isopropyl acetate
(10 L) and this was used to rinse the pump, the filter and the lines into
the receiving vessel. The filtrate and w~ching.s were combined. Total
volume = 75 L (by dipstick). LC assay gave 4.395 kg of Boc-
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_ 59 _
aminoisobutyryl O-benzylserine spiroindoline (14), i.e. 93% overall
from 7.49 moles of starting CBZ-spiroindoline sulfonamide (1).
The batch was concentrated in vacuo to low buLk (~lS L)
and the isopropyl acetate switched to ethanol by "feeding and bleeding"
5 absolute ethanol (45 L total). At the end of the solvent switch, GC
showed <1% isopropyl ~ret~te rem~inin~;. This solution (25 L)
cont~ining 4.395 kg of 14 was used for the next stage. If desired, the
inermediate compound 14 may be isolated by adding water to crystalize
it out.
EXAMPLE 17A
H H CH3~cH3
3--~~C~ ~ NH2
~N~
SO2Me
Aminoisobutyryl O-Benzylserine Spiroindoline (15)
Materials:
Boc Spiroindoline (14) 3160 g (5.03 moles)
Methanesulfonic acid (MsOH) 979 mL (lS.l moles)
EtOH 6.2 L
H2O 30 L
lNNaOH ll L
EtOAc 26 L
Darco 60 activated carbon l Kg
The Boc spiroindoline 14 was dissolved in 6.2 L of EtOH
and treated with MsOH (979 mL). The temperature rose from 20 to
30~C and the reaction was allowed to proceed overnight. After 12
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- 60 -
hours at 20~C there was still 15 A% of starting material le~t so the
mi~ture was heated to 35~C for 6 hours. Upon completion (<0.1 A%
~L) the reaction was cooled to 20~C and 30 L of H20 were added and
the solution was filtered through a glass funnel with a polypropylene
S ~llter to ~llter off residual DCU. The mixture was transferred to a 100
L extractor and 26 L of EtOAc were added. The aqueous layer was
basi~led via addition of chilled lN NaOH (11 L) and 1 L of 50~o NaOH.
Addition of ice was required to keep the temperature below 14~C.
Higher temperatures resulted in significant emulsion problems.
The organic layer was distilled at 50~C at ca. 21" of Hg
until KF <1000 ,ug/rnL. Lower KF's result in more ef~lcient carbon
treatments and better recovery at the salt formation step. KFs of 160
,ug/mL were achieved at the 700 g scale. The solution was diluted with
ethyl acetate to a total volume of 31 L (LC assay 2.40 kg). Activated
15 carbon (Darco G-60) was added and the mixtllre was stirred for 24 h.
The mixture was filtered through SoLka FlocTM and the filter cake was
washed with ethyl acetate (16 L), assay 2.34 Kg.
~XAMPLE 17B
H H CH CH3
O--~~ i= ~NH2
~N~
~J 15
N'SO2Me
~ninoisobutyryl O-Benzylserine Spiroindoline (15)
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Materials:
Boc Spiroindoline (14) 4.395 kg (6.99 mol)
Meth~nPsulfonic acid 2.017 kg (20.99 mol)
Ethyl acetate 185 L
lM Aqueous sodium hydroxide 16 L
50% Aqueous sodium hydroxide 2.6 L
Darco G-60 900 g
SoL~a FlocTM 2.5 kg
Methanesulfonic acid (2.017 kg, 1.36 L, ~3 equivs.) was
added to the stirred solution of the Boc spiroindoline 14 (4.395 kg) in
ethanol (total volume ~25 L) in a reaction vessel at room temperature.
The batch was warmed to 35-40~C, and stirred overnight. On the next
day, the batch contained ~1.1 A% of starting m~teri~l and so the
reaction was continued for a further 4 hours, then LC showed ratio of
product/ starting material to be 99.6/0.4. The batch was concentrated in
vacuo to ~15 L volume and then diluted with water (44 L). The batch
was cooled to 5~C, stirred for 30 minutes and then filtered through a
Sparkler in-line filter (cont~ining a 10,u cartridge) using a pump to
another vessel to remove a small amount of residual DCU.
The vessel, the pump, the filter and the lines were rinsed
with water (10 L), and this was added to the vessel. Ethyl acetate (36 L)
was added to the vessel and the stirred mixture was cooled to 10~C. A
solution of cold (5-10~C) lM aqueous sodium hydroxide solution (16 L)
and cold (5-10~C) 50% aqueous sodium hydroxide solution (2.6 L) were
added at 10~C and the temperature rose to 14~C. The resulting mixture
was stirred for 15 minutes at <14~C and then the lower aqueous layer
separated off.
The batch was concentrated in vacuo to ~20 L volume and
then a mixture of ethyl acetate (35 L) and ethanol (5 L) was fed in while
m~int~ining the volume at ~20 L. At the end of this distillation the KF
- was 9160 mgml~l. The batch was solvent switched to ethyl acetate by"fee-linp and bleeding" ethyl acetate (40 L total). At the end of this
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distillation, K~ was 446 mgml~l. The batch was diluted with ethyl
acetate (10 L).
Darco G-60 (900 g) was added to the hazy mixture. This
was rinsed in with ethyl acetate (6 L). This rnixture was stirred at room
5 tempel~Lule overni~ht Next day, Solka FlocTM (0.5 kg) was added to
the stirred batch in the vessel and then SoLka FlocTM (2.0 kg) was stirred
in a little ethyl acetate and loaded into an Estrella filter . The excess
solvent was pumped away through a Sparkler in-line filter cont~inin~ a
lO,u cartridge. The slurry was transferred from the vessel through a
10 flter using a pump and then through another filter to 2 x 40 L stainless
steel bins. Visual inspection showed the liquors to be clear and clean.
The vessel was rinsed with ethyl acetate (22 L) and this was used to
rinse through the route outlined above tc) the stainless steel cans. The
contents of both cans was transferred into a reaction vessel and the
15 solution was rnixed thoroughly.
The batch (58 L) had a KF of 2950 mgml~l and so was
redried by concentrating in vacuo to 20-25 L volume. The batch was
diluted to 46 L volume (dipstick) by the addition of ethyl acetate (25 L).
The KF was 363 mgml~l. The batch was ~ t~.~l to 62 L volume by the
20 addition of ethyl acetate (17 L) and was used for the final stage of the
process.
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EXAMPLE 18A
H H CH3~cH3
3--~ ~ ~ NH2- CH3SO3H
~N~ 16
~ 'SO2Me
Spiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyl-
oxy)ethyll-2-amino-2-methylpropanamide Methanesulfonate (16)
s
Materials:
Amine (15) 2340 g (4.43 moles)
Methane sulfonic acid (MsOH) 316 mL (4.88 moles)
EtOAc 60 L
EtOH 4.8 L
8% EtOH in EtOAc 20 L
The volume of the solution of 15 from the previous step
was adjusted to 60 L with ethyl acetate and EtOH (4.8 L) was ~ e~l~
The MsOH (316 mL) was added in 3 L of EtOAc at 45~C. To the deep
red homogeneous solution was added 496 g of the title compound Form
I seed (10% seed based on the weight of the free amine was employed).
The temperature rose to ca. 48~C and the reaction was aged at 52~C for
1.5 hours. Analysis indicated complete conversion to the title compound
(Form I). (At less than 10% seed longer age (> 3 hours) was required).
The slurry was allowed to cool to 20~C overnight and was filtered in a
centrifuge under N2. The cake was washed with 20 L of 8% EtOH in
EtOAc. N2 is essential during filtration because the wet crystals are
very hygroscopic. The batch was dried at 35~C under vacuum to afford
2.7Kg (56% overall yield) of the title compound (Form I) (99.9 A%
purity; < 0.1% enantiomer).
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The conversion of Porm II to Form I is also accomplished
where the salt is formed in EtOAc-EtOH by addition of MsOH as above
and the initial solution of the salt (at 55~C) is cooled to 45~C. Crystals
start appearing at that temperature and the slurry becomes thicker with
S time. The tempeLalure is then raised to 51~C and the slurry is aged
overnight. Complete conversion to Form I of 16 should be expected.
This procedure may also be employed to ~ 3ale seed crystals of Form
I of 16.
FXAMPLE 18B
H H CH3~cH3
3--~ C-- ~ NH2~ CH3SO3H
SO2Me
Spiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl~-2-(phenylmethyl-
oxy~ethyl~-2-amino-2-methylpropanamide Methanesulfonate (16)
Materials:
Amine (15) 3.1 kg (5.86 mol)
Meth~nesulfonic acid 620 g (6.45 mol)
Fthyl acetate 37 L
Absolute etharlol 8.7 L
20 Spiro~3H-indole-3,4'-piperdin]- 1 '-yl)-
carbonyl] -2-(phenylmethyl-o~y)ethyl] -
2-amino-2-methylprop~n~mide
methanesulfonate (Form I) 70 g (0.11 mol)
Absolute ethanol (6.4 L) was added to the solution of the
amine (1~) (3.1 kg) in ethyl acetate (total volume ~62 L) in a reacttion
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vessel. The batch was warmed to 50~C and a solution of
~ methanesulfonic acid (620 g, 412 ml, 1.1 equivs.) in ethyl acetate (11
L) was added over ~5 minlltes at 50-54~C. The batch was seeded with
spiro[3H-indole-3,4'-piperdin]- 1 '-yl)-carbonyl]-2-(phenylmethyl-
S oxy)ethyl]-2-alnino-2-methylprop~n~mide methanesulfonate (Form I)
(70 g) and the resulting slurry was stirred and h~.~te-l at 55~C under
nitrogen atmosphere overnight.
The next day, the slurry was cooled to 15-20~C, held for 2
hours and then dropped to the 50 cm polypropylene filter under
10 nitrogen atmosphere. The solid product was washed with a mixture of
absolute ethanol (2.3 L) in ethyl acetate (26 L). The white, solid
product was dug off and dried in an Apex oven in vacuo at 35~C for an
~p~ iate time (approx. two days). The dried spiro[3H-indole-3,4'-
piperdin]- 1 '-yl)-carbonyl]-2-(phenylmethyl-oxy)ethyl]-2-amino-2-
15 methylpropanamide methanesulfonate (3.352 kg) was sieved using aJackson-Crockatt sieve to give 3.347 kg (including seed, 70 g) } yield =
3.277 kg.
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HPLC Conditions:
LC Retention times on Zorbax RX-C8 (4.6 mm x 25 cm),
~ = 210 nm, flow rate = 1.5 ml/min.
S Compound 1: 60:40 CH3CN-H20 (1 % H3PO4) RT = 5.0 rnin
Compound lb: 35:65 CH3cN-H2o (0.1 w % NH40Ac) RT - 6.2 min.
Compound 10: 60:40 CH3CN-H2O (0.1 H3PO4) RT = 2.9 min.
Compound 11: 60:40 CH3CN-H20 (0.1% H3PO4) RT = 5.4 rnin.
Compound 12: 40:60 CH3CN-H20 [pH 5.25 NaH2P04 (6.9 glL of H20)
(adjust pH with NaOH)] RT = 5.6 min
Compound 14: 60:40% CH3CN-H20 (0.1% H3PO4) RT = 4.65 min
Compound 15: 40:60% CH3CN-H20 [pH = 5.25 NaH2P04 (6.9 g/L
of H2O)] adjust pH with NaOH)RT = 4.9 min
LC Retention times on Zorbax RX-C8 (4.6 mm x 25 cm),
~ = 210 nm, flow rate = 1.2 ml/min, column temperature = 48~C
Solvent A = 0.05% Phosphoric acid ~ 0.01% Triethyl~mine in water
Solvent B = Acetonitrile
Gradient system:
Time % A % B
0 min 95 5
35 min 10 90
38min 95 5
40 rnin 95 5
Retention time (mins)
Compound 1 25.2
Compound lb 8.5
Compound 10 20.5
Compound 11 26.3
Compound 12 14.8
Compound 14 25.6
Compound lS 15.7
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~XAMPLE 19
Procedure for Manufacturing 1.0 mg Potency Tablets of N-[l(R)-
[(1,2-dihydro- 1 -methanesulfonylspiro[3H-indole-3,4'-piperdin]- 1 '-
5 yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-methyl-
prop~n~ide methanesulfonate
In redient Per Tablet Per 2600 Tablets
Active Ingredient 1.18 mg 3.068 g
(N-[ l (R)-[(1,2-dihydro- 1 -methane-
sulfonylspiro[3H-indole-3,4'-piperdin]-
l'-yl)carbonyl]-2-(phenylmethyloxy)-
ethyl]-2-amino-2-methylprop~n~rnide
meth~n~sulfonate)
Calcium Phosphate Dibasic47.32 mg 123.03 g
Starch Pregel~tini7e~1 30.00 mg 78.0 g
.NF 1500
Microcrystalline Cellulose NF15.00 mg 39.0 g
Avicel PH 101
Magnesium Stearate Impalpable0.50 mg 1.3 g
Powder NF
Croscarmellose Sodium NF12.75 mg 33.15 g
Ethanol 95% 7.5 ,ul 19.5 ml
Water pllrifie-1 22.5 ~1 58.5 ml
(Tablet Weight = 100 g)
The active ingredient (equivalent to 1.0 mg anhydrous free
base per tablet) was mixed with the calcium phosphate dibasic, the starch
pregel~tini~e.l NF 1000, the microcrystalline cellulose NF, and half of
the croscarmellose sodium NF in a high shear granulator for 5 minutes.
The 25% ethanol/water gran~ ting solution was slowly added to the
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powder mixture with the mixer rllnnin~ over a period of about 1.5
minlltes then granulated for about 7 minutes to form granules. The wet
granules were dried at about 47~C (range 46 to 48~C) in a tray dryer or
a f~uid bed dryer for approxim~t~ly 3.0 hours. The dried granules were
S then milled using a Quadro Comill to achieve fime granules. After
milling, the rem~in(1er of the croscarmellose sodium NFS was added to
the fine granules and mixed in a V blender for about 10 mimltes.
Magnesium stearate impalpable powder N~ was added to this blend
through a 60 mesh stainless steel screen and blended in ~e V blender
10 for about 1 minute. The lubricated mixture was compressed to provide
tablets of 1.0 mg active ingredient (free base equivalent).
E~AMPLE 20
15 Procedure for Manufacturing 1.0 mg Potency Coated Tablets of
N-[ 1 (R)-[( 1 ,2-dihydro- 1 -methanesulfonylspiror3H-indole-3,4'-piperdin]-
1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl~ -2-amino-2-
methylpropanamide methanesulfonate
In~redient Per Tablet Per 2600 Tablets
Hydroxypropyl Me~ylcellulose0 80 mg 2 08 g
USP (HPMC)
Hydroxypropyl Cellulose NF0.80 mg 2.08 g
with < 0.3% Silica (HPC)
Titanium Dioxide USP 0.32 mg 0.83 g
Talc USP Purified 0.08 mg 0.21 g
Water Purified To 20 ,ul To 52 ml
(Film Coated Tablet Weight = 102 g)
The titanium dioxide and talc, USP were mixed and passed
35 through a 60 mesh stainless steel screen. This mixture was mixed with
HPMC and HPC to form a dry blend. The dry blend was added to water
(20 ml) which was previously heated to 90~C with mild agitation to
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ensure that the blend is wetted to form a slurry. The rem~in~ler of the
water (up to 32 ml) was added to the slurry at ambient temperature with
gentle agitation to form a suspension. The suspension was then applied
to the tablets from the previous Example using the following guidelines
5 to provide the coated tablets.
Pan: suitable size
Pan Speed: 20 RPM
Nozzles: 2850 liquid/120 air
Inlet Tempel~Lure: 85~C
Bed Temperature: 47~C
Spray Rate: ca. 2.0 g/minute/kg Tablets
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EXAMPLE 21
Procedure for Manufacturing 5.0 mg Potency Tablets of N-[l(R)-
[( 1 ,2-dihydro- 1 -methanesulfonylspiro[3H-indole-3,4'-piperdin]- 1'-
5 yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-methyl-
propanamide methanesulfonate
In~redient Per Tablet Per 25.000Tablets
Active Ingredient 5.91 mg 147.8 g
(N- [ 1 (R)- [( 1 ,2-dihydro- 1 -methane-
sulfonylspiro[3H-indole-3 ,4'-piperdin]-
1 '-yl)carbonyl]-2-(phenylmethyloxy)-
ethyl]-2-amino-2-methylpropanamide
methanesulfonate)
Calcium Phosphate Dibasic188.10 mg 4.70 kg
Starch Pregel~tini7ed 120.00 mg 3.00 kg
NF 1500
Microcryst~lline Cellulose NF60.00 mg 1.50 kg
Avicel PH 101
Magnesium Stearate Impalpable2.00 mg 50.0 g
Powder NF
Croscarmellose Sodium NF24.00 mg 600 g
Ethanol 95% 30 ~1 750 ml
Water purified 90 ,ul 2.251
(Tablet Weight = 400 g)
The active ingredient (equivalent to 5.0 mg anhydrous free
base per tablet) was mixed with the calcium phosphate dibasic, the starch
pregel~tini7ed NF 1000, the microcrystalline cellulose NF, and half of
the croscarmellose sodium NF in a high Fielder 10/25 mixer for about 6
minutes. The 25% ethanol/water grannl~ting solution was slowly added
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to the powder mixture with the mixer r~-nnin~ over a period of about
1.5 minlltes then gr~n~ ted for about 8 minutes to form granules. The
wet granules were dried at about 47~C (range 46 to 48~C) in a tray
dryer or a fluid bed dryer for approximately 3.0 hours. The dried
5 granules were then milled using a Quadro Comill to achieve fine
granules. After milling, the remainder of the croscarmellose sodium
NFS was added to the fine granules and mixed in a V blender for about
10 minlltto,s. M[agnesium stearate impalpable powder NF was added to
this blend through a 60 mesh stainless steel screen and blended in the V
10 blender for about 1 minute. The lubricated mixture was compressed to
provide tablets of 5.0 mg active ingredient (free base equivalent).
EXAMPLE 22
15 Procedure for Manufacturing 5.0 mg Potency Coated Tablets of
N-[l(R)-[(1,2-dihydro-1-methanesulfonylspiro[3H-indole-3,4'-piperdin]-
1 '-yl)carbonyl] -2-(phenylmethyloxy)ethyl] -2-amino-2-
methylpropanamide methanesulfonate
In~redient Per Tablet Per 25.000 Tablets
Hydroxypropyl Methylcellulose3.2 mg 80 g
USP (HPMC)
Hydroxypropyl Cellulose NF3.2 mg 80.0 g
with < 0.3% Silica (HPC)
Titanium Dioxide USP 1.28 mg 32.0 g
Talc USP Purified 0.32 mg 8.0 g
Water Purified To 80 ~l To 200 ml
(Film Coated Tablet Weight = 408 g)
-
Using essentially the procedure of the prior Example 20
35 and applying the suspension to the tablets from the previous Example,
5.0 mg potency coated tablets were formed.
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F.XAMPLE 23
Procedure for Manufacturing 25 mg Potency Tablets of N-[l(R)-
[(1,2-dihydro- 1 -methanesulfonylspiro[3H-indole-3,4'-piperdin]- 1 '-
S yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-methyl-
prop~n~rnide methanesulfonate
In~redient Per Tablet Per 25.000Tablets
Active Ingredient 29.55 mg 738.75 g
(N-[l(R)-[(1,2-dihydro-1-methane-
sulfonylspiro[3H-indole-3,4'-piperdin]-
1 '-yl)carbonyl] -2-(phenylmethyloxy)-
ethyl] -2-amino-2-methylpropanamide
methanesulfonate)
Calcium Phosphate Dibasic174.46 mg 4.361 kg
Starch Pregel~tini7ed 113.00 mg 2.825 kg
NF 1500
Microcrystalline Cellulose NF57.00 mg 1.425 kg
Avicel PH 101
Magnesium StearateImpalpable2.00 mg 50.0 g
Powder NF
Croscarmellose Sodium NF24.00 mg 600 g
Ethanol 95% 30 ~11 750 rnl
Water purified 90 ,ul 2.25 1
(Tablet Weight = 400 g)
The active ingredient (equivalent to 25 mg anhydrous free
base per tablet) was mixed with the calcium phosphate dibasic, the starch
pregel~tini7ed NF 1000, the rnicrocrystalline cellulose NF, and half of
the croscarmellose sodium NF in a high shear gr~n~ tor Fielder 10/25
mixer for about 6 minutes. The 25% ethanol/water granlll~tin~ solution
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was slowly added to the powder mixture with the mixer rllnnin~ over a
period of about 1.5 minutes then gr~n~ ted for about 8 minutes to form
granules. The wet granules were dried at about 47~C (range 46 to
48~C) in a tray dryer or a fluid bed dryer for approximately 3.0 hours.
S The dried granules were then milled using a Quadro Comill to achieve
fine granules. After millin~, the rem~incler of the croscarmellose
sodium NFS was added to the fine granules and mixed in a V blender
for about 10 minllte.s. Magnesium stearate imp~lriqkle powder NF was
added to this blend through a 60 mesh stainless steel screen and blended
10 in the V blender for about 1 minute. The lubricated mixture was
compressed to provide tablets of 25 mg active ingredient (free base
equivalent).
E;XAMPLE 24
Procedure for Manufacturing 25 mg Potency Coated Tablets of
N- [ 1 (R)-[( 1 ,2-dihydro- 1 -methanesulfonylspiro[3H-indole-3 ,4'-piperdin]-
1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-
methylprop~n~mide methanesulfonate
In~redient Per Tablet Per 25.000 Tablets
Hydroxypropyl Methylcellulose3.2 mg 80 g
USP (HPMC)
Hydroxypropyl Cellulose NF3.2 mg 80.0 g
with < 0.3% Silica (HPC)
Titanium Dioxide USP 1.28 mg 32.0 g
Talc USP Purified 0.32 mg 8.0 g
Water Purified To 80 ~1 To 200 ml
(Film Coated Tablet Weight = 408 g)
Using essentially the procedure of Example 20 and applying
the suspension to the tablets from the previous Example,
25 mg potency coated tablets were formed.
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EXAMPLE 25
Procedure for Manufacturing 100 mg Potency Tablets of N-t1(R)-
[(1,2-dihydro- 1 -methanesulfonylspiro[3H-indole-3,4'-piperdin]- 1 '-
yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-methyl-
prop~n~mide ~ethanesulfonate
Ingredient Per Tablet Per 2600 Tablets
Active Ingredient 118.20 mg 307.3 g
(N-[ l (R)-[(1,2-dihydro- 1 -methane-
sulfonylspiro[3H-indole-3,4'-piperdin]-
1 '-yl)carbonyl]-2-(phenylmethyloxy)-
15 ethyl]-2-amino-2-methylprop~n~Tnide
methanesulfonate)
Calcium Phosphate Dibasic 81.80 mg 212.7 g
Starch Pregel~tini7e~1 78.00 mg 202.8 g
NF 1500
Microcrystalline Cellulose NF 60.00 mg 156.0 g
Avicel PH 101
Magnesium StearateImpalpable 2.00 mg 5.20 g
Powder NF
Croscarmellose Sodium NF 60.00 mg 156.0 g
Ethanol 95% 30.0 ,ul 78.0 ml
Water puri~led 90.0 ,ul 234.0 ml
(Tablet Weight = 400 g)
The active ingredient (equivalent to 100 mg anhydrous free
base per tablet) was mixed with the calcium phosphate dibasic, the starch
pregel~tini7ed NF 1000, the microcrystalline cellulose NF, and half of
the croscarmellose sodium NF in a high shear granulator for S minutes.
- =
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The 25% ethanol/water granulating solution was slowly added to the
powder mixture with the mixer r~lnnin.~ over a period of about 1.5
minutes then granulated for about 7 minutes to form granules. The wet
granules were dried at about 47~C (range 46 to 48~C) in a tray dryer or
5 a fluid bed dryer for approximately 3.0 hours. The dried granules were
then milled using a Quadro Comill to achieve ~me granules. ~fter
milling, the rem~incl~r of the croscarmellose sodium NFS was added to
the fine granules and mixed in a V blender for about 10 minutes.
Magnesium stearate impalpable powder NF was added to this blend
10 through a 60 mesh st~inles.s steel screen and blended in the V blender
for about 1 minute. The lubricated mixture was compressed to provide
tablets of lO0 mg active ingredient (free base equivalent).
FXAMPLE 26
Procedure for ~nllf~cturing 100 mg Potency Coated Tablets of
N-[l(R)-[(1,2-dihydro-l-methanesulfonylspiro[3H-indole-3,4'-piperdin]-
l '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-
methylprop~n~mi~1e methanesulfonate
In~redient Per Tablet Per 2600 Tablets
Hydro~yl,r~yl Methylcellulose3.2 mg 8.32 g
USP (HPMC)
Hydroxypropyl Cellulose NF 3.2 mg 8.32 g
with < 0.3% Silica (HPC)
Titanium Dioxide USP 1.28 mg 3.33 g
Talc USP Purified 0.32 mg 0.83 g
Water Purified To 80.0 ~1 To 208 ml
(Film Coated Tablet Weight = 408 g)
Using essentially the procedure of Example 20 and applying
the suspension to the tablets from the previous Example,
100 mg potency coated tablets were formed.
_ _ _
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E~AMPLE 27
Preparation of amorphous form of N-[l(R)-[(1,2-dihydro-1-methane-
sulfonylspiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyl-
~5 oxy)ethyll-2-amino-2-methylpropanamide methanesulfonate
As a mimic of the tablet formulation process, a
concentrated solution of N-[l(R)-[(1,2-dihydro-1-methanesulfonyl-
spiro[3H-indole-3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)-
ethyl]-2-amino-2-methylprop~n~tnide methanesulfonate (118 mg) in 120
,uL 25% aqueous ethanol (980 mg/ml) was evaporated at 40~C to give a
solid. The lack of crystallinity was con~lrmed by X-ray analysis. The
X-ray diffraction pattern showed an amorphous halo. Ex~min~tion of
the solid under microscopy showed no bioler~ gence.
The solid state chemical stability of the amorphous form
was studied after 12 weeks at 40~C, 60~C and 80~C, and it was found to
exhibit excellent stability. After 12 weeks at 40~C, 100% of the initial
compound was present; after 12 weeks at 60~C, 99.7% of the initial
compound was present; after 12 weeks at 80~C, 97.8% of the initial
20 compound was present.
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E,XAMPLE 28
Chemical Stability of Film Coated Tablets of N-[l(R)-[(1,2-dihydro-1-
methanesulfonylspiro[3H-indole-3,4'-piperdin]-1'-yl)carbonyl]-2-
5 (phenylmethyloxy)ethyl]-2-amino-2-methylpropanamide
m~thanesulfonate Follow;n~ 6.0 Months Stability Study
The results of a 6.0 month stability study of film coated
tablets of N- [ 1 (R)- [( 1 ,2-dihydro- 1 -methanesulfonylspiro [3H-indole-3 ,4'-
piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-
10 methylprop~n~mide methanesulfonate are presented below.
% of initial % of initial
Stability Chamber 1.0 mg dose 100 mg dose
30~C/Amient 99 99
~llmi~lity
30~C/75% Relative 99 99
E~llmiflity
40~C/75% Relative 99 99
Humidity
In particular, no degradates were observed for the 100 mgdose. Degradates were observed for the 1.0 mg dose varying from 0.1
to 0.7 area % relative to active only at 40~C/75% relative humidity.
25 Furthermore, the tablet dissolution, ~li.cint~gration, and hardness for
both the 1.0 mg dose and the 100 mg dose were s~ti~f~ctory following
6.0 months storage under the above conditions.
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EXAMPLE 29
Tablet Crll~hing Strength and Disintegration Times of 100 mg Potency
Tablets of N-[l(R)-[(1,2-dihydro-1-methanesulfonylspiro[3H-indole-
5 3,4'-piperdin]- 1 '-yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-
methylprop~n~mic1e methanesulfonate
Tablets of 400 mg co~ ,ession weight were prepared by
the procedures of the above examples using 118.2 mg of N-[l(R)-[(1,2-
dihydro- 1 -methanesulfonylspiro[3H-indole-3,4'-piperdin]- 1'-
10 yl)carbonyl]-2-(phenylmethyloxy)ethyl]-2-amino-2-methylprop~n~mi~le
methanesulfonate per tablet (29.6% tablet weight). All gr~n~l]~tions
were prepared on a several gram scale by pestle and morter using 25%
ethanol/75% water as a gran ll~tin~ solution. Tablets were compressed
on a Carver Press under 1000 lb force (uncoated tablet cores), unless
15 other~,vise noted.
Formulation A: wet gr~n~ tion - Starch Pregel~tini7~-1 NF
1500 (34.8%); Microcrystalline Cellulose NF Avicel PH 101 (34.8%);
Talc USP (0.6%); Magnesium Stearate (0.3%).
Formulation B: direct compression - Calcium Phosphate
20 Dibasic (26.3%); Microcrystalline Cellulose NF Avicel PH 102 (39%);
Talc USP (1.2%); Magnesium Stearate (0.6%); Croscalmellose Sodium
NF (3%)-
Formulation C: wet granulation, compressed under 500 lbforce - Starch Pregel~tini7ed NF 1500 (33.3%); Microcrystalline
25 Cellulose NF Avicel PH 101 (33.3%); Talc USP (0.6%); Magnesium
Stearate (0.3%); Croscarmellose Sodium NF (3% = 1.5% intragranular
+ 1.5% extragranular).
Formulation D: wet granulation - Starch Pregel~tini7ed NF
1500 (33.3%); Microcrystalline Cellulose NF Avicel PH 101 (33.3%);
30 Talc USP (0.6%); Magnesium Stearate (0.3%); Croscarmellose Sodium
NF (3% extragranular).
Formulation E: wet gr~n~ tion - Microcrystalline
Cellulose NF Avicel PH 101 (36.4%); Calcium Phosphate (26.3%); Talc
-
CA 022348l7 l998-04-08
W ~ 97~15191 PC~nUS96~196
- 79 -
USP (1.2%); Magnesium Stearate (0.6%); Croscarmellose Sodium NF
(6% = 3% intragranular + 3% extragranular).
Formulation F: wet granulation - Microcrystalline
Cellulose NF Avicel PH 101 (l5.0~o); Calcium Phosphate (29.5%);
S Starch Pregel~tini7ed NF 1500 (19.5%); Magnesium Stearate (0.5%);
Croscarmellose Sodium NF (6% = 3% intragranular + 3%
extragranular) .
Formulation G: wet granulation - ~icrocrystalline
Cellulose NF Avicel PH 101 (15.0%); Calcium Phosphate (26.5~o);
10 Starch Pregel~tini7erl NF 1500 (19.5%); Magnesium Stearate (O.5~o);
Croscarmellose Sodium NF (9% = 3% intragranular + 6%
extragranular) .
Formulation H: wet granulation - Microcrystalline
Cellulose NF Avicel PH 101 (15.0%); Calcium Phosphate (26.5%);
Starch Pregel~tini7~1 NF 1500 (19.5%); Magnesium Stearate (0.5%);
Croscarmellose Sodium NF (9~o = 3% intragranular + 6%
extragranular); Super Disintegrant (12% extragranular).
Tablet Disintegration
Form~ tion Hardness (kP) Time (min)
A Does not break 13.0
B 23.3 (one tablet) 7.0
C 23.7 (one tablet) 14.0
D 29.2 i 1.2 (three tablets) 13.5 + 0.25
E 14.1 + 0.7 (three tablets) 12.5 + 1.6
F 13.5 i 1.1 (three tablets) 20.5 i 0-0
G 18.5 (two tablets) 14.0
H 14.9 (two tablets) 7.5
As demonstrated above, the present formulations have
superior properties regarding strength and stability. The tablet hardness
is suitable for film coating and disintegration time is not too long.
CA 02234817 1998-04-08
W O 97/1~191 PCTrUS96/17196
- 80 -
While the invention has been described and illustrated with
reference to certain particular embodiments thereof, those skilled in the
art will appreciate that various adaptations, changes, modifications,
substitutions, deletions, or additions of procedures and protocols may be
5 made without departing from the spirit and scope of the invention. For
exarnple, effective dosages other than the particular dosages as set forth
herein above may be applicable as a consequence of variations in ~e
responsiveness of the m~mm~l being treated for any of the in~ tions
with the compound of the invention indicated above. Likewise, the
10 specific ph~ cological responses observed may vary according to and
depending upon the particular active compound selected or whether
there are present pharmaceutical carriers, as well as the type of
formulation and mode of ~-1mini.~tration employed, and such expected
variations or differences in the results are contemplated in accordance
15 with the objects and practices of the present invention. It is intended,
therefore, that the invention be defined by the scope of the claims which
follow and that such claims be interpreted as broadly as is reasonable.
