Note: Descriptions are shown in the official language in which they were submitted.
~ WO94/0~496 214311 7 PCI/US93/09152
TITLE OF THE rNVENTION
TOCOLYTIC OXYTOCIN RECEPTOR ANTAGONISTS
FIELD OF THE ~VENTION
The present invention provides novel compounds, novel
compositions, methods of their use and methods of their m~mlf~cture, such
compounds generally pharmacologically useful as agents in obstetric and
gynecologic therapy. The aforementioned pharmacologic activities are
useful in the treatment of m~mm~l~. More specifically, ~e compounds of
the present invention can be used in the treatment of preterm labor,
stopping labor preparatory to Caesarean delivery, and in the treatment of
dysmenorrhea. At the present time, there is a need in the area of obstetric
and gynecologic therapy for such agents.
BACKGROUND OF THE INVENTION
In the field of obstetrics, one of the most i~nportant problems
is the m~n~gement of preterm labor. A significant number of the
pregnancies progressing past 20 weeks of gestation experience premature
labor and delivery, which is a le~clin~ cause of neonatal morbidity and
mortality. Despite major advances in neonatal care, retention of the fetus
in utero is preferred in most instances.
Tocolytic (uterine-relaxing) agents ~at are cullelllly in use
include ~2-adrenergic agonists, m~gnesium sulfate and ethanol. Ritodrine,
the le~(lin~ ~2-adrenergic agonist, causes a number of cardiovascular and
metabolic side effects in the mother, including tachycardia, increased renin
secretion, hyperglycemia (and reactive hypoglycemia in the infant). Other
~2-adrenergic agonists, including ~ b~ line and albuterol have side effects
simil~r to those of ritodrine. Magnesium sulfate at plasma concentrations
above the therapeutic range of 4 to 8 mg/dL can cause inhibition of cardiac
conduction and neuromuscular tr~n~mi~sion, respiratory depression and
cardiac arrest, thus m~kinp this agent unsuitable when renal function is
impaired. Ethanol is as effective as ritodrine in preventing premature
WO 94/07496 PCI /US93/09152
21~3117
labor, but it does not produce a corresponding reduction in the inciden ce of
fetal respiratory distress that ~tlmini~tration of ritodrine does.
It has been proposed that a selective oxytocin antagonist would
be the ideal tocolytic agent. In the last few years, evidence has acc1lm~ ted
5 to strongly suggest that the hormone oxytocln may be a physiological
initi~tor of labor in several m~mm~ n species including hllm~n~.
Oxytocin is believed to exert this effect in part by directly contracting the
uterine myometrium and in part by enhancing the synthesis and release of
contractile prost~ n-lin~ from the uterine endometriumldecidua. These
prost~gl~n~lin~ may, in addition, be important in the cervical ripening
process. By these mech~ni~m~, the process of labor (term and preterm) is
initi~ted by a heightened sensitivity of the uterus to oxytocin, resulting in
part as a result of a well-documented increase in the number of oxytocin
receptors in this tissue. This "up-regulation" of oxytocin receptors and
5 enhanced uterine sensitivity appears to be due to trophic effects of rising
plasma levels of estrogen towards term. By blocking oxytocin, one would
block both the direct (contractile) and indirect (enhanced prost~gl~nclin
synthesis) effects of oxytocin on the uterus. A selective oxytocin blocker,
or antagonist, would likely be more efficacious for treating preterm labor
20 than current regimens. In addition, since oxytocin at term has major
effects only on the uterus, such an oxytocin antagonizing compound would
be expected to have few, if any, side effects.
The compounds of the present invention can also be useful in
the treatment of dysmenorrhea. This condition is characterized by cyclic
25 pain associated with menses during ovulatory cycles. The pain is thought
to result from uterine contractions and ischemia, probably mediated by the
effect of prostaglandins produced in the secretory endometriumL. By
blocking both the direct and indirect effects of oxytocin on the uterus, a
selective oxytocin antagonist can be more efficacious for treating
3 dysmenorrhea then current regimens. An additional use for the present
invention is for the stoppage of labor preparatory to Caesarean delivery.
~ W094/07496 21~117 PCr/US93/09152
It is, therefore, a purpose of this invention to provide
substances which more effectively antagonize the function of oxytocin in
disease states in ~nim~l.c, ~lefe~dbly m~mm~l~, especially in hllm~n~. It is
ano~er purpose of this invention to prepare novel compounds which more
5 selectively inhibit oxytocin. It is still another purpose of this invention toprovide a method of antagonizing the functions of oxytocin in disease states
in m~mm~l~. It is also a purpose of this invention to develop a method of
preventing or treating oxytocin-related disorders of preterm labor and
dys~nenorrhea by antagonizing oxytocin.
It has now been found that compounds of the present invention
are antagonists of oxytocin and bind to the oxytocin receptor. VVhen the
oxytocin receptor is bound by the compounds of the present invention,
oxytocin is antagonized by being blocked from its receptor and thus being
unable to exert its biologic or ph~rm~cologic effects. These compounds
15 are useful in the treatment and prevention of oxytocin-related disorders of
~nim~ , preferably m~mm~l~ and especially hllm~n~. These disorders are
primarily preterm labor and dysmenorrhea. The compounds would also
find usefulness for stoppage of labor preparatory to Caesarean delivery.
20 SUMMARY OF THE ~VENTION
The compounds and their pharmaceutically acceptable salts and
esters of the present invention are those of the general structural formlll7~:
X-Y-Z-Rl, wherein
xis
WO 94/07496 PCI /US93/09152 ~
21~3117
- 4 -
¢~W F~
I
W is hydrogen or acetate;
Y is -CO-, -SO2- -CO(CH2)m- or-(CH2)m-;
Z is an optionar substituent that, when present, is one or more of N, O, S,
-CHR-, -CR=CH-, -CH=, -(CH2)m- or-CHCHOH-;
R is hydrogen, Cl 5 aLkyl or Cl 5 alkoxycarbonylamino,
quinuclidinylaminocarbonylamino;
R1 is -CH3, -CH(CH3)2, C1 5 alkoxycarbonyl,
~ W094/07496 21~3117 PCI~/US93/09152
R3 ' ~ , ~0 .
~\~3, ~ --~ ~ R3 ~ ~ R2
~;3 , ~, ~13 ~,~ R2
_~ ~ 2 ~ ~(CH2)m~C 3 <r/~H3
20 -NR4R5 or-NCOR6;
R2 is hydrogen, hydroxy, carboxyl, acetyl, nitro, cis or trand oximino,
halogen,
mono-, di- or tri-Cl 3 aL~cyl, spirocyclic indenyl,
25 N-~iruilldanepiperidinyl, O-R where R is as defined above, O-Het where
Het is imidazole or benzimidazole or azimidobenzene, or where R2 is
fur~er defined as -COR6, -(CH2)m-NHCOR7, -(CH2)mNHCOOR7~
-(CH2)m-NR8R9, -(CH2)m-NHCo-(cH2)mR7, -(CH2)~,-NHCo-cHR7R7,
-(CH2)m-NHCo-CH=CHR7, -(CH2)m-CO-O -R7, -(CH2)m-CO-O-
3 (CH2)mR7, -(CH2)~m-C0-0-CHR7R7, -(CH)m-Co-o-cH=cHR7~
-NHS02R-where R is as de~ned above, NHSo2R7, -(CH2)m-O-R10,
-S02R10, CoRl 1
WO 94/07496 PCI`/US93/09152~
2143117
-CONH(CH2)m~ -(CH2)m~ HO~)
CH3
~SO2~ NH2
CH3 . or (CH2)m 2~cH
o or one to two substituents selected from the group consisting of
/~¢~ or
15 R3 is one or two of hydrogen, hydroxyl or Cl S aLkyl;
with the proviso that when R1 is cyclohexyl, then R2 and R3 are limited to
being hydroxyl or Cl 5 aLkyl;
20 R4 is hydrogen, C1 5 aLkyl, or C6 10 cycloalkyl;
R5 is hydrogen or acetyl;
O CH3
R6 is SO2~ 3 or ~ SOz~ 3;
R7is
~ w094/07496 21~311 7 PCr/US93/09152
~,~Rl2 ~NH ~(CH2)m ~C3
-(CH2)m \=~ Rl2 ~ ~(CH2)m N , NH ~
~(CH2)m--~ \> -(CH2)m--~H . -(cH2)m~1,
-(C~ ~ -(CH2)m~
CH CH~,~ -(CH2)m ~0--1<CH
O NH2
-(CH2) ~ <C ~(CH2)m ~ ~~CH2)m ~cH2
hydrogen, Cl 4 aLkyl, NS02R1 2 or NHO-C1 -4 alkyl;
R8 is hydrogen or C1 5 alkyl;
R9 is hydrogen or Cl 5 aLkyl;
3 0 R l is -CH3,
W O 94/07496 PC~r/US93/09lS2 ~
2~3~l~
.
~CH3 CH2~cH3
CH=CH~CH3 ~ ~ CH2C
N~
Rl 1 is -CH3, H
--CH=CH~ (CH2)m~, ~(CH2)m N
N--N N=\
~(CH ) ~N~> or ~CH=CH~
R12 is hydrogen, C1 5 alkyl or Cl 5 alkoxy; and
25 m is an integer of from O to 5;
~W
with the proviso ~at when X is N
~ WO 94/07496 2 1 ~ 31 1 7 PCl/US93/0915'
g
and when Rl is disubstituted phenyl when the phenyl substit--ent~ are any
of hydroxyl, carboxyl, nitro, halogen, mono-, di- or tri-Cl 3 alkyl, Cl 5
aL~coxy; or when Rl is pyridyl; or when Rl is -CH3 or -CH(CH3)2; or
when Rl is unsubstituted bicyclo loweralkyl of 9 carbons or unsubstituted
5 or substituted cyclohexyl and the substituent is hydroxyl; then Y is
-(CH2)m- where m has a value of from 1 to 5.
Salts and esters encompassed within the term
"ph~ ceutically acceptable salts and esters" refer to non-toxic salts of the
compounds of this invention which are generally prepared by reacting the
free base with a suitable organic or inorganic acid. Representative salts
include the following salts:
Acetate Lactobionate
Benzenesulfonate Laurate
Benzoate Malate
Bicarbonate Maleate
Bisulfate Mandelate
Bitartrate Mesylate
Borate Methylbromide
2 0 Bromide Methylnitrate
Calcium Edetate Methylsulfate
Camsylate Mucate
Carbonate Napsylate
Chloride Nitrate
2 5 Clav~ n~te N-methylglllc~mine
Citrate ammonium salt
Dihydrochloride Oleate
Edetate Oxalate
- Edisylate Pamoate (Embonate)
3 Estolate P~lmit~te
- Esylate Pantothenate
Fumarate Phosphate/diphosphate
WO 94/07496 PCl /US93/09152 ~
2~4~ Ll~
-
- 10-
Gluceptate Polygalacturonate
Gluconate Salicylate
Glllt~m~te Stearate
Glycollylars~nil~te Sulfate
Hexylresorcinate Subac~tate
Hydrabamine Succinate
Hydrobromide Tannate
Hydrochloride Tartrate
Hydroxynaphthoate Teoclate
Iodide Tosylate
Isothionate Triethiodide
Lactate Valerate
5
The term "pharmacologically effective amount" shall mean
that amount of a drug or pharmaceutical agent that will elicit the biological
or medical response of a tissue, system, ~nim~l or hllm~n that is being
sought by a researcher or clinician.
The term "alkyl" shall mean straight or branched chain
alkanes of one to ten total carbon atoms, or any number within this range.
The term "alkenyl" shall mean straight or branched chain
aLkenes with one or more degrees of lm.~ lration at any position on the
chain, of two to ten total carbon atoms, or any number within this range.
The term "alkynyl" shall mean straight or branched chain
alkynes with one or more degrees of lmc~lllration at any position on ~e
chain, of two to ten total carbon atoms, or any number within ~is range.
The term ""aryl" shall mean phenyl, naphthyl or fluorenyl.
The term "cycloalkyl" shall mean cyclic rings of alkanes of
three to eight total carbon atoms.
Whenever the terrns "alkyl" or "aryl" or either of their prefix
roots appear in a name of a substituent (e.g. aralkoxyaryloxy) they shall be
inte~ ed as including those limitations given above for "alkyl" and
~ WO94/07496 21~3117 PCl/US93/09152
"aryl". Designated numbers of carbon atoms (e.g. C1 1o) shall refer
independently to the number of carbon atoms in an alkyl or cyclic alkyl
moiety or to the alkyl portion of a larger substituent in which alkyl appears
as its prefix root.
The term "oxo" shall refer to the substituent =0.
The term "azimidobenzene" (also known as benzotriazole)
shall refer to the moiety
1 o N~ ~
The term "spirocyclic indenyl" shall refer to the moiety
1~1
The term "halogen" shall include iodine, br~ e, chlorine
and fluorine.
The term "preterm labor" shall mean expulsion from the
uterus of a viable infant before the normal end of gestation, or more
particularly, onset of labor with effacement and dilation of the cervix
before the 37th week of gestation. It may or may not be associated with
vaginal bleeding or ~ e of the membranes.
The term "dysmenorrhea" shall mean painful menstruation.
The term "Caesarean delivery" shall mean incision through the
abdominal and uterine walls for delivery of a fetus.
The term "substituted" shall be deemed to include multiple
degrees of substitution by a named substitutent.
WO94/07496~ 21~311'Z PCl/US93/09152~
- 12 -
Where multiple substituent moieties are disclosed or claimed,
the substituted compound can be independently substituted by one or more
of the disclosed or claimed substituent moieties, singly or plurally.
The ability of the compounds o~ ~ormula I to antagonize
5 oxytocin makes these compounds useful as ph~ cologic agents for
m~mm~l~, especially for hllm~n~, for the treatment and prevention of
disorders wherein oxytocin may be involved. F.x~mples of such disorders
include preterm labor and especially dysmenorrhea. These compounds
may also find usefulness for stoppage of labor preparatory to Cesarean
1 delivery-
Because of the known relationship of vasopressin to oxytocin,the compounds of the present invention are also useful as vasopressin
antagonists. Vasopressin antagonists are useful in the treatment or
prevention of disease states involving vasopressin disorders, including their
15 use as diuretics and their use in congestive heart failure.
The compounds of the present invention can be ~lmini~tered
in such oral dosage forms as tablets, capsules (each including timed release
and sustained release formulations), pills, powders, granules, elixers,
tinctures, suspensions, syrups and em~ ions. Likewise, they may also be
20 ?~flmini~tered in intravenous (both bolus and infusion), intraperitoneal,
subcutaneous or intramuscular form, all using forms well known to those
of ordinary skill in the ph~ ceutical arts. An effect*e but non-toxic
amount of the compound desired can be employed as a tocolytic agent.
The dosage regimen l~tili7in~ the compounds of the present
25 invention is selected in accordance with a variety of factors including type,species, age, weight, sex and medical condition of the patient; the severity
of the condition to be treated; the route of ~(lmini~tration; the renal and
hepatic function of the patient; and the particular compound or salt thereof
employed. An ordinarily skilled physician or veterinarian can readily
30 determine and prescribe the effective amount of the drug required to
prevent, counter or arrest the progress of the condition.
2I~311 7
WO 94/07496 - PCr/US93/09152
Oral dosages of the present invention, when used for the
indicated effects, will range between about 0.3-6.0 gm/day orally.
Intravenously, the most preferred doses will range from 0.1 to about 10
mg/mimlt~ during a constant rate infusion. Advantageously, compounds of
5 the present invention may be ~lministered in a single daily dose, or the
total daily dosage may be ~(lmini~tered in divided doses of two, three or
four times daily. Furthermore, preferred compounds for the present
invention can be ~1mini~tered in intranasal form via topical use of suitable
intranasal vehicles, or via transdermal routes, using those forms of
transdermal skin patches well known to those of ordinary skill in that art.
To be ~lmini~tered in the form of a transdermal delivery system, the
dosage a~mini~tration will, of course, be continuous rather than
intermittant throughout the dosage regimen.
In the methods of the present invention, the compounds herein
15 described in detail can form the active ingredient, and are typically
~lmini~tered in ~ ix~ e with suitable pharmaceutical diluents, excipients
or carriers (collectively referred to herein as "carrier" materials) suitably
selected with respect to the intended form of ~lmini~tration, that is, oral
tablets, capsules, elixirs, syrups and the like, and consistent with
20 conventional ph~ ceutical practices.
For instance, for oral ~lmini~tration in the form of a tablet or
capsule, the active drug component can be combined with an oral, non-
toxic ph~m~ceutically acceptable inert carrier such as ethanol, glycerol,
water and the like. Moreover, when desired or necessary, suitable binders,
25 lubricants, disintegrating agents and coloring agents can also be
incorporated into the mix~lre. Suitable binders include starch, gelatin,
natural sugars such as glucose or beta-lactose, corn sweeteners, natural and
synthetic gums such as acacia, tr~g~r~ntl or sodium alginate,
carboxymethylcellulose, polyethylene glycol, waxes and the like.
3 Lubricants used in these dosage forms include sodium oleate, sodium
stearate, m~gnesium stearate, sodium benzoate, sodium acetate, sodium
wo 94,07496 2 1 ~3 11~ PCI/US93/09152~
- 14 -
chloride and the like. Disintegrators include, without limitation, starch,
methyl cellulose, agar, bentonite, 7~nth~n gum and the like.
The compounds of the present invention can also be
~lmini~tered in the form of liposome delivery systems, such as small
5 lmil~mellar vesicles, large lmil~mellar vesicles and mllltil~mellar vesicles.
Liposomes can be formed from a variety of phospholipids, such as
cholesterol, stearyl~mine or phosphatidylcholines.
Compounds of the present invention may also be delivered by
the use of monoclonal antibodies as individual carriers to which the
o compound molecules are coupled. The compounds of the present invention
may also be coupled with soluble polymers as targetable drug carriers.
Such polymers can include polyvinylpyrrolidone, pyran copolymer,
polyhydroxypropyl- methacrylamide-phenol, polyhydroxyethyl-
aspartamidephenol, or polyethyleneoxidepolylysine substituted with
15 palmitoyl residues. Furthermore, the compounds of the present invention
may be coupled to a class of biodegradable polymers useful in achieving
controlled release of a drug, for example, polylactic acid, polepsilon
caprolactone, polyhydroxy butyric acid, polyorthoesters, polyacetals,
polydihydropyrans, polycyanoacrylates and cross-linked or amphipathic
20 block copolymers of hydrogels.
The compounds of formula I can be prepared readily
according to ~e following reaction Schemes (in which all variables are as
defined before) and Examples or modifications thereof using readily
available starting materials, reagents and conventional synthesis
25 procedures. ~ these reactions, it is also possible to make use of variants
which are themselves known to those of ordinary skill in this art, but are
not mentioned in greater detail.
The most preferred compounds of the invention are any or all
of those specifically set forth in these examples. These compounds are not,
3 however, to be construed as forming the only genus that is considered as
the invention, and any combination of the compounds or their moieties may
itself form a genus. The following examples fur~er illustrate details for
~ WO 94/07496 2 1 ~ 3 1 1 7 PCr/US93/0915'
- 15 -
the preparation of the compounds of the present invention. Those skilled
in the art will readily understand that known variations of the conditions
and processes of the following preparative procedures can be used to
prepare these compounds. All temperatures are degrees Celsius unless
5 noted other~,vise.
WO 94/07496 PCI/US93/09~52 ~
2~3l~l
- 16 -
SCHFME 1 R
Cl-CO CH2~R
N EDC/HBT/DMF or BOP
R~
/ ~+~X-CO-CI ~X
~ R3/`3~ j2N~o-co~
N ~ /~(H)n
N (WhereX=N,OorS)
X = O, N, S, CH2 o~O
n=1,2
NO2
~ wo 94/07496 2 1 ~ 3 1 1 7 PCl /US93/09152
SCHFMF ?
N Hydro9enation N
O~CH2 ~O~cH2
¢~ ~
N2 / ~ NH2
[OH--]/
~ / BOP / \ BOP
20 ~C~ ~ ~
NHCOCH3 / CH2
+ HOOCcH2~ NH2 NHCOR
H
WO 94/07496 PCI`/US93/0915''
2~43~
- 18 -
SCHEM~ 3
CH Where W is a
~ 3 ~W~W~ suitable leaving group
NH2 N and Z is a protecting
z group such as BOC or
CBZ
[~ N~l Deprotection [~CH3
~/ N~z / ~R/ N~H
3// R~
/ Cl-CO~/,~
~CI 13 / R'
R O~ ~N~
~ WO 94/07496 2 1 4 3 1 1 7 PCI~/US93/09152
- 19 -
SCHFME 4 ~=~/R
Cl-CO-CH2~
s ~CH3 or HOOCCH2~R
EDC/HBT/DMF or BOP
~ Cl CO~,~ ¢N~CH3
l 5 I~CH3 X-CO-CI~ o X
~ ~ H
Z H3CJ~ ~/)n
R3 ~ ~ (Where X= N,O or S)
X=O,N,S,CH2
n=1,2 ~
NO2
WO 94/07496 PCl/lJS93/09152~
2~3il~
- 20 -
Abbreviations used in the Examples are as follows:
EDC = l-ethyl-3-(3-dimethylaminQpropyl)carbodiimide
hydrochloride .
BOC = tert-butoxycarbonyl
TEA = triethyl~mine
DIEA = diisopropylethyl~mine
BOP = benzotriazolyloxytris(dimethylamino)
phosphonium hexafluorophosphate
THF = tetrahydrofuran
DMF = dimethylformamide
LAH = lithium all-minllm hydride
TFA = trifluoroacetic acid
HPLC Method A = 15 min. linear gradient
95:5 A:B to 0:100 A:B
A - H20 cont~inin~ 0.1% by vol. TFA
B = CH3CN cont~inin~ 0.1% by vol. TFA
2.0 mL/min flow rate
12 cm Clg reverse phase column
W detection (215 nm)
TLC was performed on 20 cm plates coated with silica gel
(250 microns) from Analtech.
~ wo 94/07496 2 1 ~ 3 1 1 7 PCI /US93/09152
21
EXAMPLE 1
s 0~
0~
70 mg (0.316 mmol) of spiro(lH-indene-1,4'-piperidine HC1)
was dissolved in 3 ml DMF and the solution treated with 52.5 mg (0.347
mmol) of 4-aminophenyl acetic acid followed by 161 mg (0.363 mmol) of
benzotriazol-1-yl-oxy(dimethylamino)phosphonium hexafluorophosphate
(BOP Reagent). The pH of the solution was adjusted to 9.5 with 142 ~11
15 (0 794 mmol) of diisopropylethyl~mine and the mixture stirred at 25 for 1
hour.
DMF was removed in vacuo and the crude residue treated with
water and extracted with ethyl acetate (3x). The organic extracts were
combined, washed with water (lx), brine (lx), dried over Na2S04, filtered
20 and stripped to dryness in vacuo. Flash chromatography of the crude
product on silica gel (1:1 of ethyl acetate: CH2Cl2) gave the title compound
as a white foam (41 mg, 40.6% yield) upon coevaporation with ether (3x)
in vacuo.
M.W.: 318.402.
m.p.: 60-87C (sinter)
HPLC: 96.3%
PMR: Consistent with structure plus ether and water
M.S.: M+H = 319 (FAB).
- TLC: Rf = 0.34, Silica GF (1:1 of EtOAc: CH2Cl2).
CHN: Calc'd as C21H22N2O-0.05 C4H100-0.30 H20
(F.W. = 327.532):
W O 94/07496 43~ PC~r/US93/09152
- 22 -
C, 77.74; H, 7.11; N, 8.55.
Found: C, 77.83; H, 6.80; N, 8.23.
.~ , .
EXAMPLE 2
~ ¢~NH CH3
l.lgm (3.45 mmol) of the product of Fx~mrle 1 was dissolved
in 15 ml of CH2C12 and ~e solution treated with 0.27 ml (3.80 rnrnol) of
acetyl chloride. The pH was adjusted to 9.5 with 0.55 ml (3.82 mmol) of
triethyl~mine and the mixture stirred at 25C for 1 hour.
Flash chromatography of ~e reaction mixture directly on
silica gel (3:1 of ethyl acetate: CH2C12) gave the title compound as a
crystalline solid (992 mg, 79.3% yield) from ethyl acetate.
M.W. 360.438.
m.p.: 217-9C.
HPLC: 99.5%
PMR: Consistent with structure.
M.S.: M+H=361(PAB).
TLC: Rf = 0.31, Silica GF (3:1 of ethyl acetate: CH2C12).
CHN: Calc'd as C23H24N202:
C, 76.64; H, 6.71; N, 7.77.
Found: C, 76.26; H, 6.80; N, 7.76.
2143117
WO 94/07496 - PClr/US93/09152
- 23 -
EXAMPLE 3
0~
NO2
125 mg (0.564 mmol) of spiro(lH-indene-1,4'-piperidine)
hydrochloride was dissolved in 3 ml DMF and the solution treated
with 112 mg (0.620 mmol) of o-nitrophenyl acetic acid followed by
286 mg (0.649 mmol) of benzotriazol-l-yl-oxytris-(dimethylamino)-
phosphonium hexafluorophosphate (BOP reagent). The pH of the solution
was adjusted to 9.5 with 307 ,ul (1.75 mmol) of diisopropyl ethyl~mine and
the mixture stirred at 25C for 1 hour.
DMF was removed in vacuo and the crude residue treated with
H20 and extracted with ethyl acetate (3x). The organic extracts were
20 combined, washed wi~ water (lx), brine (1x), dried over Na2SO4, filtered
and stripped to dIyness in vacuo. Flash chromatography of the crude
product on silica gel (3% Et2O in CH2Cl2) gave 155.7 mg (79.4% yield)
of the title compound as a white foam after evaporation in vacuo.
M.W.: 348.386.
m.p.: 60-75C (sinter).
HPLC: 93.4%.
PMR: Consistent with structure plus water.
M.S.: M+H = 349.2 (FAB).
TLC: Rf = 0.29, Silica GF (4% Et2O in CH2C12).
CHN: Calc'd as C21H20N2O3-0.30 H20 (~.W. = 353.8):
C, 71.28; H, 5.87; N, 7.92.
WO 94/07496 21 ~3 11~ PCI/US93/09152~
- 24 -
Found: C, 71.35; H, 5.64; N, 7.91.
FXAMPLE~
.,
H o
' 0~~
250 mg (1.13 mmol) spiro(lH-indene-1,4'-piperidine) hydro-
chloride was dissolved in 10 ml DMF and the solution treated with
15 350 mg (1.24 mmol) of 2-(t-butyloxyaminomethyl)-phenylacetic acid
followed by 575 mg (1.30 mmol) of benzotriazol-l-yloxytris (dimethyl-
amino)phosphonium hexafluorophosphate (BOP Reagent). The pH of the
solution was adjusted to 9.5 with diisopropylethyl~mine and the mixture
stirred at 25C for 1 hour.
DMF was removed in vacuo and the crude residue treated with
water and extracted with ethyl acetate (3x). The organic extracts were
combined, washed with water (lx), brine (lx), dried over Na2SO4, filtered
and stripped to dryness in vacuo. Flash chromatography of the crude
product on silica gel (13% Et2O in CH2C12) gave the title compound
25 isolated as a crystalline solid (285 mg, 56% yield) from diethyl ether.
M.V.: 448.542.
m.p.: 165-7C.
HPLC: 97.2%.
30 PMR: Consistent wi~ structure plus ether.
M.S.: M+H + Thioglycerol (M.W. = 108) = 5573 (FAB).
TLC: Rf = 0.48, Silica GF (20% ET2O in CH2C12)-
~WO 94/07496 2 1 ~ 3 1 1 7 PCI`/US93/09152
- 25 -
CHN: Calc'd as C27H32N204-0.10 C4HloO (F.W. = 455.954)
C, 72.17; H, 7.30; N, 6.14.
Found: C, 72.12; H,7.33; N, 6.29.
s EXAMPLE S
~ 1
~N
O ~ N O
H
250 mg (1.13 mmol) of spiro(lH-indene-1,4'-piperidene)
hydrochloride was dissolved in 10 ml DMF and the solution treated with
350 mg (1.24 mmol) of 3-(t-butyloxyaminomethyl)-phenyl acetic acid
followed by 575 mg (1.30 mmol) of benzotriazol-1-yloxytris(dimethyl-
amino)phosphonium hexafluorophosphate (BOP Reagent). The pH of the
20 solution was adjusted to 9.5 with 550 ,ul (3.14 mmol) of diisopropyl-
ethyl~mine and the mixture stirred at 25C for 1 hour.
DMF was removed in vacuo and the crude residue treated with
water and extracted with ethyl acetate (3x). The organic extracts were
combined, washedl with water (lx), dried over Na2S04, filtered, and
25 stripped to dryness in vacuo. Flash chromatography of the crude product
on silica gel (20% Et2O in CH2Cl2) gave the title compound as a white
foam (328 mg, 65.0% yield) upon coevaporation with ether (3x) in vacuo.
M.W.: 448.542.
30 m.p.: SS-72C (sinter).
HPLC: 99.6%
PMR: Consistent with structure plus water.
W094/07496 2~43l~rt PCI/US93/09152
- 26 -
M.S.: M+H + Thioglycerol (M.W. - 108) = 557.6 (FAB).
TLC: Rf = 0.52, Silica GF (25% Et2O in Ch2cl2).
CHN: Calc'd as C27H32N2O4-0 15 H20 (F.W. = 451.269):
C, 71.86; H, 7.21; `-N, 6.21.
Found: C, 71.83; H, 7.09; N, 6.45.
EXAMPLE 6
l o ~>~ ,~¢N>
70 mg (0.220 mmol) of the product of Fx~mple 1 was
dissolved in 5 ml DMF and the solution treated with 39.3 mg (0.242 mmol)
of imidazole-4-acetic acid-hydrocloride followed by 32.7 mg (0.242 mmol)
of l-hydroxybenzotriazole hydrate (HBT) and 46.4 mg (0.242 mmol) of 1-
ethyl-3-(3-dimethylaminopropyl)-carbodiimidehydrochloride (EDC). The
pH was adjusted to 9.5 with triethyl~mine and the mixture stirred at 25C
or 4 hours.
DMF was removed in vacuo and the crude residue treated with
water and extracted with e~yl acetate (3x). The organic extracts were
combined, washed with water (1x), brine (1x), dried over Na2SO4, filtered
and stripped to dryness in vacuo. Flash chromatography of the crude
product on silica gel (100:10:1 of CH2C12:MeOH: Conc NH40H) gave the
title compound as a white solid (50.8 mg, 54.2% yield) after evaporation in
vacuo and trituration with ether.
M.W.: 426.50.
m.p.: 104-36C (sinter).
21~3117
WO 94/07496 PCI /US93/09152
- 27 -
HPLC: 99.3%
PMR: Consistent with structure plus ether, CH2C12, and water.
M.S.: M+H + 427.1 (FAB).
TLC: Rf = 0.32, Silica GF (80:10:1 of CH2Cl2: MeOH:
Conc. NH40H).
CHN: Calc'd a C26H26N4)2-0.10 CH2C12-0.10:
C4H1oO-0.50 H20 (F.W. = 451.212):
C, 70.53; H, 6.26; N, 12.42.
Found: C, 70.56; H, 6.04; N, 12.64.
EXAMPLE 7
~ NH2
85 mg (0.267 mmol) of the product of Example 1 was
dissolved in S ml DMF and the solution treated with 72.3 mg (0.294 mmol)
of N-t-butyloxycarbonyl-L-glllt~mine followed by 39.7 mg (0.294 mmol)
of l-hydroxybenzotriazole hydrate (HBT) and 56.3 mg (0.294 mmol of 1-
ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC). The
25 pH was adjusted to 9.5 with 55 ,ul (0.395 mmol) of triethyl~mine (Et3N)
and the ~ Ul~ stirred at 25C for 1.5 hours.
A second portion of 72.3 mg (0.294 mmol) of N-t-butyloxy-
carbonyl-L-~ t~mine, 39.7 mg (0.294 mmol) of H B T, 56.3 mg (0.294
mmol) of EDC and 55 ,ul (0.395 mmol) of Et3N was added and the
30 mixture stirred at 25C for 18 hours.
DMF was removed in vacuo and the residue treated with H20
and extracted with ethyl acetate (3x). The organic extracts were com-
WO 94/07496~ 3~ PCr/US93/0915
- 28 -
bined, washed with water (lx), brine (lx), dried over Na2S04, filtered and
stripped to dryness in vacuo. Flash chromatography of ~e crude product
on silica gel (6% MeOH in CH2C12) ga~ the title compound
as a white solid (57.4 mg, 39.3% yiel~d) after evaporation in vacuo and
5 trituration with ether.
M.W.: 546.646.
m.p.: 104-47C (sinter).
HPLC: 99.1 %.
PMR: Consistent with structure plus water and ether.
M.S.: M+H = 547 (FAB).
TLC: Rf = 0.18, Silica GF (4% MeOH in CH2C12).
CHN: Calc'd as C31H3gN4O5-0.10 C4HloO-0.75 H20
(F.W. = 5657.597):
C, 66.44; H, 7.19; N, 9.87.
Found: C, 66.42; H, 6.98; N, 10.04.
EXAMPLE 8
N ~Br
O~J
75 mg (0.338 mmol) of spiro(lH-indene-1,4'-piperidine)
hydrochloride was dissolved in 3 ml DMF and the solution treated with
80.0 mg (0.372 mmol) of 4-bromophenyl acetic acid followed by 171.9 mg
30 (0.389 mmol) of benzotriazol-l-yloxytris(dimethylamino)-phosphonium
hexafluorophosphate (BOP Reagent). The pH of ~e solution was adjusted
~ WO94/07496 21~3117 PCI/US93/09152
- 29 -
to 9.5 with 125 ml (0.710 mmol) of diisopropylethyl~mine and the mixture
stirred at 25C for 1 hour.
DMF was removed in vacuo and the crude residue treated with
water and extracted with ethyl acetate (3x). The organic extracts were
5 combined, washed with water (lx), brine (lx), dried over Na2SO4, ~1ltered
and stripped to dryness in vacuo. Flash chromatography of the crude
product on silica gel (5% Et20 in CH2Cl2) gave the title compound as a
white foam (30 mg, 23.3% yield) upon coevaporation with ether (3x) in
vacuo.
M.W.: = 382.282.
m.p.: 44-58C (sinter).
HPLC: 99.5%.
PMR: Consistent with structure plus water.
M.S.: M+H = 382.1/384.1 at 1/1 (FAB).
TLC: Rf = 0.44, Silica GF (6% Et2O in ch2c12)-
CHN: Calc'd as C21H20BrNo-o.4o H2O (F.W. = 389.513):
C, 64.75; H, 5.38; N, 3.60.
Found: C, 64.51; H, 5.06; N, 3.60.
FXAMPLE 9
~NH~ >
375 mg (1.18 mmol) of the product of Example 1 was
dissolved in 10 ml DMF and the solution treated with 179 mg (1.30 mmol)
of urocanic acid followed by 175 ml (1.29 mmol of 1-hydroxybenzo-
wo 94/07496 ~ ~3~ ~'1 PCI`/US93/0915
- 30 -
triazole hydrate (HBT) and 248.5 mg (1.30 mmol) of 1-ethyl-3-(3-
dimethyl-aminopropyl) carbodiimide hydrochloride (EDC). The pH of the
solution was adjusted to 9.5 with 414 ,ul (0.298 mmol) of triethyl~mine
(Et3N) and the reaction stirred at 25C for~6~hours.
A second portion consisting of 179 mg (1.30 rnmol) of
urocanic acid, 175 mg (1.29 mmol) of HBT, 249 (1.30 mmol) of EDC and
414 ~11 (0.298 mmol) of Et3N was added and the reaction stirred at 25C
for 18 hours.
DMP was removed in vacuo and the residue treated with water
and extracted with ethyl acetate (3x). The organic extracts were combined,
washed with water (1x), brine (1x), dried over Na2SO4, filtered and
stripped to dryness in vacuo. Flash chromatography of the crude product
on silica gel (100:10:1 of CH2Cl2: MeOH: Conc. NH40H) gave the title
compound as a crystalline solid (106 mg, 20.5% yield) from ethyl acetate.
M.W.: = 438.51.
m.p.: 170-85C (Physical Change); 228-30C (melt).
HPLC: 99.0%.
PMR: Consistent with structure plus water.
2 0 M.S.: M+H = 439 (FAB).
TLC: Rf = 0.36, silica GF (80:10:1 of CH2Cl2:
MeOH: Conc. NH40H).
CHN: Calc'd as C27H26N402-0.95 H20 (F.W.455.468):
C, 71.17; H, 6.17; N, 12.30.
25 Found: C, 71.05; H, 5.78; N, 11.91.
WO 94/07496 PCl /US93/09152
2143117
EXAMPLE 10
.,
. 0~0>
60 mg (0.271 mmol) of spiro(lH-indene-1,4'-piperidine)
hydrochloride was dissolved in 4 ml DMF and the solution treated with
53.7 mg (0.298 mmol) of 3,4-methylenedioxy phenyl acetic acid followed
by 137.8 mg (0.312 mmol) of benzotriazol-1-yloxytris (dimethylamino)
phosphonium hexafluorophosphate (BOP Reagent). The pH of the solution
was adjusted to 9.5 wi~ 100,ul (0.569 mmol) of diisopropylethyl~mine and
the mixture stirred at 25C for 1 hour.
DMF was removed in vacuo and the crude residue treated with
water and extracted with ethyl acetate (3x). The organic extracts were
combined, washed wi~ water (lx), brine (lx), dried over Na2SO4, fîltered
20 and stripped to dryness in vacuo. Flash chromatography of the crude
product on silica gel (6% Et2O in CH2Cl2) gave the title compound as a
c~ystalline solid (32.4 mg, 34.3% yield) from ether.
M.W.: 347.396.
m.p.: 123-5C.
HPLC: 99.4%
PMR: Consistent with structure plus water.
M.S.: M+H = 348.1 (FAB).
TLC: Rf = 0.30, Silica GF (6% Et2O in Ch2cl2)
- CHN: Calc'd as C22H21NO3-0.15 H2O (F.W. = 350.119):
C, 75.47; H, 6.13; N, 4.00.
Found: C, 75.51; H, 5.79; N, 4.01.
WO 94/07496 PCr/US93/0915~
3~
- 32 -
EXAMPLE 11 ~;
N
300 mg (1.35 mmol) of spiro(lH-indene-1,4'-piperidine)
hydrochloride was dissolved in 10 ml DMF and the solution treated with
341 mg (1.49 mmol) of p-bromoethyl phenyl acetic acid followed by 687
15 mg (1.55 mmol) of benzotriazol-l-yloxy(dimethylamino) phosphonium
hexafluorophosphate (BOP Reagent). The pH of the solution was adjusted
to 9.5 with 769 ~11 (4.39 mml) of diisopropylethyl~mine and the mixtllre
stirred at 25C for 18 hours.
DMF was removed in vacuo and the crude residue treated with
20 water and extracted with ethyl acetate (3x). The oganic extracts were
combined, washed with water (lx), brine (lx), dried over Na2SO4, filtered
and stripped to dryness in vacuo. Flash chromatography of
the crude product on silica gel (12% Et2O in CH2Cl2) gave the title
compound as a white foam (318 mg, 59.3% yield) upon coevaporation with
25 ether (3x) in vacuo.
M.W.: 450.52.
m.p.: 50-64C.
HPLC: 94.9%.
PMR: Consistent with structure plus water.
M.S.: M+H = 451 (FAB).
TLC: Rf = 0.27, silica GF (15% Et2O in CH2C12)-
.
WO 94/07496 2 1 ~ 3 1 ~ 7 PCI/US93/09152
- 33 -
CHN: Calc'd as C28H26N402-1.30 H20 (F.W. = 473.965):
C, 70.95; H, 6.08; N, 11.82.
Found: C,70.98; H, 5.81; N, 12.00.
EXAMPLE 12
~ N~N>
60 mg (0.188 mmol) of the product of Example 1 was
15 dissolved in 3 ml DMF and the solution treated with 36.6 mg (0.226 mmol)
of benzoimidazole-5-carboxylic acid followed by 30.5 mg (0.226 mmol) of
l-hydroxybenzotriazole hydrate (HBT) and 43.3 mg (0.226 mmol) of 1-
ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC). The
pH was adjusted to 9.5 with 63.0 ~1 (0.453 mmol) of triethyl~mine and the
20 mixture stirred at 25C for 20 hours.
A second portion consisting of 18.3 mg (0.226 mmol) of
benzoimidazole-5-carboxylic acid, 15.3 mg (0.113 mmol) HBT, 21.7 mg
(0.113 mmol) EDC and 31.5 ~11 (0.227 mmol) of triethyl~mine was added
and the reaction stirred at 25C for 6 hours.
DMF was removed in vacuo and the residue treated with
water and extracted with ethyl acetate (3x). The organic extracts were
combined, washed with water (lx), brine (1x), dried over Na2SO4, filtered
and stripped to dryness in vacuo. Flash chromatography of the crude
product on silica gel (100:10:1 of CH2Cl2:MeOH:Conc. NH40H) gave the
title compound as a crystalline solid (51.5 mg, 59.1% yield) from ethyl
acetate.
W094/07496 ~ PCr/lJS93/0915
- 34 -
M.W.: 462.53.
m.p.: 212-6C.
HPLC: 98.8%. ~ .
PMR: Consistent with structure.
M.S.: M+H=463 (FAB).
TLC: Rf = 0.34, silica GF (80:10:1 of C12:MeOH:Conc NH4OH).
CHN: Calc'd as C29H26N402: (m.w. - 462.53):
C, 75.30; H, 5.67; N, 12.11.
Found: C, 75.18; H, 5.66; N, 12.06.
EXAMPLE 13
~ I
O~N CH3
~f CH3
O
100 mg (0.451 mmol) of spiro(lH-indene-1,4'-piperidine)
hydrochloride was dissolved in 3 ml DMF N-ethoxycarbonyl-a-(4-
Acet~miclophenyl)-glycine followed by 229 mg (0.519 mrnol) of
2 5 benzotriazol- 1 -yl-oxytris(dimethylamino)phosphonium hexafluoro-
phosphate (BOP Reagent). The pH of the solution was adjusted to
9.5 with 166 ~11 (0.947 mmol) of diisopropylethyl~mine and the
mixture stirred at 25C for 1 hours.
DMF was removed in vacuo and the crude residue treated with
30 water and extracted with ethyl acetate (3x). The organic extracts were
combined, washed with water (lx), brine (1x), dried over Na2SO4, filtered
and stripped to dryness in vacuo. Flash chromatography of the crude
2143117
WO 94/07496 - PCTtUS93/091~2
product on silica gel (3% MeOH in CH2Cl2) gave the title compound
isolated as a crystalline solid (139 mg, 68.8% yield) from ether.
M.W.: 447.516.
m.p.: 216-8C.
HPLC: 98.1 %.
PMR: Consistent with structure.
M.S.: M+H=448.1 (FAB).
TLC: Rf = 0.27, silica GF (4% MeOH in CH2Cl2).
CHN: Calc'd as C26H29N304 (F.W. - 447.516):
C, 69.78; H, 6.53; N, 9.39.
Found: C, 69.67; H, 6.51; N, 9.55.
EXAMPLE 14
~ H
74.4 mg (0.170 mmol) of the product of Example 9 was
dissolved in 10 ml of absolute ethanol, treated with 15 mg 10% Pd on C
25 under a nitrogen atmosphere, and hydrogenated at 60 psi for 3 hours. The
mixhlre was filtered through SoLka Floc~ to remove catalyst and the filter
pad was washed thoroughly with fresh absolute ethanol. The filtrate was
evaporated to dryness in vacuo and the residue was flash chromatographed
on silica gel (60:10:1 of CH2Cl2:MeOH:H2O:HoAc). The product
30 fractions were combined, stripped to dryness in vacuo, and the residue
treated with dilute NaHCO3(aq) and extracted with ethyl acetate (3x). The
organic extracts were combined, washed with water (1x), dried over
WO 94/07496 PCl /US93/09~5~
~43~
- 36 -
Na2SO4, filtered and stripped to dryness in vacuo. The residue was
coevaporated with ether (3x), then tri~rated with ether and filtered to give
the title compound as a white solid (44.8 mg, 59.7% yield).
M.W.: 442.542.
m.p.: 89-128C.
HPLC: 99.6%.
PMR: Consistent wi~ structure plus e~er, ethyl acetate and water.
M.S.: M+H=443.1 (FAB).
o TLC: Rf = 0.29, silica GF (60:10:1:1 of CH2Cl2:MeOH:H2O:HoAc).
CHN: Calc'd as C27H30N4O2-0.15 C4HloO-O.10 C4HgO2-0.35
H20:
C, 71.74; H,7.10; N, 11.95.
Found: C, 71.68; H,7.05; N, 11.75.
EXAMPLE 15
~3~ N~
2s 50 mg (0.157 mmol) of the product of F~mple 1 was
dissolved in 2 ml DMF and the solution treated with 27.8 mg (0.173 mmol)
of indole-5-carboxylic acid followed by 23.3 mg (0.173 mmol of
l-hydroxybenzotriazole hydrate (HBT) and 33.1 mg (0.173 mmol) of
1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC).
The pH of the solution was adjusted to 9.5 with 49 ~1 (0.350 mmol) of
triethyl~mine and the reaction stirred at 25C for 1 hour.
WO 94/07496 2 1 4 3 1 1 7 PCr/US93/09152
A second portion consisting of 13.0 mg (0.081 mmol) of
indole-5-carboxylic acid, 12.0 mg (0.089 mmol) HBT, 15.0 mg (0.078
mmol) EDC, and 22.1 lul (0.159 mmol) of triethyl~min~ was added and ~e
reaction stirred at 25C for 1 hour. DMF was removed in vacuo and the
5 residue treated with water and extracted with ethyl acetate (3x). The
organic layers were combined, washed with water (lx), brine (1x), dried
over Na2SO4, filtered and stripped to dryness in vacuo. Flash
chromatography of the crude product on silica gel (230:10:1 of
CH2Cl2:MeOH:Conc. NH40H) gave the title compound as an off-white
solid (30.0 mg, 37.6% yield) upon trituration with ethyl acetate.
M.W.: 461.54.
m.p.: 241-3C.
HPLC: 97.7%.
PMR: Consistent with structure plus water.
M.S.: M+H=462 (FAB).
TLC: Rf = 0.25, silica GF (200:10:1 of CH2Cl2:MeOH:conc.
NH4OH)
CHN: Calc'd as C30H27N302-0.75 H2O. (F.W.=492.701):
C, 75.08; H, 6.16; N, 8.53.
Found: C, 74.97; H,. 5.80; N, 8.47.
~XAMPLE 16
~ O
~HN ~cN N
WO 94/07496 PCI/US93/09152
~43~
- 38 -
55 mg (0.173 mmol) of the product of Example 1 was
dissolved in 2 ml DMF and the solution was treated with 33.9 mg (0.208
mmol) of benzotriazole-5-carboxylic ac~d~followed by 28.1 mg (0.208
mmol) of l-hydroxybenzotriazole hydrate (HBT) and 39.9 mg (0.208
s mmol of l-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride
(EDC). The pH of the solution was adjusted to 9.5 with 66.6 ~1 (0.478
mmol) of triethylamine and the reaction stirred at 25C for 18 hours.
DMF was removed in vacuo and the residue treated with water
and extracted with ethyl acetate (3x). The organic extracts were combined,
washed with water (lx), brine (lx), dried over Na2SO4, filtered and
stripped in dryness in vacuo. Flash chromatography of the crude product
on silica gel (88:10:1 of CH2C12:MeOH: Conc. NH40H) gave the title
compound as a crystalline solid (39.8 mg, 49.8% yield from ethyl acetate.
M.W.: 463.52.
m.p.: 229-32C.
HPLC: 99.6%
PMR: Consistent with structure plus ethyl acetate.
M.S.: M+H=464 (FAB).
TLC: Rf = 0.33, silica GF (80:10:1 of CH2C12:MeOH:Conc.
NH40H)
CHN: Calc'd as C2gH25N5O2-0.05 C4H802 (F.W. = 467.948):
C, 72.38; H, 5.47; N, 14.97.
Found: C, 72.39; H, 5.33; N, 14.92.
~ WO 94/07496 2 1 ~ 3 1 1 7 PCI /US93/09152
- 39 -
EXAMPLE 17
~ ~ ~[OCH
55.0 mg (0.173 mmol) of the product of F.x~mrle 1 was
dissolved in 3 ml CH2C12 and the solution treated with 37.9 mg (0.189
mmol) of 3,4-dimethoxybenzoyl chloride. The pH of the solution was
adjusted to 9.5 with 28.0 ,ul (0.201 mmol) of triethyl~mine and the mixture
stirred at 25C for 15 ..~ les. Flash chromatography of the reaction
mixture directly on silica gel (40% Et2O in CH2C12) gave the title
compound as a white solid (20.0 mg, 24.0% yield) upon trituration with
ether.
M.W.: 482.556.
m.p.: 107-18C (shrink).
HPLC: 99.6%.
PMR: Consistent with structure plus ether and water.
M.S.: = M+H = 483 (FAB).
TLC: Rf = 0.26, silica GF (40% Et20 in CH2C12).
CHN: Calc'd as C30H30N2O4-0.15 C4HloO-0.35 H20
(F.W.=500.008):
C, 73.50; H, 6.49; N, 5.60.
Found: C, 73.48; H, 6.31; N, 5.55.
wo 94,07496 ~ 43~ ~ ~ PCI/US93/09152
- 40 -
FXAMPLE 18
s ~ J~
60 mg (0.188 mmol) of ~e product of Example 1 was
dissolved in 2 ml CH2CL2 and the solution treated wi~ 36.9 mg (0.207
mmol) of nicotinoylchloride hydrochloride. The pH of the mi~ture was
adjusted to 9.5 with 58 ~11 (0.416 mmol) of trie~yl amine and the reaction
stirred at 25C for 18 hours.
Flash chromatography of ~e reaction mixtllre directly on
silica gel (4% MeOH in CH2Cl2) gave the title compound as a white solid
(56.8 mg, 71.3% yield) crystallized from ether.
M.W.: 423.494.
m.p.: 205-6C.
HPLC: 99.7 % .
PMR: Consistent with structure.
M.S.: M+H=424 (FAB).
TLC: Rf = 0.26, silica GF (5% MeOH in CH2C12).
2 CHN: Calc'd as C27H25N302:
C, 76.57; H, 5.92; N, 9.92.
Found: C, 76.50; H, 5.90; N, 9.91.
WO 94/07496 21 ~ 31 I 7 PCr/US93tO9152
- 41 -
EXAMPLE 19
S ~
60 ml (0.188 mmol) of the product of Example 1 was
dissolved in 2 ml CH2Cl2 and the solution treated with 36.9 mg (0.207
mmol) of isonicotinoyl chloride hydrocloride. The pH of the mix~lre was
adjusted to 9.5 with 58 ,ul (0.416 mmol) of trie~yl~mine and ~e reaction
stirred at 25C for 1 hour.
Flash chromatography of the reaction mixture directly on
silica gel (5% MeOH in CH2CL2) gave the title compound as a white solid
(56.0 mg, 70.4% yield) crystallized from ether.
M.W.: 423.494.
m.p.: 224-6C.
HPLC: 99.7%
PMR: Consistent with structure.
M.S.: M+H=424 (FAB).
TLC: Rf = 0.23, silica GF (5% MeOH in CH2cl2).
C, 76.57; H, 5.92; N, 9.92.
Found: C, 76.22; H, 5.95, N, 9.74.
WO 94/07496 PCT/US93/0915~
~43~
- 42 -
TABLES
~'' .
In addition to those compounds specifically exemplified above,
additional compounds of the present invention are set forth in tabular form
5 below. These compounds are synthesized by use of the synthetic routes and
methods described in the above Schemes and Examples and variations
thereof well known to those of ordinary skill in the art, and not requiring
undue experimentation. All variables listed in Table 1 below are with
reference to the following generic structure:
~1
o~X,Rl
~ 21 ~ ~ I 1 7 PCI/US93/09152
WO 94/07496
- 43 -
TABLE 1
X
-CH2- ~
NH2
o -CH2- H3CCHN~
-CH2- H2N~3
no substituentH3CCHN~3
-CH2-
NH2
~~--NH2
-CH2- ~J
WO 94/07496~,~ 43~ PCl/US93/091
- 44 -
TABLE 1 ~CONTD)
X .~ R
-CH2- H
-CH2- ~ CH,
-CH2- ~--NH CH3
1 s -CH2-
HN ~n,CH3
-CH2- ~ , N~o CH3
2s no substituent ~3~' CH3
o
~ NH2
-CH2-
-CH2-
~ 2I93ll7
WO 94/07496 PCI`/US93/09152
- 45 -
TABLE 1 (CONT'D)
X Rl
CH3
-CH2- ~ ~CH3
o ~N>
-CH2- ~ O
~l~H3C CH3
-CH2- ~NH ~ NH2
no substituent ~NH2
-CH2- ~3`HCCH=CH~ q
NH
no substituent ~3--NH CH3
CH W~ NO2
PCI /US93/09152
WO 94/07496
~43~
- 46 -
TABLE 1 (CONT'D)
X ' Rl
no substituent ~3~H~H3
~ o
-CH2- H ~N~
H
CH; ~NH2
CH; ~NH CH3
2s -CH2`
-CH2- [~ N ~` H
~ WO 94/07496 21 ~ 31 1 7 PCI/US93/09152
- 47 -
TABLE 1 (CONT'D)
X Rl
-CH2-~ ~ NN~H
o O CH3
-CH2-
2 o -CH=CH-~1~ NO2
-CH2- H~CN
WO 94/07496 PCI`/US93/Ogl5~
3~
- 48 -
TABLE 2
The variables shown in Table 2 are with reference to the
following structure:
N
~! 214317
WO 94/07496 - PCT/US93/09152
- 49 -
Y Y
OH
/so2~3 /SO2~CI
~SO~ /SO2~F
/S2~ /S2~
OH
~S~ ~SO2~
O-CH3 HO
W094/07496 ~43~'1 PCI/US93/0915~
- 50 -
TABLE 2 (CONT'D)
Y Y
Cl NH2
~SO2~ ` /S02~
O
~ ~S CCHH33 /S2~
HN ~CH3
o
H3C CH3
H3C ~CH3 ~ NH CH3
CH3 CH3
/SO2~ `l3`NH2
Z5 ~5O2~D`NH~0 ~CH
WO 94/07496 2 1 ~ 3 1 1 7 PCI /US93/09152
- 51 -
TABLE 2 (CONT'D)
y Y
5~SO2~l3` N' 2 CH3 --~H3CH3
O H3C
~Z~ NJ~CH3 ~SO~CH3
H3C CH3 H3C CH3
,CH3
~OZ~o~cH3 S ~ 2
~SO2~ ~SO2 NH~Z~so2 CH3
H3C CH3
~SO2~,~NH2
WO 94/07496 PCTtUS93/0915~
2i9~3~
- 52 -
TABLE 2 (CONTD)
Y y
". CH3
~SO2 NH~ ~S2~_~o~CH3
H3C N
1 o--SO2~o~cH3
~H~
o,~ ,S2 ~H)~
~S2--N~fH3 ~N~NH
H3C
WO 94/07496 PCI`/US93/09152
21~3117
- 53 -
TABLE 3
The variables shown in Table 3 are with reference to the
following structure:
[~CH3
~N~
J~ Z ~
O H
o~o ~ H
2 o 1 ~,N~N~
NH H3C
2 5 ~ J~H3 H3C ~
O H3C CH3
Dg~oJ~CH3 H
WO 94/07496 PCI /US93/09151--
p'L~3~ 54-
o
0 J~ ,~H3CH
0~ ,NH
O
~3
[~ ¢~
WO 94/07496 PCI/US93/09lS2
21~311 7
- 55 -
O Z Z
5 J~ ~
o~N~ SO2
N CH3
O H
10 ,J~
od` ~N~ N~
N
O H
Il O
2 o S2 N~
OH
o CH3
~
~--13
WO 94/07496 PCI /US93/09152--
2~43~
- - 56 -
TABLE 3 (CONT'D)
7 `` 7.
O . O
O~N~0
'J~ J~
H ~[3 od~r SO2~CH3
HN O~,CH3
~ rCH3
, O CH3
O~CH3
pH HO~
O N O N
~CH3 ~CH3
PCl~/US93/09152
WO 94/07496 2 1 ~ 3 1 1 7
J~l
~I~O~,CH3
o
~,
O~N
N
H
~NJ
o~,SO
NH2
SO2
W O 94/07496PC~r/US93/0915 ~
~43~
- 58 -
FXAMPLE 20
RADIOLIGAND BINDING ASSAYS
5The high affinity binding of [3H] Oxytocin (OT)([tyrosyl, 3,5-
[3H]oT; 30-60 Ci/mmol; New Fn~l~nd Nuclear. Boston, MA) to uterine
OT receptors was based on an assay (Fuchs, A-R; Fuchs, F; Soloff, MS.
1985 J. Clin. Endocrinol. Metab. 60:37) using a crude membrane
preparation of uteri taken from diethylstilbestrol dipropionate (DES)-
treated (0.3 mg/kg, ip; 18-24) rats. Cornpetition studies were conducted at
equilibrium (60 minlltes; 22C) using 1 nM~3H]OT in the following assay
rrel. 50 mM Tris-HCl, 5 mM MgC12, and 0.1% BSA, pH 7.4.
Nonspecific binding (10% of the total binding) was determined using 1 ,uM
unlabeled OT and the binding reaction was termin~ted by filtration through
glass fiber filters using a cell harvester (model 7019, Skatron, l[nc.,
Sterling, VA). ICso (the concentration of tested compound that inhibits
50% of OT) was reported, unless otherwise noted.
The measurement of [3H]Vasopressin (AVP) ([phenylalanyl-
3,4,5-3H]AVP; 80-90 Ci/mmol, New Fngl~nd Nuclear)binding to a crude
membrane preparation of male rat liver (AVP-Vl sites) or kidney medulla
(AVP-V2 sites) was determined according to the method of Butlen, et al.
(Butlen, D; Guillon, G; Rajerison, R.M.;Jard, S; Sawyer, W.H.;~I~nning,
M. 1978 Mol Ph~ col 14:1006).
Competition assays were conducted at equilibrium (30 minlltes
at 30C) using 1 nM [3H]AVP (liver) or 2 nM [3H]AVP (kidney) in the
following assay burrel. 100 mM Tris-HCl, S mM MgC12, 0.1% BSA, 50
mM phenylmethylsulfonylfluoride, and 50 mg/ml bacitracin, pH 8Ø
Nonspecific binding (5-10% of the total binding) was determined using 10
,uM unlabeled AVP, and the binding reaction was termin~ted by filtration
as described above for the [3H]oT binding assay.
2l~3ll 7
WO 94/07496 PCI /US93/09152
_ 59 _
ICso values were determined for both [3H]oT and [3H]AVP
binding assays by linear regression of the relation log concentration of
compound vs. percent inhibition of specific binding.
Example ICso[3HlOT (nM)
3~700
12 150
14 170
310
16 520
While the invention has been described and illustrated with
5 reference to certain preferred embodiments thereof, those skilled in the art
will appreciate that various changes, modifications and substitutions can be
made therein without departing from the spirit and scope of the invention.
For example, effective dosages other than the preferred dosages as set forth
hereinabove may be applicable as a consequence of variations in ~e
20 responsiveness of the m~mm~l being treated for prevention of prete~n
labor, or for other indications for the compounds of the invention indicated
above. Likewise, the specific ph~rm~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
25 the type of form~ tion and mode of ~(lmini~tration employed, and such
expected variations or differences in the results are collt~nl~lated in
accordance with the objects and practices of the present invention. It is
intended, therefore, that the invention be limited only by the scope of the
claims which follow and that such claims be interpreted as broadly as is
reasonable.
