Note: Descriptions are shown in the official language in which they were submitted.
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TITLE OF THE INVENTION
HETEROCYCLIC THIOESTERS AND KETONES
This application is a divisional of Canadian patent application Serial No.
2,263,927 filed internationally on September 9, 1997 and entered nationally on
February 23, 1999.
BACKGROUND OF THE INVENTION
1. Field of Invention
This invention relates to neurotrophic low molecular weight, small
molecule heterocyclic thioesters and ketones having an affinity for FKBP-type
immunophilins, and their use as inhibitors of the enzyme activity associated
with immunophilin proteins, particularly peptidyl-prolyl isomerase, or
rotamase,
enzyme activity.
2. Description of Related Art
The term immunophilin refers to a number of proteins that serve as
receptors for the principal immunosuppressant drugs, cyclosporin A (CsA),
FK506 and rapamycin. Known classes of immunophilins are cyclophilins and
FK506 binding proteins, or FKBPs. Cyclosporin A binds to cyclophilin A while
FK506 and rapamycin bind to FKBP12. These immunophilin-drug complexes
interface with various intracellular signal transduction systems, especially
the
immune and nervous systems.
Immunophilins are known to have peptidyi-prolyl isomerase (PPIase), or
rotamase, enzyme activity. It has been determined that rotamase enzyme
activity plays a role in the catalyzation of the interconversion of the cis
and
trans isomers of peptide and protein substrates for the immunophilin proteins.
Immunophilins were originally discovered and studied in the immune
tissue. It was initially postulated by those skilled in the art that
inhibition of the
immunophilins' rotamase activity leads to inhibition of T-cell proliferation,
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thereby causing the immunosuppressive activity exhibited by immuno-
suppressant drugs, such as cyclosporin A, FK506 and rapamycin. Further study
has shown that the inhibition of rotamase activity, in and of itself, does not
result in immunosuppressive activity. Schreiber et al., Science, 1990, vol.
250,
pp. 556-559. Instead, immunosuppression appears to stem from the formula-
tion of a complex of immunosuppressant drug and immunophilin. It has been
shown that the immunophilin-drug complexes interact with ternary protein
targets as their mode of action. Schreiber et al., Cell, 1991, vol. 66, pp.
807-
815. In the case of FKBP-FK506 and cyclophilin-CsA, the immunophilin-drug
complexes bind to the enzyme calcineurin and inhibit the T-cell receptor
signalling which leads to T-cell proliferation. Similarly, the immunophilin-
drug
complex of FKBP-rapamycin interacts with the RAFT1/FRAP protein and inhibits
the IL-2 receptor signalling.
Immunophilins have been found to be present at high concentrations in
the central nervous system. Immunophilins are enriched 10-50 times more in
the central nervous system than in the immune system. Within neural tissues,
immunophilins appear to influence nitric oxide synthesis, neurotransmitter
release and neuronal process extension.
It has been found that picomolar concentrations of an immunosuppres-
sant such as FK506 and rapamycin stimulate neurite outgrowth in PC12 cells
and sensory neurons, namely dorsal root ganglion cells (DRGs). Lyons et al.,
Proc. of Natl. Acad. Sci., 1994, vol. 91, pp. 3191-3195. In whole animal
experiments, FK506 has been shown to stimulate nerve regeneration following
facial nerve injury.
Surprisingly, it has been found that certain compounds with a high affinity
for FKBPs are potent rotamase inhibitors and exhibit excellent neurotrophic
effects. Furthermore, these rotamase inhibitors are devoid of immuno-
suppressive activity. These findings suggest the use of rotamase inhibitors in
treating various peripheral neuropathies and enhancing neuronal regrowth in
the central nervous system (CNS). Studies have demonstrated that neuro-
degenerative disorders, such as Alzheimer's disease, Parkinson's disease and
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amyotrophic lateral sclerosis (ALS), may occur due to the loss, or decreased
availability, of a neurotrophic substance specific for a particular population
of
neurons affected in the disorder.
Several neurotrophic factors affecting specific neuronal populations in the
central nervous system have been identified. For example, it has been hypo-
thesized that Alzheimer's disease results from a decrease or loss of nerve
growth factor (NGF). It has thus been proposed to treat SDAT patients with
exogenous nerve growth factor or other neurotrophic proteins, such as brain
derived growth factor, glial derived growth factor, ciliary neurotrophic
factor and
neurotropin-3, to increase the survival of degenerating neuronal populations.
Clinical application of these proteins in various neurological disease states
is hampered by difficulties in the delivery and bioavailability of large
proteins to
nervous system targets. By contrast, immunosuppressant drugs with neuro-
trophic activity are relatively small and display excellent bioavailability
and
specificity. However, when administered chronically, immunosuppressant drugs
exhibit a number of potentially serious side effects including nephrotoxicity,
such as impairment of glomerular filtration and irreversible interstitial
fibrosis
(Kopp et al., J. Am. Soc. Nephrol., 1991, 1:162); neurological deficits, such
as
involuntary tremors, or non-specific cerebral angina, such as non- localized
headaches (De Groen et al., N. Engl. J. Med., 1987, 317:861); and vascular
hypertension with complications resulting therefrom (Kahan et al., N. Engl. J.
Med., 1989, 321:1725).
To prevent the side effects associated with use of the immunosuppressant
compounds, the present invention provides non-immunosuppressive compounds
containing small molecule FKBP rotamase inhibitors for enhancing neurite out-
growth, and promoting neuronal growth and regeneration in various neuropath-
ological situations where neuronal repair can be facilitated, including:
peripheral
nerve damage caused by physical injury or disease state such as diabetes;
physical damage to the central nervous system (spinal cord and brain); brain
damage associated with stroke; and neurological disorders relating to
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neurodegeneration, such as Parkinson's disease, SDAT (Alzheimer's disease)
and amyotrophic lateral sclerosis.
SUMMARY OF THE INVENTION
The present invention relates to neurotrophic low molecular weight, small
molecule compounds having an affinity for FKBP-type immunophilins. Once
bound to these proteins, the neurotrophic compounds are potent inhibitors of
the enzyme activity associated with immunophilin proteins, particularly
peptidyl-
prolyl isomerase, or rotamase, enzyme activity. A key feature of the compounds
of the present invention is that they do not exert any significant immuno-
suppressive activity in addition to their neurotrophic activity.
Specifically, the present invention relates to a compound of formula II:
[CH2] n
CZ-Rl
II
D x
O
R2
or a pharmaceutically acceptable salt thereof, wherein:
n is 1 or 2;
XisOorS;
Z is selected from the group consisting of S, CH2, CHR1, and C(Rl)Z;
Rl is selected from the group consisting of C1-C5, straight or branched
chain alkyl, C2-C5 straight or branched chain alkenyl, Ar1 and mixtures
thereof,
wherein said Ri is unsubstituted or substituted with halo, nitro, C1-C6
straight or
branched chain alkyl, C2-C3 straight or branched chain alkenyl, hydroxy, C1-C4
alkoxy, C2-C4
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alkenyloxy, phenoxy, benzyloxy, amino, Ari or a mixture thereof;
R2 is selected from the group consisting of C1-C9 straight or branched
chain alkyl, C2-C9 straight or branched chain alkenyl, C3-C8 cycloalkyl, C5-C7
cycloalkenyl and Arl; and
Arl is phenyl, benzyl, pyridyl, fluorenyl, thioindolyl or naphthyl wherein
said Arl is unsubstituted or substituted with halo, hydroxy, nitro, C1-C6
straight
or branched chain alkyl, C2-C6 straight or branched chain alkenyl, C1-C4
alkoxy,
C2-C4 alkenyloxy, phenoxy, benzyloxy, amino or a mixture thereof.
The present invention also relates to a pharmaceutical composition
comprising:
(i) an effective amount of the compound of formula II for effecting a
neuronal activity; and
(ii) a pharmaceutically acceptable carrier.
The present invention further relates to a method of effecting a neuronal
activity in an animal, comprising:
administering to the animal an effective amount of the compound of
formula II.
This is a second divisional of application No. 2,263,927 and relates to
processes to synthesize pyrrolidine derivatives having use as intermediates.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1(A) is a representative photomicrograph of untreated sensory
neurons.
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FIG. 1(B) is a representative photomicrograph of compound 1 (10 pM)
promoting neurite outgrowth in sensory neurons.
FIG. 1(C) is a representative photomicrograph of compound 1 (1 nM)
promoting neurite outgrowth in sensory neurons.
FIG. 1(D) is a representative photomicrograph of compound 1 (1 pM)
promoting neurite outgrowth in sensory neurons.
FIG. 2(A) is a representative photomicrograph of untreated sensory
neurons.
FIG. 2(B) is a representative photomicrograph of compound 9 (10 pM)
promoting neurite outgrowth in sensory neurons.
FIG. 2(C) is a representative photomicrograph of compound 9 (1 nM)
promoting neurite outgrowth in sensory neurons.
FIG. 2(D) is a representative photomicrograph of compound 9 (100 nM)
promoting neurite outgrowth in sensory neurons.
FIG. 3(A) is a representative photomicrograph of untreated sensory
neurons.
FIG. 3(B) is a representative photomicrograph of compound 10 (10 pM)
promoting neurite outgrowth in sensory neurons.
FIG. 3(C) is a representative photomicrograph of compound 9 (1 nM)
promoting neurite outgrowth in sensory neurons.
FIG. 3(D) is a representative photomicrograph of compound 9 (100 nM)
promoting neurite outgrowth in sensory neurons.
FIG. 4 presents quantitation for the recovery of TH-positive dopaminergic
neurons in the striatum of animals receiving compounds 1, 9 and 10.
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DETAILED DESCRIPTION OF THE INVENTION
Definitions
"Alkyl" refers to a branched or unbranched saturated hydrocarbon chain
containing 1 to 6 carbon atoms, such as methyl, ethyl, propyl, iso-propyl,
butyl,
iso-butyl, tert-butyl, n-pentyl, n-hexyl and the like, unless otherwise
indicated.
"Alkoxy" refers to the group -OR wherein R is alkyl as herein defined.
Preferably, R is a branched or unbranched saturated hydrocarbon chain
containing 1 to 3 carbon atoms.
"Halo" refers to fluoro, chloro, bromo or iodo, unless otherwise indicated.
"Isomers" are different compounds that have the same molecular
formula. "Stereoisomers" are isomers that differ only in the way the atoms are
arranged in space.
"Enantiomers" are a pair of stereoisomers that are non-superimposable
mirror images of each other. "Diastereoisomers" are stereoisomers which are
not mirror images of each other. "Racemic mixture" means a mixture containing
equal parts of individual enantiomers. "Non-racemic mixture" is a mixture con-
taining unequal parts of individual enantiomers or stereoisomers.
"Pharmaceutically acceptable salt" refers to a salt of the inventive com-
pounds which possesses the desired pharmacological activity and which is
neither biologically nor otherwise undesirable. The salt can be formed with
inorganic acids such as acetate, adipate, alginate, aspartate, benzoate,
benzenesulfonate, bisulfate butyrate, citrate, camphorate, camphorsulfonate,
cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate,
fumarate, glucoheptanoate, glycerophosphate, hemisulfate heptanoate,
hexanoate, hydrochloride hydrobromide, hydroiodide, 2-hydroxyethane-
sulfonate, lactate, maleate, methanesulfonate, 2-naphthalenesulfonate,
nicotinate, oxalate, thiocyanate, tosylate and undecanoate. Examples of a base
salt include ammonium salts, alkali metal salts such as sodium and potassium
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salts, alkaline earth metal salts such as calcium and magnesium salts, salts
with
organic bases such as dicyclohexylamine salts, N-methyl-D-glucamine, and salts
with amino acids such as arginine and lysine. Also, the basic nitrogen-
containing
groups can be quarternized with agents including: lower alkyl halides such as
methyl, ethyl, propyl and butyl chlorides, bromides and iodides; dialkyl
sulfates
such as dimethyl, diethyl, dibutyl and diamyl sulfates; long chain halides
such
as decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides; and
aralkyl halides such as benzyl and phenethyl bromides.
"Phenyl" refers to any possible isomeric phenyl radical, optionally mono-
substituted or multisubstituted with substituents selected from the group
consisting of alkyl, alkoxy, hydroxy, halo and haloalkyl.
"Treating" refers to:
(i) preventing a disease, disorder or condition from occurring in an
animal which may be predisposed to the disease, disorder and/or condition but
has not yet been diagnosed as having it;
(ii) inhibiting the disease, disorder or condition, i.e., arresting its
development; and
(iii) relieving the disease, disorder or condition, i.e., causing regression
of the disease, disorder and/or condition.
Compounds of the Invention
The neurotrophic low molecular weight, small molecule FKBP inhibitor
compounds of this invention have an affinity for FKBP-type immunophilins, such
as FKBP12. When the neurotrophic compounds of this invention are bound to an
FKBP-type immunophilin, they have been found to inhibit the prolyl-peptidyl
cis-
trans isomerase activity, or rotamase, activity of the binding protein and un-
expectedly stimulate neurite growth.
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FORMULA I
In particular, this invention relates to a compound of formula I:
B
p ~ Z~
N I R1 I
Y X
W
R2
or a pharmaceutically acceptable salt thereof, wherein:
A and B, together with the nitrogen and carbon atoms to which they are
respectfully attached, form a 5-7 membered saturated or unsaturated hetero-
cyclic ring containing any combination of CH2, 0, S, SO, SO2, NH or NR2 in any
chemically stable oxidation state;
X is either 0 or S;
Z is either S, CH2, CHRl or C(R1)2;
W and Y are independently 0, S, CH2 or HZ;
R1 is C1-C6 straight or branched chain alkyl or alkenyl, which is sub-
stituted in one or more position(s) with (Arl)rõ (Arl)n connected by a C1-C6
straight or branched chain alkyl or alkenyl, C3-C8 cycloalkyl, C3-C8
cycloalkyl
connected by a C1-C6 straight or branched chain alkyl or alkenyl, Ar2 or a
combination thereof;
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n is 1 or 2;
R2 is either C1-C9 straight or branched chain alkyl or alkenyi, C3-Cg
cycloalkyl, C5-C7 cycloalkenyl or Arl, wherein said alkyl, alkenyl, cycloalkyl
or
cycloalkenyl is either unsubstituted or substituted in one or more position(s)
with C1-C4 straight or branched chain alkyl or alkenyl, hydroxyl or a
combination
thereof; and
Arl and Ar2 are independently a mono-, bi- or tricyclic, carbo- or hetero-
cyclic ring, wherein the ring is either unsubstituted or substituted in one to
three
position(s) with halo, hydroxyl, nitro, trifluoromethyl, CI-C6 straight or
branched
chain alkyl or alkenyl, C1-C4 alkoxy, C1-C4 alkenyloxy, phenoxy, benzyloxy,
amino or a combination thereof; wherein the individual ring sizes are 5-6
members; and wherein the heterocyclic ring contains 1-6 heteroatom(s)
selected from the group consisting of 0, N, S and a combination thereof.
Suitable mono- and bicyclic, carbo- and heterocyclic rings include, with-
out limitation, naphthyl, indolyl, furyl, thiazolyl, thienyl, pyridyl,
quinolinyl,
isoquinolinyl, fluorenyl and phenyl.
FORMULA II
A preferred embodiment of this invention is a compound of formula II:
[CH2] n
CZ-R1
N
II
o~ x
0
R2
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or a pharmaceutically acceptable salt thereof, wherein:
nislor2;
XisOorS;
Z is selected from the group consisting of S, CH2, CHRl and C(Rl)2;
Rl is selected from the group consisting of C1-C5 straight or branched
chain alkyl, C2-C5 straight or branched chain alkenyl, Ar1 and mixtures
thereof,
wherein said Rl is unsubstituted or substituted with halo, nitro, C1-C6
straight or
branched chain alkyl, C2-C6 straight or branched chain alkenyl, hydroxy, C1-C4
alkoxy, C2-C4
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alkenyloxy, phenoxy, benzyloxy, amino, Arl or a mixture
thereof;
R2 is selected from the group consisting of C1-C9
straight or branched chain alkyl, C2-C9 straight or
branched chain alkenyl, C3-C8 cycloalkyl, C5-C7
cycloalkenyl and Arl; and
Arl is phenyl, benzyl, pyridyl, fluorenyl,
thioindolyl or naphthyl wherein said Arl is unsubstituted
or substituted with halo, trifluoromethyl, hydroxy,
nitro, C1-C6 straight or branched chain alkyl, C2-C6
straight or branched chain alkenyl, C1-C4 alkoxy, C2-C4
alkenyloxy, phenoxy, benzyloxy, amino or a mixture
thereof.
Specific examples of these embodiments are
presented in TABLE I.
TABLE I
No. n X Z R, RZ
1 1 0 CH2 3-Phenylpropyl 1,1-Dimethylpropyl
2 1 0 CH2 3-(3-Pyridyl)propyl 1,1-Dimethylpropyl
3 1 0 CH2 3-Phenylpropyl tert-Butyl
4 1 0 CH2 3-(3-Pyridyl)propyl tert-Butyl
5 1 0 CH2 3-(3-Pyridyl)propyl Cyclohexyl
6 1 0 CH2 3-(3-Pyridyl)propyl Cyclopentyl
7 1 0 CH2 3-(3-Pyridyl)propyl Cycloheptyl
8 1 0 CH2 2-(9-Fluorenyl) ethyl 1, 1 -Dimethylpropyl
9 1 0 S 2-Phenethyl 1,1-Dimethylpropyl
10 2 0 S 2-Phenethyl 1,1-Dimethylpropyl
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11 1 0 S Methyl(2-thioindole) 1,1-Dimethyipropyl
12 1 0 S 2-Phenethyl Cyclohexyl
13 2 0 S 2-Phenethyl tert-Butyl
14 2 0 S 2-Phenethyl Phenyl
15 1 0 CH2 3-(4-Methoxyphenyl)propyl 1,1-Dimethylpropyl
16 2 0 CH2 4-(4-Methoxyphenyl)butyl 1, 1 -Dimethylpropyl
17 2 0 CHZ 4-Phenylbutyl 1,1-Dimethylpropyl
18 2 0 CH2 4-Phenylbutyl Phenyl
19 2 0 CH2 4-Phenylbutyl tert-Butyl
20 1 S CHZ 3-Phenylpropyl 1,1-Dimethylpropyl
21 1 S S 2-Phenethyl 1,1-Dimethylpropyl
22 2 S CH2 3-Phenylpropyl 1,1-Dimethylpropyl
23 2 S S 2-Phenethyl 1,1-Dimethylpropyl
24 2 0 CHR, 3-Phenylpropyl 1,1-Dimethylpropyl
25 2 0 CHR, 3-Phenylpropyl Cyclohexyl
26 2 0 CHRI 3-Phenylpropyl Phenyl
27 2 0 CHR, 3-Phenylpropyl 3,4,5-Trimethoxyphenyl
28 1 0 S 2-Phenethyl Cyclopentyl
29 2 0 S 3-Phenylpropyl tert-Butyl
30 1 0 S 3-Piieny ipropyl 1,1-Dimethylpropyl
31 1 0 S 3-.(3-Pyridyl)propyl 1,1-Dimethylpropyl
32 1 0 S 3-Phenylpropyl Cyclohexyl
33 1 0 S 4-Phenylbutyl Cyclohexyl
34 1 0 S 4-Phenylbutyl 1,1-Dimethylpropyl
35 1 0 S 3-(3-Pyridyl)propyl Cyclohexyl
36 1 0 S 3,3-Diphenylpropyl 1,1-Dimethylpropyl
37 1 0 S 3,3-Diphenylpropyl Cyclohexyl
38 1 0 S 3-(4-Methoxyphenyl) 1,1-Dimethylpropyl
propyl
39 2 0 S 4-Phenylbutyl tert-Butyl
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40 2 0 S 1,5-Diphenyl-3-pentyl 1,1-Dimethylpropyl
41 2 0 S 1,5-Diphenyl-3-pentyl Phenyl
42 2 0 S 3-(4-Methoxyphenyl) 1,1-Dimethylpropyl
propyl
43 2 0 S 3-(4-Methoxyphenyl) Phenyl
propyl
44 2 0 S 3-(1-Naphthyl)propyl 1,1-Dimethylpropyl
45 1 0 S 3,3-Di(4-fluoro)phenyl- 1, 1 -Dimethylpropyl
propyl
46 1 0 S 4,4-Di(4-fluoro)phenyl- 1,1-Dimethylpropyl
butyl
47 1 0 S 3-(1-Naphthyl)propyl 1,1-Dimethylpropyl
48 1 0 S 2,2-Diphenylethyl 1,1-Dimethylpropyl
49 2 0 S 2,2-Diphenylethyl 1,1-Dimethylpropyl
50 2 0 S 3,3-Diphenylpropyl I,I-Dimethylpropyl
51 1 0 S 3-(4-{Trifluoromethyl}- 1,1-Dimethylpropyl
phenyl)propyl
52 1 0 S 3-(2-Naphthyl)propyl I,I-Dimethylpropyl
53 2 0 S 3-(1-Naphthyl)propyl 1,1-Dimethylpropyl
54 1 0 S, --'3-(3-Chloro)phenylpropyl 1,1-Dimethylpropyl
55 1 0 S 3-(3-{Trifluoromethyl}- 1,1-Dimethylpropyl
phenyl)propyl
56 1 0 S 3-(2-Biphenyl)propyl I,I-Dimethylpropyl
57 1 0 S 3-(2-Fluorophenyl)propyl I,I-Dimethylpropyl
58 1 0 S 3-(3-Fluorophenyl)propyl 1,1-Dimethylpropyl
59 2 0 S 4-Phenylbutyl 1,1-Dimethylpropyl
60 2 0 S 3-Phenylpropyl 1, 1-Dimethylpropyl
61 1 0 S 3-(2-Chloro)phenylpropyl 1,1-Dimethylpropyl
62 2 0 S 3-(3-Chloro)phenylpropyl 1,1-Dimethylpropyl
63 2 0 S 3-(2-Fluoro)phenylpropyl 1,1-Dimethylpropyl
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64 2 0 S 3-(3-Fluoro)phenylpropyl 1,1-Dimethylpropyl
65 1 0 S 3-(3,4-Dimethoxyphenyl)- 1, 1 -Dimethylpropyl
propyl
66 1 0 CH2 3-Phenylpropyl Cyclohexyl
67 1 0 CH2 2-Phenylethyl tert-Butyl
68 2 0 CH2 4-Phenylbutyl Cyclohexyl
69 2 0 CHR, 2-Phenylethyl tert-Butyl
70 1 0 CH2 3,3-Di(4-fluorophenyl)- 1,1-Dimethylpropyl
propyl
71 2 0 CH2 3-Phenylpropyl 1, l-Dimethylpropyl
The most preferred examples of TABLE I are named as
follows:
1 (2S)-3,3-dimethyl-l-[2-(5-phenylpentanoyl)
pyrrolidinyl]pentane-l,2-dione
2 (2S)-3,3-dimethyl-l-[2-(5-(3-pyridyl)pentanoyl)
pyrrolidinyl]pentane-l,2-dione
3 (2S) -2- (1-oXo7.5-phenyl) -pentyl-l- (3, 3-dimethyl-1, 2-
dioxobutyl)pyrrolidine
9 2-Phenyl-l-ethyl (2S)-1-(3,3-dimethyl-1,2-
dioxopentyl)-2-pyrrolidinecarbothioate
10 2-Phenyl-l-ethyl 1-(3,3-dimethyl-l,2-dioxopentyl)-
2-piperidinecarbothioate
11 1-{2-benzo[b]thiophen-3-ylmethyl.thio)carbonyl]
pyrrolidinyl}-3,3-dimethylpentane-1,2-dione
12 2-Phenyl-l-ethyl (2S)-l-(2-cyclohexyl-1,2-
dioxoethyl)-2-pyrrolidinecarbothioate
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14 2-Phenyl-l-ethyl 1-(2-phenyl-1,2-dioxoethyl)-2-
piperidinecarbothioate
28 2-Phenyl-l-ethyl (2S)-1-(2-cyclopentyl-1,2-
dioxoethyl)-2-pyrrolidinecarbothioate
29 3-Phenyl-l-propyl 1-(3,3-dimethyl-1,2-dioxobutyl)-
2-piperidinecarbothioate
30 3-Phenyl-l-propyl (2S)-1-(3,3-dimethyl-1,2-
dioxopentyl)-2-pyrrolidinecarbothioate
31 3-(3-Pyridyl)-1-propyl (2S)-1-(3,3-dimethyl-1,2-
dioxopentyl)-2-pyrrolidinecarbothioate
32 3-Phenyl-l-propyl (2S)-1-(2-cyclohexyl=1,2-
dioxoethyl)-2-pyrrolidinecarbothioate
33 4-Phenyl-l-butyl (2S)-1-(2-cyclohexyl-1,2-
dioxoethyl)-2-pyrrolidinecarbothioate
34 4-Phenyl-l-butyl (2S)-1-(3,3-dimethyl-1,2-
dioxopentyl)-2-pyrrolidinecarbothioate
35 3-(3-Pyridyl)-1-propyl (2S)-1-(2-cyclohexyl-1,2-
dioxoethyl)-2-pyrrolidinecarbothioate
36 3,3-Diphenyl-l-propyl (2S)-1-(3,3-dimethyl-1,2-
dioxopentyl)-2-pyrrolidinecarbothioate
37 3,3-Diphenyl-l-propyl (2S)-1-(2-cyclohexyl-1,2-
dioxoethyl)-2-pyrrolidinecarbothioate
38 3-(para-Methoxyphenyl)-1-propyl (2S)-1-(3,3-
dimethyl-1,2-dioxopentyl)-2-pyrrolidinecarbothioate
39 4-Phenyl-l-butyl 1-(1,2-dioxo-3,3-dimethylbutyl) -
2-piperidinecar.bothioate
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40 1,S-Diphenyl-3-pentyl 1-(3,3-dimethyl-1,2-
dioxopentyl)-2-piperidinecarbothioate
41 1,5-Diphenyl-3-pentyl 1-(2-phenyl-1,2-dioxoethyl)-
2-piperidinecarbothioate
42 3-(para-Methoxyphenyl)-1-propyl 1-(1,2-dioxo-3,3-
dimethylpentyl)piperidine-2-carbothioate
43 3-(para-Methoxyphenyl)-1-propyl 1-(2-phenyl-1,2-
dioxoethyl)piperidine-2-carbothioate
44 3-(1-Naphthyl)-1-propyl 1-(3,3-dimethyl-1,2-
dioxopentyl)piperidine-2-carbothioate
45 3,3-Di(para-fluoro)phenyl-l-propyl (2S)-1-(3,3-
dimethyl-1,2-dioxopentyl)-2-pyrrolidinecarbothioate
46 4,4-Di(para-fluorophenyl)butyl 1-(3,3-dimethyl-2-
oxopentanoyl)-2-pyrrolidinecarbothioate
47 3-(1-Naphthyl)-1-propyl (2S)-1-(3,3-dimethyl-2-
oxopentanoyl)-2-pyrrolidinecarbothioate
48 2,2-Diphenylethyl (2S)-1-(3,3-dimethyl-2-
oxopentanoyl)-2-pyrrolidinecarbothioate.
49 2,2-Diphenylethyl (2S)-1-(3,3-dimethyl-2-
oxopentanoyl)-2-piperidinecarbothioate
50 3, 3-Diphenylpropyl 1-(3,3-dimethyl-2-oxopentanoyl)-
2-piperidinecarbothioate
51 3- [4- (Trifluoromethyl) phenyl] propyl (2S) -1- (3, 3-
dimethyl-2-oxopentanoyl)-2-pyrrolidinecarbothioate
52 3-(2-Naphthyl)propyl (2S)-1-(3,3-dimethyl-2-
oxopentanoyl)-2-pyrrolidinecarbothioate
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53 3-(1-Naphthyl)-1-propyl 1-(3,3=dimethyl-2-
oxopentanoyl)-2-piperidinecarbothioate
54 3-(3-Chlorophenyl)propyl (2S)-1-(3,3-dimethyl-2-
oxopentanoyl)-2-pyrrolidinecarbothioate
55 3-[3-(Trifluoromethyl)phenyl]propyl (2S)-1-(3,3-
dimethyl-2-oxopentanoyl)-2-pyrrolidinecarbothioate
56 3-(2-Biphenyl)propyl (2S)-1-(3,3-dimethyl-2-
oxopentanoyl)-2-pyrrolidinecarbothioate
57 3-(2-Fluorophenyl)propyl (2S)-1-(3,3-dimethyl-2-
oxopentanoyl)-2-pyrrolidinecarbothioate
58 3-(3-Fluorophenyl)propyl (2S)-1-(3,3-dimethyl-2-
oxopentanoyl)-2-pyrrolidinecarbothioate
59 4-Phenylbutyl 1-(3,3-dimethyl-2-oxopentanoyl)-2-
piperidinecarbothioate
60 3-Phenylpropyl 1-(3,3-dimethyl-2-oxopentanoyl)-2-
piperidinecarbothioate
61 3-(2-Chlorophenyl)propyl (2S)-1-(3,3-dimethyl-2-
oxopentanoyl)-2-pyrrolidinecarbothioate
62 3-(3-Chlorophenyl)-1-propyl 1-(3,3-dimethyl-2-
oxopentanoyl)-2-piperidinecarbothioate
63 3-(2-Fluorophenyl)propyl 1-(3,3-dimethyl-2-
oxopentanoyl)-2-piperidinecarbothioate
64 3-(3-Fluorophenyl)propyl 1-(3,3-dimethyl-2-
oxopentanoyl)-2-piperidinecarbothioate
65 3-(3,4-Dimethoxyphenyl)propyl (2S) -1- (3, 3-dimethyl-
2-oxopentanoyl)-2-pyrrolidinecarbothioate
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66 (2S)-2-( 1-Oxo-5-phenyl) pentyl-l- (2-Cyclohexyl-1, 2-
dioxoethyl)pyrrolidine
67 2-(1-Oxo-4-phenyl)-butyl-l-(3,3-dimethyl-1,2-
dioxobutyl)pyrrolidine
68 2-(1-Oxo-6-phenyl)-hexyl-l-(2-Cyclohexyl-1,2-
dioxoethyl)piperidine -
69 2-({1-Oxo-[2-{2'-phenyl}ethyl]-4-phenyl}-butyl-l-
(3,3-dimethyl-l,2-dioxobutyl)piperidine
70 (2S)-2-[5,5-di(4-Fluorophenyl)pentanoyl]-1-(3,3
dimethyl-1,2-pentanedione)pyrrolidine
71 3,3-Dimethyl-l-[2-(5-phenylpentanoyl)piperidino]-
1,2-pentanedione
FORMULA III
Another preferred embodiment is a compound of
formula III:
B-C
A Z Ri
=N
0 III
2
or a pharmaceutically acceptable salt thereof, wherein:
CA 02602791 2007-09-19
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A, B, C and D are independently CH2, 0, S, SO, SO2, NH or NR2;
X is 0 or S;
Z is S, CH2, CHRl or C(Rl)Z;
Rl is C1-C6 straight or branched chain alkyl or alkenyl, which is substi-
tuted in one or more position(s) with (Arl)n, (Arl)õ connected by a C1-C6
straight
or branched chain alkyl or alkenyl, C3-C8 cycloalkyl, C3-C8 cycloalkyl
connected
by a C1-C6 straight or branched chain alkyl or alkenyl, Ar2 or a combination
thereof;
n is 1 or 2;
R2 is either C1-C9 straight or branched chain alkyl or alkenyl, C3-C9 cyclo-
alkyl, C5-C7 cycloalkenyl or Ar1, wherein said alkyl, alkenyl, cycloalkyl or
cyclo-
alkenyl is either unsubstituted or substituted in one or more position(s) with
C1-
C4 straight or branched chain alkyl or alkenyl, hydroxyl or a combination
thereof; and
Ari and Ar2 are independently a mono-, bi- or tricyclic, carbo- or hetero-
cyclic ring, wherein the ring is either unsubstituted or substituted in one to
three
position(s) with halo, hydroxyl, nitro, trifluoromethyl, C1-C5 straight or
branched
chain alkyl or alkenyl, C1-C4 alkoxy, C1-C4 alkenyloxy, phenoxy, benzyloxy,
amino or a combination thereof; wherein the individual ring sizes are 5-6
members; and wherein the heterocyclic ring contains 1-6 heteroatom(s)
selected from the group consisting of 0, N, S and a combination thereof.
Particularly preferred compounds of formula III are presented in
TABLE II.
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TABLE II
No. A B C X Z Rl R2
72 CH2 S CH2 0 S 2-phenethyl 3, 3-dimethyl -
pentyl
73 CH: S CH2 0 CH2 3-phenylpropyl 3, 3-dimethyl-
pentyl
74 C-1-:_ Ch, N'ri 0 S 2-phenethyl 3, 3-dimethyl -
pentyl
75 CH, S CH2 S S 2-phenethyl 3, 3-dimethyl-
pentyl
FORMULA IV
A further preferred embodiment of this invention is a compound of
formula IV:
BCD
A Z Rl
O IV
O x
R2
CA 02602791 2007-09-19
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or a pharmaceutically acceptable salt thereof, wherein;
A, B, C and D are independently CH2, 0, S, SO, SOZ, NH or NR2;
XisOorS;
Z is S, CH2, CHR1 or C(R1)2;
Rl is C1-C5 straight or branched chain alkyl or alkenyl, which is substi-
tuted in one or more position(s) with (Arl)n, (Ari)n connected by a C1-C6
straight
or branched chain alkyl or alkenyl, C3-C8 cycloalkyl, C3-C8 cycloalkyl
connected
by a C1-C6 straight or branched chain alkyl or alkenyl, Ar2 or a combination
thereof;
n is 1 or 2;
R2 is either C1-C9i straight or branched chain alkyl or alkenyl, C3-C9 cyclo-
alkyl, C5-C7 cycloalkenyl or Arl, wherein said alkyl, alkenyl, cycloalkyl or
cyclo-
alkenyl is either unsubstituted or substituted in one or more position(s) with
C1-C4, straight or branched chain alkyl or alkenyl, hydroxyl or a combination
thereof; and
Arl and Ar2 are independently a mono-, bi- or tricyclic, carbo- or hetero-
cyclic ring, wherein the ring is either unsubstituted or substituted in one to
three
position(s) with halo, hydroxyl, nitro, trifluoromethyl, C1-C5 straight or
branched
chain alkyl or alkenyl, C1-C4 alkoxy, C1-C4 alkenyloxy, phenoxy, benzyloxy,
amino or a combination thereof; wherein the individual ring sizes are 5-6
members; and wherein the heterocyclic ring contains 1-6 heteroatom(s)
selected from the group consisting of 0, N, S and a combination thereof.
Particularly preferred compounds of formula IV are presented in
TABLE III.
CA 02602791 2007-09-19
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TABLE III
No. A B C D X Z R, R2
76 CH, CH1 0 CH1 0 CH1 3-phenylpropyl 3,3-dimethylpentyl
77 CH, CH, 0 CH, 0 S 2-phenethyl 3,3-dimethylpentyl
78 CH, CH, S CH1 0 CH, 3-phenylpropyl 3,3-dimethylpenryl
79 CH, CH, S CH, 0 S 2-phenethyl 3,3-dimethylpentyl
The compounds of this invention possess asymmetric centers and thus
can be produced as mixtures of stereoisomers or as individual stereoisomers.
The individual stereoisomers may be obtained by using an optically active
start-
ing material, by resolving a racemic or non-racemic mixture of an intermediate
at some appropriate stage of the synthesis, or by resolving the compound of
formula (I). It is understood that the individual stereoisomers as well as
mixtures (racemic and non-racemic) of stereoisomers are encompassed by the
scope of the present invention. The compounds of this invention possess at
least
one asymmetric center and thus can be produced as mixtures of stereoisomers
or as individual R- and S-stereoisomers. The individual enantiomers may be
obtained by resolving a racemic or non-racemic mixture of an intermediate at
some appropriate stage of the synthesis. It is understood that the individual
R-
and S-stereoisomers as well as mixtures of stereoisomers and encompassed by
this invention. The S-stereoisomer is most preferred due to its greater
activity.
FORMULAE IA, IIA, IIIA and IVA
According to an aspect, the invention relates to compounds of formulae
IA, IIA, IIIA and IVA outlined below. The invention also relates to pharma-
ceutical compositions comprising these compounds. Moreover, the invention
relates to administering to an animal a neurotrophically effective amount of a
compound of formulae IA, IIA, IIIA or IVA.
CA 02602791 2007-09-19
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B
rA,,, N
N
O 0
O
R2
IA
(CH2)n
N
N
O 0
O
R2 IIA
B--C
~ I Y--~~ N N
O 0
O
R2
IIIA
CA 02602791 2007-09-19
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B/ C D
I I
AN N
O O
O
R2 IVA
Method of Using the Compounds of the Invention
The compounds of the present invention have an affinity for the FK506
binding protein, particularly FKBP12, which is present in the brain. When the
inventive compounds bind to FKBP in the brain, they exhibit excellent neuro-
trophic activity. This activity is useful in the stimulation of damaged
neurons,
the promotion of neuronal regeneration, the prevention of neurodegeneration,
and the treatment of several neurological disorders known to be associated
with neuronal degeneration and peripheral neuropathies.
For the foregoing reasons, the present invention further relates to a
method of effecting a neuronal activity in an animal, comprising:
administering to the animal a neurotrophically effective amount of a
compound of formulae I, II, III or IV.
In a preferred embodiment, the neuronal activity is selected from the
group consisting of stimulation of damaged neurons, promotion of neuronal
regeneration, prevention of neurodegeneration and treatment of neurological
disorder.
The neurological disorders that may be treated include but are not
limited to: trigeminal neuralgia; glossopharyngeal neuralgia; Bell's Palsy;
CA 02602791 2007-09-19
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myasthenia gravis; muscular dystrophy; amyotrophic lateral sclerosis;
progressive muscular atrophy; progressive bulbar inherited muscular atrophy;
herniated, ruptured or prolapsed invertebrate disk syndromes, cervical
spondylosis; plexus disorders; thoracic outlet destruction syndromes;
peripheral neuropathies such as those caused by lead, dapsone, ticks,
porphyria or Guillain-Barre syndrome; Alzheimer's disease; and Parkinson's
disease.
The compounds of the present invention are particularly useful for
treating a neurological disorder selected from the group consisting of:
peripheral neuropathy caused by physical injury or disease state, traumatic
injury to the brain, physical damage to the spinal cord, stroke associated
with
brain damage, and neurological disorder relating to neurodegeneration.
Examples of neurological disorders relating to neurodegeneration are
Alzheimer's Disease, Parkinson's Disease and amyotrophic lateral sclerosis.
For these purposes, the compounds may be administered orally,
parenterally, by inhalation spray, topically, rectally, nasally, buccally,
vaginally or via an implanted reservoir in dosage formulations containing
conventional non-toxic pharmaceutically-acceptable carriers, adjuvants and
vehicles. The term parenteral as used herein includes subcutaneous, intra-
venous, intramuscular, intraperitoneally, intrathecally, intraventricularly,
intrasternal and intracranial injection or infusion techniques.
To be effective therapeutically as central nervous system targets, the
compounds should readily penetrate the blood-brain barrier when peripherally
administered. Compounds which cannot penetrate the blood-brain barrier can
be effectively administered by an intraventricular route.
The compounds may be administered in the form of sterile injectable
preparations, for example, as sterile injectable aqueous or oleaginous sus-
pensions. These suspensions may be formulated according to techniques
known the art using suitable dispersing or wetting agents and suspending
agents. The sterile injectable preparations may also be sterile injectable
CA 02602791 2007-09-19
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solutions or suspensions in non-toxic parenterally-acceptable diluents or
solvents, for example, as solutions in 1, 3-butanediol. Among the acceptable
vehicles and solvents that may be employed are water, Ringer's solution and
isotonic sodium chloride solution. In addition, sterile, fixed oils are conven-
tionally employed as solvents or suspending mediums. For this purpose, any
bland fixed oil such as a synthetic mono- or di-glyceride may be employed
Fatty acids such as oleic acid and its glyceride derivatives, including olive
oil
and castor oil, especially in their polyoxyethylated versions, are useful in
the
preparation of injectables. These oil solutions or suspensions may also
contain
long-chain alcohol diluents or dispersants.
Additionally, the compounds may be administered orally in the form of
capsules, tablets, aqueous suspensions or solutions. Tablets may contain
carriers such as lactose and corn starch, and/or lubricating agents such as
magnesium stearate. Capsules may contain diluents including lactose and
dried corn starch. Aqueous suspensions may contain emulsifying and sus-
pending agents combined with the active ingredient. The oral dosage forms
may further contain sweetening and/or flavoring and/or coloring agents.
The compounds may also be administered rectally the form of supposi-
tories. These compositions can be prepared by mixing the drug with a suitable
non-irritating excipient which is solid at room temperature, but liquid at
rectal
temperature and, therefore, will melt in the rectum to release the drug. Such
materials include cocoa butter, beeswax and polyethylene glycols.
Furthermore, the compounds may be administered topically, especially
when the conditions addressed for treatment involve areas or organs readily
accessible by topical application, including neurological disorders of the
eye,
the skin or the lower intestinal tract. Suitable topical formulations can be
readily prepared for each of these areas.
For topical application to the eye, or ophthalmic use, the compounds
can be formulated as micronized suspensions in isotonic, pH adjusted sterile
saline, or, preferably, as a solution in isotonic, pH adjusted sterile saline,
CA 02602791 2007-09-19
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either with or without a preservative such as benzylalkonium chloride.
Alternatively, the compounds may be formulated into ointments, such as
petrolatum, for ophthalmic use.
For topical application to the skin, the compounds can be formulated
into suitable ointments containing the compounds suspended or dissolved in,
for example, mixtures with one or more of the following: mineral oil, liquid
petrolatum, white petrolatum, propylene glycol, polyoxyethylene polyoxypro-
pylene compound, emulsifying wax and water. Alternatively, the compounds
can be formulated into suitable lotions or creams containing the active com-
pound suspended or dissolved in, for example, a mixture of one or more of
the following: mineral oil, sorbitan monostearate, polysorbate 60, cetyl ester
wax, cetearyl alcohol, 2-octyidodecanol, benzyl alcohol and water.
Topical application to the lower intestinal tract can be effected in rectal
suppository formulations (see above) or in suitable enema formulations.
Dosage levels on the order of about 0.1 mg to about 10,000 mg of the
active ingredient compound are useful in the treatment of the above condi-
tions, with preferred levels of about 0.1 mg to about 1,000 mg. The amount
of active ingredient that may be combined with the carrier materials to
produce a single dosage form will vary depending upon the host treated and
the particular mode of administration.
It is understood, however, that a specific dose level for any particular
patient will depend upon a variety of factors, including the activity of the
specific compound employed; the age, body weight, general health, sex and
diet of the patient; the time of administration; the rate of excretion; drug
combination; the severity of the particular disease being treated; and the
form of administration.
The compounds can be administered with other neurotrophic agents
such as neurotrophic growth factor (NGF), glial derived growth factor, brain
derived growth factor, ciliary neurotrophic factor and neurotropin-3. The
CA 02602791 2007-09-19
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dosage level of other neurotrophic drugs will depend upon the factors pre-
viously stated and the neurotrophic effectiveness of the drug combination.
Pharmaceutical Compositions of the Invention
The present invention also relates to a pharmaceutical composition
comprising:
(i) a neurotrophically effective amount of the compound of formula
I, II, III or IV, and
(ii) a pharmaceutically acceptable carrier. The above discussion
relating to the utility and administration of the compounds of the present
invention also applies to the pharmaceutical compositions of the present
invention.
Examnles
The following examples are illustrative of the present invention and are
not intended to be limitations thereon. Unless otherwise specified, all
percentages are based on 100% by weight of the final compound.
CA 02602791 2007-09-19
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EXAMPLE 1
Synthesis of (2S) -2- (1-oxo-5-phenyl) -pentyl-l- (3 , 3-
dimethyl-l,2-dioxopentyl)pyrrolidine (1)
(2S)-2-(1-oxo-5-phenyl)pentyl-N-benzylpyrrolidine.
1-chloro-4-phenylbutane (1.78 g; 10.5 mmol) in 20 mL of
THF was added to 0.24 g(10 mmol) of magnesium turnings
in 50 mL of refluxing THF. After the addition was
complete, the mixture was refluxed for' an additional 5
hours, and then added slowly to a refluxing solution of
N-benzyl-L-proline ethyl ester (2.30 g (10 mmol) in 100
mL of THF. After 2 hours of further reflux, the mixture
was cooled and treated with 5 mL of 2 N HCl. The
reaction mixture was diluted with ether (100 mL) and
washed with saturated NaHCO3, water and brine. The
organic phase was dried, concentrated and
chromatographed, eluting with 5:1 CH2C12:EtOAc to obtain
2.05 g (640) of the ketone as an oil, 1H NMR (CDC13; 300
MHz): 1.49-2.18 (m, 8H) ; 2.32-2.46 (m, 1H) ; 2.56-2.65
(m, 2H); 2.97-3.06 (m, 1H); 3.17-3.34 (m, 1H); 3.44-3.62
(m, 1H); 4.02-4.23 (m, 2H); 7.01-7.44 (m, 10H).
(2S) -2- (1-oxo-5-phenyl ) pentylpyrrolidine. The ketone
compound (500 mg) and palladium hydroxide (20% on carbon,
50 mg) was hydrogenated at 40 psi in a Paar shaker
overnight. The catalyst was removed by filtration and
the solvent was removed in vacuo. The free amine was
obtained as a yellow oil (230 mg; 100 0), 1H NMR (CDC13;
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300 MHz): 1.75-2.34 (m, 10H) ; 2.55 (m, 2H) ; 2.95 (dm,
1H); 3.45-3.95 (m, 1H); 4.05 (m, 1H); 7.37 (m, 5H).
(2S)-2-(1-oxo-5-phenyl)pentyl-l-(1,2-dioxo-2-
methoxyethyl)pyrrolidine. To a solution of (2S)-2-(1-
oxo-5-phenyl)pentylpyrrolidine (230 mg; 1.0 mmol) in
CH2C12(20 mL) at 0 C was added dropwise methyloxalyl
chloride (135 mg; 1.1 mmol). After stirring at 0 C for
3 hours, the reaction was quenched with saturated NH4Cl
and the organic phase was washed with water and brine and
dried and concentrated. The crude residue was purified
on a silica gel column, eluting with 20:1 CH2C12:EtOAc to
obtain 300 mg of the oxamate as a clear oil (98%), 'H NMR
(CDC13; 300 MHz): 1.68 (m, 4H); 1.91-2.38 (m, 4H); 2.64
(t, 2H); 3.66-3.80 (m, 2H); 3.77, 3.85 (s, 3H total);
4.16 (m, 2H); 4.90 (m, 1H); 7.16 (m, 3H); 7.27 (m, 2H).
(2S)-2-({1-oxo-5-phenyl}-pentyl-1-(3,3-dimethyl-1,2-
dioxopentyl)pyrrolidine (1). To a solution of the
oxamate above (250 mg; 0.79 mmol) in anhydrous ether (15
mL), cooled to - 78 C, was added 1,1-dimethylpropyl-
magnesium chloride (0.8 mL of a 1.0 M solution in ether;
0.8 mmol). After stirring the resulting mixture at -78 C
for 2 hours, the reaction was quenched by the addition of
2 mL of saturated NH4C1, followed by 100 mL of EtOAc.
The organic phase was washed with brine, dried,
concentrated, and purified on a silica gel column,
eluting with 50:1 CH2C12:EtOAc. Compound 1 was obtained
CA 02602791 2007-09-19
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as a clear oil, 120 mg, 'H NMR (CDC13 , 300 MHz) : b 0.87
(t, 3H, J=7.5); 1.22 (s, 3H); 1.25 (s, 3H); 1.67 (m, 4H);
1.70-2.33 (m, 6H); 2.61 (t, 2H, J=7.1); 3.52 (m, 2H); 4.17
(t, 2H, J=6.2); 4.52 (m, 1H); 7.16-7.49 (m, SH). Anal.
Calcd. for CZZH31N03 - H20: C, 70.37; H, 8.86; N, 3.73.
Found: 70.48; H, 8.35; N, 3.69.
EXAMPLE 2
Synthesis of 2-phenyl-l-ethyl 1- (3 3-dimethyl-1 2-
dioxopentyl)-2-pyrrolidinecarbothioate (9)
Methyl(2S)-1-(1,2-dioxo-2-methoxyethyl)-2-
pyrrolidinecarboxylate. A solution of L-proline methyl
ester hydrochloride (3.0'8 g; 18.60 mmol) in dry methylene
chloride was cooled to 0 C and treated with triethylamine
(3.92 g; 38.74 mmol; 2.1 eq). After stirring the formed
slurry under a nitrogen atmosphere for 15 min, a solution
of methyl oxalyl chloride (3.20 g; 26.12 mmol) in
methylene chloride (45 mL) was added dropwise. The
resulting mixture was stirred at 0 C for 1.5 hour. After
filtering to remove solids, the organic phase was washed
with water, dried over MgSO4 and concentrated. The crude
residue was purified on a silica gel column; eluting with
50% ethyl acetate in hexane, to obtain 3.52 g( 88 0) of
the product as a reddish oil. Mixture of cis-trans amide
rotamers; data for,trans rotamer given. 'H NMR (CDC13
6.1.93 (dm, 2H); 2.17(m, 2H); 3.62(m, 2H); 3.71 (s, 3H);
CA 02602791 2007-09-19
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3.79, 3.84 (s, 3H total); 4.86 (dd, 1H, J=8.4, 3.3).
Methyl (2S)-1-(1,2-dioxo-3,3-dimethylpentyl)-2-pyrrolidinecarbox ly ate.
A solution of methyl (2S)-1-(1,2-dioxo-2-methoxyethyl)-2-pyrrolidinecar-
boxylate (2.35 g; 10.90 mmol) in 30 mL of tetrahydrofuran (THF) was cooled
to -78 C and treated with 14.2 mL of a 1.0 M solution of 1,1-dimethylpropyl-
magnesium chloride in THF. After stirring the resulting homogeneous mixture
at -78 C for three hours, the mixture was poured into saturated ammonium
chloride (100 mL) and extracted into ethyl acetate. The organic phase was
washed with water, dried, and concentrated, and the crude material obtained
upon removal of the solvent was purified on a silica gel column, eluting with
25% ethyl acetate in hexane, to obtain 2.10 g(75%) of the oxamate as a
colorless oil, 'H NMR (CDCI3): b 0.88 (t, 3H); 1.22, 1.26 (s, 3H each); 1.75
(dm, 2H); 1.87-2.10 (m, 3H); 2.23 (m, 1H); 3.54 (m, 2H); 3.76 (s, 3H); 4.52
(dm, 1H, 3=8.4, 3.4).
(2S)-1-(1,2-dioxo-3,3-dimethylpentyl)-2-pyrrolidine-carboxylic acid. A
mixture of methyl (2S)-1-(1,2-dioxo-3,3-dimethylpentyl)-2-pyrrolidine-
carboxylate (2.10 g; 8.23 mmol), 1 N LiOH (15 mL), and methanol (50 mL)
was stirred at 0 C for 30 minutes and at room temperature overnight. The
mixture was acidified to pH 1 with 1 N HCI, diluted with water, and extracted
into 100 mL of methylene chloride. The organic extract was washed with
CA 02602791 2007-09-19
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brine and concentrated to deliver 1.73 g(870) of snow-
white solid which did not require further purification,
1H NMR (CDC13) 6 0.87 (t, 3H); 1.22, 1.25 (s, 3H each);
1.77 (dm, 2H); 2.02 (m, 2H); 2.17 (m, 1H); 2.25 (m, 1H);
3.53 (dd, 2H, J=10.4, 7.3); 4.55 (dd, 1H, J=8.6, 4.1).
2-phenyl-l-ethyl 1-(3,3-dimethyl-1,2-dioxopentyl)-2-
Qyrrolidinecarbothioate (9). To a solution of (2S)-1-
(1,2-dioxo-3,3-dimethylpentyl)-2-pyrrolidinecarboxylic
acid (241 mg; 1.0 mmol) in CHZC12 (10 mL) was added
dicyclohexylcarbodiimide (226 mg; 1.1 mmol). After
stirring the resulting mixture for 5 minutes, the
solution was cooled to 0 C and treated with a solution of
phenethyl mercaptan (138 mg; 1.0 mmol) and 4-
dimethylaminopyridine (6 mg) in 5 ml of CH2ClZ. The
mixture was allowed to warm to room temperature with
stirring overnight. The solids were removed by
filtration and the filtrate was concentrated in vacuo;
the crude residue was purified by flash chromatography
(10:1 hexane:EtOAc) to obtain 302 mg (84%) of 9 as an
oil, 'H NMR (CDC13, 300 MHz): 6 0.85 (t, 3H, J=7.5); 1.29
(s, 3H); 1.31 (s, 3H); 1.70-2.32 (m, 6H); 2.92 (t, 2H,
J=7.4); 3.22(t, 2H, J=7.4); 3.58 (m, 214); 4.72 (m, 1H);
7.23-7.34 (m, 5H). Anal. Calcd. for C20H27N03S - 0.4H20:
C, 65.15; H, 7.60; N, 3.80. Found: C, 65.41; H, 7.49;
N, 3.72.
CA 02602791 2007-09-19
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EXAMPLE 3
Synthesis of 2-phenyl-l-ethyl (2S)-1-(3,3-
dimethyl-1,2-dioxopentyl)-2_piperidinecarbothioate (10)
Methyl 1-(1,2-dioxo-2-methoxyethyl)-2-piperidine-
carboxylate. A solution of methyl pipecolate
hydrochloride (8.50 g; 47.31 mmol) in dry methylene
chloride (100 mL) was cooled to 0 C and treated with
triethylamine (10.5 g; 103 mmol; 2.1 eq) After stirring
the formed slurry under a nitrogen atmosphere for 15
minutes, a solution of methyl oxalyl chloride (8.50 g;
69.4 mmol) in methylene chloride (75 mL) was added
dropwise. The resulting mixture was stirred at 0 C for
1.5 hours. After filtering to remove solids, the organic
phase was washed with water, dried over MgSOq and
concentrated. The crude residue was purified on a silica
gel column, eluting with 50% ethyl acetate in hexane, to
obtain 9.34 g(860) -of the product as a reddish oil.
Mixture of cis-trans amide rotamers; data for trans
rotamer given. iH NMR (CDC13): b 1.22-1.45 (m, 2H);
1.67-1.78 (m, 3H); 2.29 (m, 1H); 3.33 (m, 1H); 3.55 (m,
1H); 3.76 (s, 3H); 3.85, 3.87 (s, 3H total); 4.52 (dd,
1H).
Methyl 1-(1,2-dioxo-3,3-dimethylpentyl)-2-piperidine
-carboxylate. A solution of methyl 1-(1,2-dioxo-2-
methoxyethyl)-2-piperidinecarboxylate (3.80 g; 16.57
mmol) in 75 mL of tetrahydrofuran (THF) was cooled to -
CA 02602791 2007-09-19
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78 C and treated with 20.7 mL of a 1.0 M solution of 1,1-dimethyl-propyl-
magnesium chloride in THF. After stirring the resulting homogeneous mixture
at -78 C for three hours, the mixture was poured into saturated ammonium
chloride (100 mL) and extracted into ethyl acetate. The organic phase was
washed with water, dried, and concentrated, and the crude material obtained
upon removal of the solvent was purified on a silica gel column, eluting with
25% ethyl acetate in hexane, to obtain 3.32 g (74%) of the oxamate as a
colorless oil, 'R NMR (CDC13): 6 0.88 (t, 3H); 1.21, 1.25 (s, 3H each); 1.35-
1.80 (m, 7H); 2.35 (m, 1H); 3.24 (m, 1H); 3.41 (m, 1H); 3.76 (s, 3H); 5.32
(d, 1H).
1-(1,2-dioxo-3,3-dimethylpentyl)-2-piperidine-carboxylic acid. A
mixture of methyl 1-(1,2-dioxo-3,3-dimethylpentyl)-2-piperidinecarboxylate
(3.30 g; 12.25 mmol), 1 N LiOH (15 mL), and methanol (60 mL) was stirred
at 0 C for 30 minutes and at room temperature overnight. The mixture was
acidified to pH 1 with 1 N HCI, diluted with water, and extracted into 100 mL
of methylene chloride. The organic extract was washed with brine and con-
centrated to deliver 2.80 g (87%) of snow-white solid which did not require
further purification, 'H NMR (CDC13): 6 0.89 (t, 3H); 1.21, 1.24 (s, 3H each);
1.42- 1.85 (m, 7H); 2.35 (m, 1H); 3.22 (d, 1H); 3.42 (m, 1H); 5.31 (d, 1H).
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2-phenyl-l-ethyl (2S)-1-(3,3-dimethyl-1,2-
dioxopentyl)-2-piperidinecarbothioate (10).. To a
solution of 1-(1,2-dioxo-3,3-dimethylpentyl)-2-
piperidine-carboxylic acid (255 mg; 1.0 mmol) in CHZC12
(10 mL) was added dicyclohexylcarbodiimide (226 mg; 1.1
mmol). After stirring the resulting mixture for 5
minutes, the solution was cooled to 0 C and treated with
a solution of phenethyl mercaptan (138 mg; 1.0 mmol) and
4-dimethylaminopyridine (6 mg) in 5 ml of CH2Clz. The
mixture was allowed to warm to room temperature with
stirring overnight. The solids were removed by
filtration and the filtrate was concentrated in vacuo;
the crude :residue was purified by flash chromatography
(10:1 hexane:EtOAc) to obtain 300 mg (80%) of 10 as an
oil, 'H NMR (CDC13, 300 MHz): d 0.94 (t, 3H, J=7.5); 1.27
(s; 3H) ; 1.30 (s, 3H) ; 1 .34-1 .88 (m, 7H) ; 2.45 (m, 1H) ;
2.90 (t, 2H, J=7.7); 3:26-(t, 2-H; J=7.7); 3.27- (m, 1H) ;
3.38 (m, 1H); 5.34 (m, 1H); 7.24-7.36 (m, 5-H). Anal.
Calcd. for C21H29N03S: C, 67.17; H, 7.78; N, 3.73. Found:
C, 67.02; H, 7.83; N, 3.78.
As discussed above, the compounds of the present
invention have an affinity for the FK506 binding protein,
particularly FKBP12. The inhibition of the prolyl
peptidyl cis-trans isornerase activity of FKBP may be
measured as an indicator of this affinity.
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K,. Test Procedure
Inhibition of the peptidyi-prolyl isomerase (rotamase) activity of the
inventive compounds can be evaluated by known methods described in the
literature (Harding et al., Nature, 1989, 341:758-760; Holt et al. J. Am.
Chem. Soc., 115:9923-9938). These values are obtained as apparent Kl's and
are presented for representative compounds in Table IV. The cis-trans iso-
merization of an alanine-proline bond in a model substrate, N-succinyl-Ala-
Ala-Pro-Phe-p-nitroanilide, is monitored spectrophotometrically in a chymo-
trypsin-coupled assay, which releases para-nitroanilide from the trans form of
the substrate. The inhibition of this reaction caused by the addition of
differ-
ent concentrations of inhibitor is determined, and the data is analyzed as a
change in first-order rate constant as a function of inhibitor concentration
to
yield the apparent Kl values.
In a plastic cuvette are added 950 mL of ice cold assay buffer (25 mM
HEPES, pH 7.8, 100 mM NaCI), 10 mL of FKBP (2.5 mM 10 mM Tris-CI pH 7.5,
100 mM NaCI, 1 mM dithiothreitol), 25 mL of chymotrypsin (50 mg/ml in
1 mM HCI) and 10 L of test compound at various concentrations in dimethyl
sulfoxide. The reaction is initiated by the addition of 5 mL of substrate
(succinyl-Ala-Phe-Pro-Phe-para-nitroanilide, 5 mg/mL 2.35 mM LiCI trifluoro-
ethanol).
The absorbance at 390 nm versus time is monitored for 90 seconds
using a spectrophotometer and the rate constants are determined from the
absorbance versus time data files.
The data for these experiments for representative compounds are
presented in Table IV under the column "Ki".
The neurotrophic effects of the compounds of the present invention can
be demonstrated in cellular biological experiments in vitro, as described
below.
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Chick Dorsal Root Ganglion Cultures and Neurite Outgrowth
The neurotrophic effects of the FKBP inhibitor compounds were demon-
strated by evaluating the ability of the compounds to promote neurite out-
growth in cultured chick sensory neurons from dorsal root ganglia. Dorsal root
ganglia were dissected from chick embryos of ten day gestation. Whole
ganglion explants were cultured on thin layer Matrigel-coated 12 well plates
with Liebovitz L15 plus high glucose media supplemented with 2 mM gluta-
mine and 10% fetal calf serum, and also containing 10 pM cytosine (3-D
arabinofuranoside (Ara C) at 37 C in an environment containing 5% COZ.
Twenty-four hours later, the DRGs were treated with various concentrations of
nerve growth factor, immunophilin ligands or combinations of NFG plus drugs.
Forty-eight hours after drug treatment, the ganglia were visualized under
phase contrast or Hoffman Modulation contrast with a Zeiss Axiovert inverted
microscope. Photomicrographs of the explants were made, and neurite out-
growth was quantitated. Neurites longer than the DRG diameter were
counted as positive, with total number of neurites quantitated per each
experimental condition. Three to four DRGs are cultured per well, and each
treatment was performed duplicate.
Dose-response curves were generated from which ED50 values were
obtained. The results of these experiments are presented in Table IV under
the column "ED50". Representative photomicrographs of untreated (control)
sensory neurons and of compounds 1 (10 pM, 1 nM, 1 pM) , 9 (10 pM, 1 nM,
100 nM) and 10 (10 pM, 1 nM, 100 nM) promoting neurite outgrowth in
sensory neurons are shown m FIG.'s 1(A-D), 2(A-D) and 3(A-D), respectively.
MPTP Model of Parkinson's Disease
The remarkable neurotrophic and neuroregenerative effects of the
present inventive compounds were further demonstrated in an animal model
of neurodegenerative disease. MPTP lesioning of dopaminergic neurons in
mice was used as an animal model of Parkinson's Disease. Four week old
male CD1 white mice were dosed i.p. with 30 mg/kg of MPTP for 5 days. Test
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compounds (4 mg/kg), or vehicle, were administered s.c. along with the MPTP
for 5 days, as well as for an additional 5 days following cessation of MPTP
treatment. At 18 days following MPTP treatment, the animals were sacrificed
and the striata were dissected and homogenized. Immunostaining was per-
formed on saggital and coronal brain sections using anti-tyrosine hydroxylase
1 g to quantitate survival and recovery of dopaminergic neurons. In animals
treated with MPTP and vehicle, a substantial loss of functional dopaminergic
terminals was observed as compared to non-lesioned animals. Lesioned
animals receiving test compounds showed a significant recovery of TH-stained
dopaminergic neurons.
The results of these experiments are presented in TABLE IV under the
column "% TH recovery". Quantitation for the recovery of TH-positive dopa-
minergic neurons in the striatum of animals receiving compounds 1, 9 and 10,
and for representative control and lesioned animals not receiving the test
drugs, are presented in FIG. 4.
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TABLE IV
In Vitro Test Results
Example # Ki. nM ED50, nM % TH recovery
1 31 0.4 23
2 210 - --
3 85 - --
9 104 0.5 61
12 0.8 54
11 299 0.36 53
12 442 0.025 --
14 313 0.9 48
28 108 0.9 41
29 59 0.003 50
30 11 0.00025 65
- 1 8.7 -- 31
32 362 -- 52
33 1698 -- --
34 34 0.9 48
35 62 -- --
36 7 -- 56
37 68 -- -
38 8.9 0.011 37.32
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39 347 -- -
40 1226
41 366 - -
42 28 -- --
43 259 -- -
44 188 - 25
45 31 -- --
46 757 -- --
47 21 -- 50
48 127 -- 28
49 1334 -- --
50 55 - 62
51 33 -- -
52 6 -- --
53 261 -- --
54 37
-- --
55 30 - -
56 880 - -
57 57 -- --
58 79 - --
59 962 -- -
60 90 -- --
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61 139 - -
62 196 - -
63 82 - -
64 163 - -
65 68 - -
66 306 5 38
67 177 - --
68 284 -- --
69 49 -- 23
70 457 -- 25
71 788 -- --
The invention being thus described, it will be obvious that the same
may be varied in many ways. Such variations are not to be regarded as a
departure from the spirit and scope of the invention and all such
modifications
are intended to be included within the scope of the following claims.
