U.S. patent application number 17/428505 was filed with the patent office on 2022-01-20 for methods for treating symptoms and disorders associated with lysosomal storage diseases.
This patent application is currently assigned to GENZYME CORPORATION. The applicant listed for this patent is GENZYME CORPORATION. Invention is credited to Nigel Patrick Somerville CRAWFORD, Tanya Zaremba FISCHER.
Application Number | 20220016092 17/428505 |
Document ID | / |
Family ID | 1000005931749 |
Filed Date | 2022-01-20 |
United States Patent
Application |
20220016092 |
Kind Code |
A1 |
CRAWFORD; Nigel Patrick Somerville
; et al. |
January 20, 2022 |
METHODS FOR TREATING SYMPTOMS AND DISORDERS ASSOCIATED WITH
LYSOSOMAL STORAGE DISEASES
Abstract
This disclosure to methods for treating or preventing particular
symptoms and disorders which are associated with lysosomal storage
diseases using quinuclidine compounds of formula (I), optionally in
combination with enzyme replacement therapy. This includes
supranuclear gaze palsies, including horizontal and vertical
saccadic gaze palsies, and cognitive deficits or gait disorders,
such as in a patient having Gaucher disease or Niemann-Pick disease
Type C. Also disclosed is a pharmaceutical composition comprising a
quinuclidine compound for use in said methods.
Inventors: |
CRAWFORD; Nigel Patrick
Somerville; (Bridgewater, NJ) ; FISCHER; Tanya
Zaremba; (Bridgewater, NJ) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
GENZYME CORPORATION |
Cambridge |
MA |
US |
|
|
Assignee: |
GENZYME CORPORATION
Cambridge
MA
|
Family ID: |
1000005931749 |
Appl. No.: |
17/428505 |
Filed: |
February 3, 2020 |
PCT Filed: |
February 3, 2020 |
PCT NO: |
PCT/US2020/016441 |
371 Date: |
August 4, 2021 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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62962647 |
Jan 17, 2020 |
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62937618 |
Nov 19, 2019 |
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62894167 |
Aug 30, 2019 |
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62851433 |
May 22, 2019 |
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62800996 |
Feb 4, 2019 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K 38/47 20130101;
A61P 25/02 20180101; A61K 31/439 20130101 |
International
Class: |
A61K 31/439 20060101
A61K031/439; A61P 25/02 20060101 A61P025/02; A61K 38/47 20060101
A61K038/47 |
Claims
1. A method for treating or preventing supranuclear gaze palsies
(e.g., associated with a lysosome storage disease), in a subject in
need thereof, the method comprising administering to the subject an
effective amount of a compound of formula (I), ##STR00015## or a
pharmaceutically acceptable salt or prodrug thereof, wherein:
R.sup.1 is selected from hydrogen, halogen (e.g., fluorine), cyano,
nitro, hydroxy, thio, amino, C.sub.1-6-alkyl (e.g., methyl or
ethyl), C.sub.2-6-alkenyl, C.sub.2-6-alkynyl, C.sub.1-6-alkyloxy,
C.sub.2-6-alkenyloxy, and C.sub.2-6-alkynyloxy, wherein said alkyl,
alkenyl, alkynyl, alkyloxy, alkenyloxy, or alkynyloxy is optionally
substituted with one or more (e.g., 1, 2 or 3) groups selected from
halogen, cyano, nitro, hydroxy, thio or amino; R.sup.2 and R.sup.3
are independently selected from C.sub.1-3-alkyl, optionally
substituted by one or more (e.g. 1, 2 or 3) halogens, or R.sup.2
and R.sup.3 together form a cyclopropyl or cyclobutyl group,
optionally substituted by one or more (e.g. 1 or 2) halogens;
R.sup.4, R.sup.5 and R.sup.6 are each independently selected from
hydrogen, halogen, nitro, hydroxy, thio, amino, C.sub.1-6-alkyl,
and C.sub.1-6-alkyloxy, wherein said alkyl or alkyloxy is
optionally substituted by one or more (e.g. 1, 2 or 3) groups
selected from halogen, hydroxy, cyano, and C.sub.1-6-alkyloxy; and
A is a 5- or 6-membered aryl or heteroaryl group (e.g., phenyl or
thiazolyl), optionally substituted with 1, 2 or 3 groups
independently selected from halogen, hydroxy, thio, amino, nitro,
C.sub.1-6alkoxy and C.sub.1-6alkyl.
2. The method of claim 1, wherein R.sup.1 is selected from
hydrogen, fluorine, methyl and ethyl, wherein said methyl or ethyl
is optionally substituted by 1 or 2 groups selected from halogen,
hydroxy, thio or amino.
3. The method of claim 1 or 2, wherein R.sup.2 and R.sup.3 are each
independently selected from methyl and ethyl groups, optionally
substituted with one or more fluorines.
4. The method of any one of claims 1 to 3, wherein R.sup.4 is
selected from a halogen (e.g., fluorine), C.sub.1-3-alkyl (e.g.,
methyl) and C.sub.1-3-alkyloxy (e.g., methoxy or ethoxy), wherein
said alkyl or alkyoxy is optionally substituted by one or more
(e.g., 1, 2 or 3) groups selected from a halogen and
C.sub.1-3-alkyloxy (e.g., methoxy or ethoxy).
5. The method of any one of claims 1 to 4, wherein R.sup.5 and
R.sup.6 are each hydrogen.
6. The method of any one of claims 1 to 5, wherein R.sup.4 is
fluorine or 2-methoxyethoxy, and R.sup.5 and R.sup.6 are
hydrogen.
7. The method of any one of claims 1 to 6, wherein R.sup.4 is
positioned at the 4-position of the phenyl ring to which it is
attached (i.e., para to the A substituent).
8. The method of any one of claims 1 to 7, wherein A is phenyl,
optionally substituted with 1, 2 or 3 groups independently selected
from halogen, hydroxy, thio, amino, nitro, C.sub.1-6alkoxy and
C.sub.1-6alkyl (e.g., methyl).
9. The method of claim 8, wherein the two groups attached to the A
substituent are positioned in a 1,3- or a 1,4-relationship to each
other (i.e., meta or para).
10. The method of any one of claims 1 to 7, wherein A is a
5-membered heteroaryl group which contains 1 or 2 heteroatoms
selected from N and S.
11. The method of claim 10, wherein the two groups attached to the
A substituent are positioned in a 1,3-relationship to each other
(i.e., meta).
12. The method of any one of claims 1 to 11, wherein said compound
is a compound of formula (II), (III) or (IV), ##STR00016## or a
pharmaceutically acceptable salt or prodrug thereof.
13. The method of any of claims 1 to 11, wherein said compound is a
compound of formula (V), ##STR00017## or a pharmaceutically
acceptable salt or prodrug thereof.
14. The method of any one of claims 1 to 11, wherein said compound
is a compound of formula (VI), (VII) or (VIII), ##STR00018## or a
pharmaceutically acceptable salt or prodrug thereof.
15. The method of any of claims 1 to 11, wherein said compound is a
compound of formula (IX) or (XI), ##STR00019## or a
pharmaceutically acceptable salt or prodrug thereof.
16. The method of claim 15, wherein R.sup.4 is fluorine.
17. The method of claim 1, wherein said compound is selected from:
quinuclidin-3-yl
(2-(4'-fluoro-[1,1'-biphenyl]-3-yl)propan-2-yl)carbamate;
(S)-quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate;
(S)-quinuclidin-3-yl
(2-(4'-(2-methoxyethoxy)-[1,1'-biphenyl]-4-yl)propan-2-yl)carbamate;
and the pharmaceutically acceptable salts and prodrugs thereof.
18. The method of claim 1, wherein said compound is
quinuclidin-3-yl
(2-(4'-fluoro-[1,1'-biphenyl]-3-yl)propan-2-yl)carbamate.
19. The method of claim 1, wherein said compound is
(S)-quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate.
20. The method of any one of claims 1 to 19, wherein the subject
has Gaucher disease Type 3 or Niemann-Pick disease Type C.
21. The method of any one of claims 1 to 20, wherein the
supranuclear gaze palsy is a conjugate gaze palsy
22. The method of any one of claims 1 to 21, wherein said subject
is a mammal, e.g. a human.
23. The method of any one of claims 1 to 22, wherein said compound,
or pharmaceutically acceptable salt or prodrug thereof, is
administered by systemic administration, e.g. via a non-parenteral
route.
24. The method of claim 23, wherein said compound, or
pharmaceutically acceptable salt or prodrug thereof, is
administered orally.
25. The method of any one of claim 1-24, wherein the subject
undergoes concurrent treatment with enzyme replacement therapy
(ERT), e.g., using a glucocerebrosidase (e.g., imiglucerase,
velaglucerase, or taliglucerase).
26. The method of any one of claims 1-25 wherein the subject is
administered a daily dose of about 1 mg to about 50 mg of the
compound, e.g., from 5 to 50 mg, or from 10 to 40 mg, or from 10 to
30 mg, or from 10 to 20 mg, or from 20 to 30 mg, or from 30 to 40
mg, or from 40 to 50 mg, or from 5 to 25 mg, or from 20 to 50 mg,
or from 5 to 15 mg, or from 15 to 30 mg, or about 15 mg;
27. A compound, or a pharmaceutically acceptable salt or prodrug
thereof, as defined in any one of claims 1 to 19 for use in a
method of treating or preventing supranuclear gaze palsies (e.g.,
associated with a lysosome storage disease), in a subject in need
thereof.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is an international application which
claims priority to and the benefit of U.S. Provisional Applications
No. 62/800,996, filed on Feb. 4, 2019, No. 62/851,433, filed on May
22, 2019, No. 62/894,167, filed on Aug. 30, 2019, No. 62/937,618,
filed on Nov. 19, 2019, and No. 62/962,647, filed on Jan. 17, 2020,
the contents of each of which are hereby incorporated in their
entireties.
FIELD
[0002] This invention relates to methods for treating or preventing
particular symptoms and disorders which are associated with
lysosomal storage diseases using quinuclidine compounds of formula
(I), optionally in combination with enzyme replacement therapy.
This includes supranuclear gaze palsies, including horizontal and
vertical saccadic gaze palsies, and cognitive deficits or gait
disorders, such as in a patient having Gaucher disease or
Niemann-Pick disease Type C.
BACKGROUND
[0003] Lysosomal Storage Diseases
[0004] Lysosomal storage diseases (LSDs) are a group of about 50
rare inherited metabolic diseases caused by defects in lysosomal
function. Generally, patients with an LSD accumulate harmful levels
of a substrate (i.e., material stored) in lysosomes due to a
deficiency or defect in an enzyme responsible for metabolizing the
substrate, or due to a deficiency in an enzymatic activator
required for proper enzymatic function. Most LSDs are caused by a
single enzymatic defect or deficiency, usually for an enzyme
involved in the metabolism of lipids or glycoproteins. Some of the
more common LSDs include Gaucher disease, Fabry disease and
Niemann-Pick disease (type C). Gaucher, Fabry and Niemann-Pick are
examples of sphingolipidoses. Each of these diseases are associated
with a constellation of symptoms which are directly or indirectly
caused by the underlying genetic defects. As a result, it is often
difficult to predict which symptoms or disorders associated with
these can be effectively treated with different treatment methods.
Symptoms which are common across several LSDs include alterations
in saccadic eye movements, cognitive dysfunction, and gait
disorders, such as ataxia. These symptoms are particularly common
in Gaucher disease (e.g., type 3) and in Neiman-Pick disease (type
C).
[0005] Saccadic Eye Movement Defects in LSDs
[0006] Several functional classes of eye movements exist, including
saccades, smooth pursuit, optokinetic nystagmus (OKN), vestibular
reflexes, and vergence, each controlled by distinct cortical,
brainstem and cerebellar supranuclear networks. Failure of
brainstem supranuclear saccade centers results in supranuclear gaze
palsy, also called saccadic gaze palsy. "Supranuclear" refers to
the location of the defect being superior to the relevant cranial
nerve nuclei in the midbrain of the brainstem (oculomoter nerve,
trochlear nerve) or the pons of the brainstem (abducens nerve). The
oculomoter, trochlear and abducens nerves are the only cranial
nerves controlling the small muscles which move the eye, and
lesions to the nerves themselves do not result in conjugate gaze
palsies.
[0007] Saccades are quick, simultaneous movements of both eyes
between two or more phases of fixation in the same direction.
Saccades are in contrast to smooth pursuit movements, in which the
eyes move smoothly without jumps, usually while tracking objects in
the visual field. Saccades serve as a mechanism for visual
fixation, rapid eye movement, and the fast phase of optokinetic
nystagmus. Saccades are controlled cortically by the frontal eye
fields region of the frontal cortex, or subcortically by the
superior colliculus (a region of the midbrain). Saccades are
particularly important during reading and when scanning the
immediate surroundings. Because the high-resolution region of the
retina, the fovea, is very small (about 1-2 degrees of vision
wide), saccadic eye movement is critical in resolving small objects
in the field of view. Skilled readers move their eyes during
reading on the average of every 250 milliseconds, and during each
saccade, lasting 20-40 milliseconds, the gaze target moves across
7-9 characters on average (range 1-20 characters).
[0008] The peak angular speed of the eye during a saccade can reach
up to 900 degrees per second in humans. Saccades in response to an
unexpected stimulus normally take only about 200 milliseconds to
initiate, and they last from about 20 milliseconds to 200
milliseconds depending upon amplitude. The amplitude of a saccade
is the angular distance the eye travels during the eye movement.
Head-fixed saccades can have amplitudes up to 90 degrees, but under
most condition, any shift of gaze larger than 20 degrees is
accompanied by head movement. During these gaze saccades, the eye
first undergoes saccade to shift the gaze to the target, and the
head follows more slowly while the eyes maintain focus on the
target. The latter is referred to as the vestibulo-ocular reflex
(VOR), and it function to slowly shift the eyes in the opposite
direction to the movement of the head in order to maintain visual
focus in the retinas. Because the head is almost always in slight
motion, VOR is essential to stabilizing vision under nearly all
circumstances, but especially during reading.
[0009] Saccadic gaze palsy may result in the slowing of saccades
either horizontally, vertically, or both, and may be either with or
without range limitations. Whether horizontal or vertical saccadic
palsy exists depends on the exact region of the brain involved in
pathology.
[0010] Gaucher disease (GD) is a rare, autosomal recessive,
lysosomal storage disease. GD patients have a mutation in the GBA1
gene which encodes glucosylceramidase (GC), also known as
beta-glucocerebrosidase. This enzyme is responsible for breaking
down glycosphingolipids into their components, such as breaking
down glucosylceramide (GLC; also known as glucocerebroside) into
glucose and ceramide. Monocytes and macrophages have a particularly
high content of lysosomes containing GLC, and in GD patients these
cells become enlarged and accumulate toxic concentrations of GLC.
These so-called "Gaucher cells" accumulate in several organs,
including the bone, bone marrow, spleen, liver, lung and brain.
Systemically, this results in splenomegaly, hepatomegaly, anemia,
thrombocytopenia, leukopenia, osteopenia, osteonecrosis, and other
pathologic abnormalities.
[0011] There are three subtypes of Gaucher disease, which differ in
the age of onset, severity, and presence of neurological
manifestations. Type 1 Gaucher disease (GD-1), non-neuronopathic
GD, is the most common form, with median age at diagnosis of 28,
and mildly reduced life expectancy. In GD-1, the GC enzyme retains
some functionality, and there is no neurological involvement.
Type-2 GD is acute neuronopathic GD, with diagnosis during infancy,
severe neurological involvement and death usually within the first
two years of life. The GC enzyme in a Type-2 patient is more
severely compromised in function compared to in GD-1. Type-3 GD is
chronic neuronopathic GD, with diagnosis during childhood,
gradually worsening neurological involvement, and life expectancy
usually not more than 30 years. Symptoms of GD-3 include spleen and
liver abnormalities, fatigue, bleeding, seizures and supranuclear
gaze palsy. The neurological manifestations in GD-3 patients
gradually develops over the course of the disease. One of the more
debilitating features is gaze palsy, which is a defect in the
neuronal pathways controlling saccadic eye movement. During early
stages of the disease, there is a slowing in horizontal saccades.
The disease progresses to complete horizontal saccadic palsy along
with varying degrees of vertical saccadic palsy. The VOR may also
be impaired in GD-3 patients. These features of the disease have a
profound impact on the quality of life of GD-3 patients, and can
hinder education and employment prospects.
[0012] Existing treatment for GD-1 and GD-3 are limited to
recombinant enzyme replacement therapy (ERT) using imiglucerase,
velaglucerase, or taliglucerase, and substrate reduction therapy
(SRT) using miglustat or eliglustat. See, e.g., Lunawati L. Bennett
& Chris Fellner, Pharmacotherapy of Gaucher Disease: Current
and Future Options, P&T 43(5): 274-280, 309 (2018).
Imiglucerase, the leading treatment regimen, is a recombinant
version of human GC, made in Chinese hamster ovary cells and
administered by slow intravenous injection (typically over 1-2
hours) every 1-2 weeks. It has been available since 1998 in the
U.S. Velaglucerase is another recombinant human GC analog, this one
made in a fibrosarcoma cell line, and it was FDA-approved in 2010.
Taliglucerase is similar, made using genetically modified carrot
plant root cells, and has been approved since 2012. These
treatments all require IV administration in a hospital or other
medical setting and the recombinant enzymes do not cross the
blood-brain barrier, and therefore, are not capable of treating the
neurological symptoms of GD. Thus, while these ERT regimes have
proven effective in treating GD-1 patients, in GD-3 patients they
are only effective in treating the non-neurological symptoms of the
disease.
[0013] Substrate-reduction therapy is an alternative approach to
treating GD. The goal of this therapy is to reduce the accumulation
of GLC by inhibiting the enzyme which is responsible for
synthesizing GLC. Glucosylceramide synthase (GCS), also known as
UDP-glucose ceramide synthase, is the enzyme which catalyzes the
initial glycosylation step of ceramide to form
glucosylceramide.
[0014] GCS inhibitors have been proposed for the treatment of a
variety of diseases, including glycolipid storage diseases and
lysosomal storage diseases, including Gaucher disease. See for
example, WO 2005/068426 (Actelion Pharm. Ltd.). Miglustat
(Zavesca), is an iminoglucose GCS inhibitor. It is an N-alkylated
iminosugar and acts as a reversible competitive inhibitor of GCS,
binding in the enzyme's active site. While it was developed to
treat the neuronopathic forms of GD, GD-2 and GD-3, the FDA has
only approved it for the treatment of patients with mild to
moderate GD-1, and only as a second-line therapy (patients must be
unable to receive ERT treatment). While miglustat does cross the
blood-brain barrier, in clinical trials it was found to be
ineffective in treating the neurological manifestations of GD-3.
Eliglustat is also a GCS inhibitor, and it is an analogue of the
ceremide. It has only been FDA-approved for treatment of the
systemic symptoms in GD-1 patients.
[0015] Niemann-Pick Disease Type C (NPC) is also a lysosomal
storage disease, although it's cause is quite different than
Gaucher disease, in some ways, the net result is similar. NPC is
caused by mutations in either the NPC1 or NPC2 genes. NPC1 is a
membrane protein which mediates intracellular trafficking of
cholesterol to post-lysosomal destinations. Specifically, NPC1 acts
in concert with NPC2 to promote the egress of cholesterol from the
endosomal/lysosomal compartment. Unesterified cholesterol that has
been released from low density lipoproteins in the lumen of the
late endosomes/lysosomes is transferred by NPC2 to the
cholesterol-binding pocket of NPC1. Approximately 95% of NPC
patients have mutations in NPC1, while most of the remainder have
mutations in NPC2. One of the effects of this disrupted cholesterol
trafficking is the accumulation of cholesterol and
glycosphingolipids (including GLC), in liver, spleen and brain
cells. One of the hallmarks of NPC, like GD3, is the progressive
development of supranuclear gaze palsy, including horizontal and
vertical saccadic palsies.
[0016] Another group of diseases and disorders commonly associated
with saccadic gaze palsies include the GM2-gangliosidoses (such as
Tay Sachs disease, Sandhoff disease and AB variant GM2
gangliosidosis).
[0017] GM2 gangliosidoses are, similar to Gaucher disease,
lysosomal storage diseases marked by genetic defects in
glycosphingolipid metabolism. GM2 gangliosidoses are marked by
defects in the enzyme hexosaminidase A and/or its co-factor GM2
activator protein, which are responsible for the breakdown of GM2
to GM3. GM2 and GM3 are related gangliosides which are part of the
same metabolic pathway in glucosylceramide is degraded to ceramide.
As such, GM3 is made by a stepwise process that begins with the
conversion of ceramide to glucosylceramide (by GLC), followed by
conversion to a galactosyl-glucosylceramide, followed by conversion
to GM3 (N-acetyl-a-neuraminidyl-galactosyl-glucosylceramide),
followed by conversion to GM2 (N-acetyl-galactosyl
N-acetyl-a-neuraminidyl-galactosyl-glucosylceramide). The
pathological accumulation of GM2 that is the hallmark of GM2
gangliosidoses can thus be ameliorated by a GCS inhibitor which
inhibits the earlier synthetic step of glucosylceramide.
[0018] The quinuclidine compounds described herein have activity as
inhibitors of the enzyme glucosylceramide synthase (GCS). These
compounds have been disclosed as generally being useful in the
treatment lysosomal storage diseases such as Fabry disease, Gaucher
disease and Niemann-Pick disease. See, e.g., WO 2012/129084 and
U.S. 2016/0361301.
[0019] There is a real need in the art to develop therapeutics
effective in alleviating or managing the neurological symptoms
associated with Gaucher Disease Type 3, especially the saccadic eye
movement deficits.
SUMMARY OF THE INVENTION
[0020] The present invention relates to a quinuclidine compound
(Compound 1) according to formula (I),
##STR00001##
[0021] or a pharmaceutically acceptable salt or prodrug thereof,
wherein: [0022] R.sup.1 is selected from hydrogen, halogen (e.g.,
fluorine), cyano, nitro, hydroxy, thio, amino, C.sub.1-6-alkyl
(e.g., methyl or ethyl), C.sub.2-6-alkenyl, C.sub.2-6-alkynyl,
C.sub.1-6-alkyloxy, C.sub.2-6-alkenyloxy, and C.sub.2-6-alkynyloxy,
wherein said alkyl, alkenyl, alkynyl, alkyloxy, alkenyloxy, or
alkynyloxy is optionally substituted with one or more (e.g., 1, 2
or 3) groups selected from halogen, cyano, nitro, hydroxy, thio or
amino; [0023] R.sup.2 and R.sup.3 are independently selected from
C.sub.1-3-alkyl, optionally substituted by one or more (e.g. 1, 2
or 3) halogens, or R.sup.2 and R.sup.3 together form a cyclopropyl
or cyclobutyl group, optionally substituted by one or more (e.g. 1
or 2) halogens; [0024] R.sup.4, R.sup.5 and R.sup.6 are each
independently selected from hydrogen, halogen, nitro, hydroxy,
thio, amino, C.sub.1-6-alkyl, and C.sub.1-6-alk.sub.yloxy, wherein
said alkyl or alkyloxy is optionally substituted by one or more
(e.g. 1, 2 or 3) groups selected from halogen, hydroxy, cyano, and
C.sub.1-6-alkyloxy; and [0025] A is a 5- or 6-membered aryl or
heteroaryl group, optionally substituted with 1, 2 or 3 groups
independently selected from a halogen, hydroxy, thio, amino, nitro,
C.sub.1-6alkoxy or C.sub.1-6alkyl.
[0026] In a first aspect, the present application provides a method
for treating or preventing supranuclear gaze palsies, including
horizontal and vertical saccadic gaze palsies, in a subject in need
thereof, the method comprising administering to the subject an
effective amount of a quinuclidine compound as described herein,
e.g., a compound according to Formula I. In other aspects, the
present application further provides use of the quinuclidine
compounds described herein, for the treatment or prevention of
supranuclear gaze palsies, including horizontal and vertical
saccadic gaze palsies, and/or for the manufacture of a medicament
for the treatment or prevention of supranuclear gaze palsies,
including horizontal and vertical saccadic gaze palsies.
[0027] Additional features and advantages of compounds,
compositions and methods disclosed herein will be apparent from the
following detailed description.
BRIEF DESCRIPTION OF THE DRAWINGS
[0028] FIGS. 1 and 2 show horizontal saccadic eye movement measured
in five patients as described in Example 5 (FIG. 1 shows patients
1-3 and FIG. 2 shows patients 4-5). Saccade amplitude and peak
velocity are measured when a target moves horizontally either
15.degree. (grey dots) or 30.degree. (black dots) from the center
position in either a leftward or rightward direction. Movements of
the eye to the right are represented by positive peak velocities
and movement to the left by negative peak velocities. The grey
shaded area on each plot represents the normal range of peak
velocities at any given amplitude.
DETAILED DESCRIPTION
[0029] Although specific embodiments of the present disclosure will
now be described with reference to the preparations and schemes, it
should be understood that such embodiments are by way of example
only and merely illustrative of but a small number of the many
possible specific embodiments which can represent applications of
the principles of the present disclosure. Various changes and
modifications will be obvious to those of skill in the art given
the benefit of the present disclosure and are deemed to be within
the spirit and scope of the present disclosure as further defined
in the appended claims.
[0030] Definitions
[0031] Unless defined otherwise, all technical and scientific terms
used herein have the same meanings as commonly understood by one of
ordinary skill in the art to which this disclosure belongs.
Although any methods and materials similar or equivalent to those
described herein can be used in the practice or testing of the
present invention, exemplary methods, devices, and materials are
now described. All technical and patent publications cited herein
are incorporated herein by reference in their entirety. Nothing
herein is to be construed as an admission that the invention is not
entitled to antedate such disclosure by virtue of prior
invention.
[0032] The practice of the present disclosure will employ, unless
otherwise indicated, conventional techniques of tissue culture,
immunology, molecular biology, microbiology, cell biology and
recombinant DNA, which are within the skill of the art.
[0033] All numerical designations, e.g., pH, temperature, time,
concentration, molecular weight, including ranges, are
approximations which are varied (+) or (-) by increments of 0.1 or
1.0, where appropriate. It is to be understood, although not always
explicitly stated, that all numerical designations are preceded by
the term "about". It also is to be understood, although not always
explicitly stated, that the reagents described herein are merely
exemplary and that equivalents of such are known in the art.
[0034] As used herein, the term "optionally substituted" is meant
to be equivalent to the phrase "non-substituted or substituted
by."
[0035] As used herein, the phrase "in a method of treating or
preventing" (such as in the phrase "in a method of treating or
preventing supranuclear gaze palsies") is meant to be equivalent to
the phrase "in the treatment or prevention of" (such as in the
phrase "in the treatment or prevention of supranuclear gaze
palsies").
[0036] As used in the specification and claims, the singular form
"a", "an" and "the" include plural references unless the context
clearly dictates otherwise. For example, the term "a cell" includes
a plurality of cells, including mixtures thereof. Unless
specifically stated or obvious from context, as used herein, the
term "or" is understood to be inclusive. The term "including" is
used herein to mean, and is used interchangeably with, the phrase
"including but not limited to".
[0037] As used herein, the term "comprising" or "comprises" is
intended to mean that the compositions and methods include the
recited elements, but not excluding others. "Consisting essentially
of" when used to define compositions and methods, shall mean
excluding other elements of any essential significance to the
combination for the stated purpose. Thus, a composition consisting
essentially of the elements as defined herein would not exclude
trace contaminants from the isolation and purification method and
pharmaceutically acceptable carriers, such as phosphate buffered
saline, preservatives and the like. "Consisting of" shall mean
excluding more than trace elements of other ingredients and
substantial method steps for administering the compositions of this
invention or process steps to produce a composition or achieve an
intended result. Embodiments defined by each of these transition
terms are within the scope of this invention. Use of the term
"comprising" herein is intended to encompass "consisting
essentially of" and "consisting of".
[0038] A "subject," "individual" or "patient" is used
interchangeably herein, and refers to a vertebrate, such as a
mammal. Mammals include, but are not limited to, murines, rats,
rabbit, simians, bovines, ovine, porcine, canines, felines, farm
animals, sport animals, pets, equines, primates, and humans. In one
embodiment, the mammals include horses, dogs, and cats. In some
embodiments, the mammal is a human, e.g., a human suffering from a
particular disease or disorder, such as Gaucher disease (e.g.,
GD-3) or Niemann-Pick disease Type C.
[0039] "Administering" is defined herein as a means of providing an
agent or a composition containing the agent to a subject in a
manner that results in the agent being inside the subject's body.
Such an administration can be by any route including, without
limitation, oral, transdermal (e.g. vagina, rectum, oral mucosa),
by injection (e.g. subcutaneous, intravenous, parenterally,
intraperitoneally, into the CNS), or by inhalation (e.g. oral or
nasal). Pharmaceutical preparations are, of course, given by forms
suitable for each administration route.
[0040] "Treating" or "treatment" of a disease generally includes:
(1) inhibiting the disease, i.e. arresting or reducing the
development of the disease or its clinical symptoms; and/or (2)
relieving the disease, i.e. causing regression of the disease or
its clinical symptoms.
[0041] As used herein, "treating" and "treatment" also refer to
either a reversal of the supranuclear gaze palsy or a stabilization
of the supranuclear gaze palsy. This is because the diseases and
disorders described herein are progressive disorders--in the
absence of treatment, the supranuclear gaze palsies will continue
to deteriorate until complete palsy (i.e., paralysis) results. For
example, early in the course of disease, a patient may suffer from
slowed or inhibited saccadic eye movements, but as the disease
progresses, patients may develop complete absence of saccades.
Treatment thus embraces both a slowing of this progressive
deterioration (e.g., stabilization), as well as reversal of this
progressive deterioration (e.g., improvement.
[0042] "Preventing" or "prevention" of a disease generally includes
causing the clinical symptoms of the disease not to develop in a
patient that may be predisposed to the disease but does not yet
experience or display symptoms of the disease.
[0043] As used herein, "preventing" or "prevention" also embraces
the prevention of development of a supranuclear gaze palsy in a
patient suspected of having or diagnosed as having a disease or
disorder described herein. Because the diseases and disorders
described herein are progressive disorders, different signs and
symptoms may manifest progressively as the disease advances. Thus,
for example, a patient may be diagnosed with GD-3 or NPC before
supranuclear gaze palsy begins developing. In such a patient, the
methods of treatment described herein may be effective in
preventing the supranuclear gaze palsy from developing.
[0044] The term "palsy" is synonymous with "paralysis" and includes
any degree of loss of motor function of one or more skeletal
muscles. As used herein, the term "palsy" thus embraces both
complete palsy, i.e., complete paralysis, as well as partial palsy.
Complete palsy means that a muscle or group of muscles, for example
the extraocular muscles, have lost the ability to contract. As
such, the effected eye or eyes may be unable to move. Partial palsy
may be manifested as an inhibition of movement, a slowing of
movement, or other defects in movement. These may include a loss of
range of motion. As applied to saccades, this can include
inhibition of initiating saccades (e.g., in response to stimuli),
changes in the frequency of saccades, changes in the peak velocity
of saccades, changes in the amplitude of saccades, changes in the
latency between saccades, and/or a loss of the ability to hold gaze
or to shift gaze. As used herein, in some embodiments, palsy
includes ophthalmoparesis and/or ophthalmoplegia. As such, the term
embraces both weakness and paralysis of the extraocular muscles.
The extraocular muscles include any one or more of the superior
recti, inferior recti, medial recti, lateral recti, inferior
oblique and superior oblique muscles of the eye. Weakness and/or
paralysis may include one or more of horizontal movement, vertical
movement or rotational movement.
[0045] The term "suffering" as it relates to the term "treatment"
refers to a patient or individual who has been diagnosed with the
disease. The term "suffering" as it relates to the term
"prevention" refers to a patient or individual who is predisposed
to the disease. A patient may also be referred to being "at risk of
suffering" from a disease because of a history of disease in their
family lineage or because of the presence of genetic mutations
associated with the disease. A patient at risk of a disease has not
yet developed all or some of the characteristic pathologies of the
disease.
[0046] An "effective amount" or "therapeutically effective amount"
is an amount sufficient to effect beneficial or desired results. An
effective amount can be administered in one or more
administrations, applications or dosages. Such delivery is
dependent on a number of variables including the time period for
which the individual dosage unit is to be used, the bioavailability
of the therapeutic agent, and the route of administration. It is
understood, however, that specific dose levels of the therapeutic
agents of the present invention for any particular subject depends
upon a variety of factors including, for example, the activity of
the specific compound employed, the age, body weight, general
health, sex, and diet of the subject, the time of administration,
the rate of excretion, the drug combination, and the severity of
the particular disorder being treated and form of administration.
Treatment dosages generally may be titrated to optimize safety and
efficacy. Typically, dosage-effect relationships from in vitro
and/or in vivo tests initially can provide useful guidance on the
proper doses for patient administration. In general, one will
desire to administer an amount of the compound that is effective to
achieve a serum level commensurate with the concentrations found to
be effective in vitro. Determination of these parameters is well
within the skill of the art. These considerations, as well as
effective formulations and administration procedures are well known
in the art and are described in standard textbooks. Consistent with
this definition, as used herein, the term "therapeutically
effective amount" is an amount sufficient to treat (e.g. improve)
one or more symptoms associated with a disease or disorder
described herein (e.g., in any of Method 1 et seq., or Method 4 et
seq.) ex vivo, in vitro or in vivo.
[0047] As used herein, the term "pharmaceutically acceptable
excipient" encompasses any of the standard pharmaceutical
excipients, including carriers such as a phosphate buffered saline
solution, water, and emulsions, such as an oil/water or water/oil
emulsion, and various types of wetting agents. Pharmaceutical
compositions also can include stabilizers and preservatives. For
examples of carriers, stabilizers and adjuvants, see Remington's
Pharmaceutical Sciences (20th ed., Mack Publishing Co. 2000).
[0048] As used herein, the term "prodrug" means a pharmacological
derivative of a parent drug molecule that requires
biotransformation, either spontaneous or enzymatic, within the
organism to release the active drug. For example, prodrugs are
variations or derivatives of the quinuclidine compounds described
herein that have groups cleavable under certain metabolic
conditions, which when cleaved, become the quinuclidine compounds
described herein, e.g. a compound of Formula I. Such prodrugs then
are pharmaceutically active in vivo when they undergo solvolysis
under physiological conditions or undergo enzymatic degradation.
Prodrug compounds herein may be called single, double, triple,
etc., depending on the number of biotransformation steps required
to release the active drug within the organism, and the number of
functionalities present in a precursor-type form. Prodrug forms
often offer advantages of solubility, tissue compatibility, or
delayed release in the mammalian organism.
[0049] Prodrugs commonly known in the art include well-known acid
derivatives, such as, for example, esters prepared by reaction of
acid compounds with a suitable alcohol, amides prepared by reaction
of acid compounds with an amine, and basic groups reacted to form
an acylated base derivative. Other prodrug derivatives may be
combined with other features disclosed herein to enhance
bioavailability. As such, those of skill in the art will appreciate
that certain of the presently disclosed compounds having, for
example, free amino or hydroxy groups can be converted into
prodrugs. Prodrugs include compounds having an amino acid residue,
or a polypeptide chain of two or more (e.g. two, three or four)
amino acid residues which are covalently joined through peptide
bonds to free amino, hydroxy or carboxylic acid groups of the
presently disclosed compounds. The amino acid residues include the
20 naturally occurring amino acids commonly designated by three
letter symbols and also include 4-hydroxyproline, hydroxylysine,
demosine, isodemosine, 3-methylhistidine, norvalin, beta-alanine,
gamma-aminobutyric acid, citrulline, homocysteine, homoserine,
ornithine and methionine sulfone. Prodrugs also include compounds
having a carbonate, carbamate, amide or alkyl ester moiety
covalently bonded to any of the above substituents disclosed
herein.
[0050] As used herein, the term "pharmaceutically acceptable salt"
means a pharmaceutically acceptable acid addition salt or a
pharmaceutically acceptable base addition salt of a currently
disclosed compound that may be administered without any resultant
substantial undesirable biological effect(s) or any resultant
deleterious interaction(s) with any other component of a
pharmaceutical composition in which it may be contained.
[0051] As used herein, the term "C.sub.1-6-alkyl" means a saturated
linear or branched free radical consisting essentially of 1 to 6
carbon atoms and a corresponding number of hydrogen atoms.
Exemplary C.sub.1-6-alkyl groups include methyl, ethyl, n-propyl,
isopropyl, n-butyl, and isobutyl. Other C.sub.1-6-alkyl groups will
be readily apparent to those of skill in the art given the benefit
of the present disclosure. The terms "C.sub.1-3-alkyl",
"C.sub.1-4-alkyl", etc., have equivalent meanings, i.e., saturated
linear or branched free radical consisting essentially of 1 to 3
(or 4) carbon atoms and a corresponding number of hydrogen
atoms.
[0052] As used herein, the term "C.sub.2-6-alkenyl" means an
unsaturated linear or branched free radical consisting essentially
of 2 to 6 carbon atoms and a corresponding number of hydrogen
atoms, which free radical comprises at least one carbon-carbon
double bond. Exemplary C.sub.2-6-alkenyl groups include ethenyl,
prop-1-enyl, prop-2-enyl, isopropenyl, but-1-enyl,
2-methyl-prop-1-enyl, and 2-methyl-prop-2-enyl. Other
C.sub.2-6-alkenyl groups will be readily apparent to those of skill
in the art given the benefit of the present disclosure.
[0053] As used herein, the term "C.sub.2-6-alkynyl" means an
unsaturated linear or branched free radical consisting essentially
of 2 to 6 carbon atoms and a corresponding number of hydrogen
atoms, which free radical comprises at least one carbon-carbon
triple bond. Exemplary C.sub.2-6-alkynyl groups include ethynyl,
prop-1-ynyl, prop-2-ynyl, but-1-ynyl, and 3-methyl-but-1-ynyl.
Other C.sub.2-6-alkynyl groups will be readily apparent to those of
skill in the art given the benefit of the present disclosure.
[0054] As used herein, the term "C.sub.1-6-alkyloxy" means a
saturated linear or branched free radical consisting essentially of
1 to 6 carbon atoms (and a corresponding number of hydrogen atoms)
and an oxygen atom. A C.sub.1-6-alkyloxy group is attached via the
oxygen atom. Exemplary C.sub.1-6-alkyloxy groups include methyloxy,
ethyloxy, n-propyloxy, isopropyloxy, n-butyloxy, and isobutyloxy.
Other C.sub.1-6-alkyloxy groups will be readily apparent to those
of skill in the art given the benefit of the present disclosure.
The terms "C.sub.1-3-alkyloxy", "C.sub.1-4-alkyloxy", and the like,
have an equivalent meaning, i.e. a saturated linear or branched
free radical consisting essentially of 1 to 3 (or 4) carbon atoms
(and a corresponding number of hydrogen atoms) and an oxygen atom,
wherein the group is attached via the oxygen atom.
[0055] As used herein, the term "C.sub.2-6-alkenyloxy" means an
unsaturated linear or branched free radical consisting essentially
of 2 to 6 carbon atoms (and a corresponding number of hydrogen
atoms) and an oxygen atom, which free radical comprises at least
one carbon-carbon double bond. A C.sub.2-6-alkenyloxy group is
attached via the oxygen atom. An exemplary C.sub.2-6-alkenyloxy
group is ethenyloxy; others will be readily apparent to those of
skill in the art given the benefit of the present disclosure.
[0056] As used herein, the term "C.sub.2-6-alkynyloxy" means an
unsaturated linear or branched free radical consisting essentially
of 2 to 6 carbon atoms (and a corresponding number of hydrogen
atoms) and an oxygen atom, which free radical comprises at least
one carbon-carbon triple bond. A C.sub.2-6-alkenyloxy group is
attached via the oxygen atom. An exemplary C.sub.2-6-alkenyloxy
group is ethynyloxy; others will be readily apparent to those of
skill in the art given the benefit of the present disclosure.
[0057] As used herein, the term "heteroaryl" means an aromatic free
radical having 5 or 6 atoms (i.e. ring atoms) that form a ring,
wherein 1 to 5 of the ring atoms are carbon and the remaining 1 to
5 ring atom(s) (i.e. hetero ring atom(s)) is selected independently
from the group consisting of nitrogen, sulfur, and oxygen.
Exemplary 5-membered heteroaryl groups include furyl, thienyl,
thiazolyl (e.g. thiazol-2-yl), pyrazolyl, isothiazolyl, oxazolyl,
isoxazolyl, pyrrolyl, triazolyl, imidazolyl, oxadiazolyl and
thiadiazolyl. Exemplary 6-membered heteroaryl groups include
pyridyl, pyrimidyl, pyrazinyl, pyridazinyl, 1,2,4-triazinyl,
benzoxazolyl, benzothiazolyl, benzisothiazolyl, benzisoxazolyl, and
benzimidazolyl. Other heteroaryl groups will be readily apparent to
those of skill in the art given the benefit of the present
disclosure. In general, the heteroaryl group typically is attached
to the main structure via a carbon atom. However, those of skill in
the art will realize that certain other atoms, e.g. hetero ring
atoms, can be attached to the main structure.
[0058] As used herein, the term "aryl" means an aromatic free
radical having 5 or 6 atoms (i.e. ring atoms) that form a ring,
wherein all of the ring atoms are carbon. An exemplary aryl group
is a phenyl group.
[0059] As used herein, the term "aliphatic" means a non-aromatic
compound containing carbon and hydrogen atoms, e.g. containing 1 to
9 carbon atoms. Aliphatic compounds may be straight-chained or
branched, may contain one or more ring structures, and may contain
one or more carbon-carbon double bonds (provided that the compound
does not contain an unsaturated ring structure having aromatic
character). Examples of aliphatic compounds include ethane,
propylene, cyclobutane, and cyclohexadiene.
[0060] As used herein, the terms "halo" and "halogen" mean
fluorine, chlorine, bromine, or iodine. These terms are used
interchangeably and may refer to a halogen free radical group or to
a halogen atom as such. Those of skill in the art will readily be
able to ascertain the identification of which in view of the
context in which this term is used in the present disclosure.
[0061] As used herein, the term "cyano" means a free radical having
a carbon atom linked to a nitrogen atom via a triple bond. The
cyano radical is attached via its carbon atom.
[0062] As used herein, the term "nitro" means an --NO.sub.2 radical
which is attached via its nitrogen atom.
[0063] As used herein, the terms "hydroxy" and "hydroxyl" mean an
--OH radical which is attached via its oxygen atom. The term "thio"
means an --SH radical which is attached via its sulfur atom.
[0064] As used herein, the term "amino" means a free radical having
a nitrogen atom and 1 or 2 hydrogen atoms. As such, the term
"amino" generally refers to primary and secondary amines. In that
regard, as used herein, a tertiary amine is represented by the
general formula RR'N--, wherein R and R' are carbon radicals that
may or may not be identical. Nevertheless, the term "amino"
generally may be used herein to describe a primary, secondary, or
tertiary amine, and those of skill in the art will readily be able
to ascertain the identification of which in view of the context in
which this term is used in the present disclosure.
[0065] As used herein, the term and "oxo" means an oxygen radical
which is attached via a double bond. Where an atom bonded to this
oxygen is a carbon atom, the bond is a carbon-oxygen double bond
which may be denoted as --(C.dbd.O)-- and which may be referred to
as a ketone.
[0066] The recitation of a listing of chemical groups in any
definition of a variable herein includes definitions of that
variable as any single group or combination of listed groups. The
recitation of an embodiment for a variable or aspect herein
includes that embodiment as any single embodiment or in combination
with any other embodiments or portions thereof.
[0067] Any compositions or methods provided herein can be combined
with one or more of any of the other compositions and methods
provided herein.
[0068] The following abbreviations are used herein:
[0069] br Broad signal
[0070] CDI Carbonyldiimidazole
[0071] CNS Central Nervous System
[0072] d Doublet
[0073] DAPI 4',6-diamidino-2-phenylindole
[0074] dd Doublet of doublets
[0075] DME Dimethoxyethane
[0076] DMEM Dulbecco Modified Eagle Medium
[0077] DMSO-d.sub.6 Dimethyl sulfoxide-d6
[0078] DMF Dimethylformamide
[0079] DNA Deoxyribonucleic acid
[0080] DTBZ Carbon-11 dihydrotetrabenazine
[0081] EDTA Ethylenediaminetetraacetic acid
[0082] ELISA Enzyme-linked Immunosorbent Assay
[0083] Et.sub.2O Diethyl ether
[0084] EtMgBr Ethylmagnesium bromide
[0085] EtOAc Ethyl acetate
[0086] GL1 Glucosylceramide (GlcCer)
[0087] GM1 Monosialotetrahexosylganglioside
[0088] GM3 Monosialodihexosylganglioside
[0089] GSL Glycosphingolipid
[0090] H&E Hematoxylin and eosin stain
[0091] HPLC High pressure/performance liquid chromatography
[0092] HSA Human serum albumin
[0093] IPA Isopropyl alcohol
[0094] J Coupling constant
[0095] LCMS Liquid chromatography mass spectrometry
[0096] m Multiplet
[0097] ppm Parts per million
[0098] rHA Recombinant human albumin
[0099] s Singlet
[0100] TBME Tert-Butyl Methyl Ether
[0101] THF Tetrahydrofuran
[0102] Tris Tris(hydroxymethyl)aminomethane
[0103] TWEEN20 Polysorbate 20
[0104] TWEEN80 Polysorbate 80
[0105] WT Wild type
[0106] UPLCMS Ultra performance liquid chromatography mass
spectrometry
[0107] Compounds
[0108] The present disclosure relates to quinuclidine compounds for
use in therapeutic methods relating to the treatment or prevention
of the diseases and disorders discussed herein. In all of its
various aspects, the invention relates to a quinuclidine compound
(Compound 1) according to formula (I),
##STR00002##
[0109] or a pharmaceutically acceptable salt or prodrug thereof,
wherein: [0110] R.sup.1 is selected from hydrogen, halogen (e.g.,
fluorine), cyano, nitro, hydroxy, thio, amino, C.sub.1-6-alkyl
(e.g., methyl or ethyl), C.sub.2-6-alkenyl, C.sub.2-6-alk.sub.ynyl,
C.sub.1-6-alkyloxy, C.sub.2-6-alkenyloxy, and C.sub.2-6-alkynyloxy,
wherein said alkyl, alkenyl, alkynyl, alkyloxy, alkenyloxy, or
alkynyloxy is optionally substituted with one or more (e.g., 1, 2
or 3) groups selected from halogen, cyano, nitro, hydroxy, thio or
amino; [0111] R.sup.2 and R.sup.3 are independently selected from
C.sub.1-3-alkyl, optionally substituted by one or more (e.g. 1, 2
or 3) halogens, or R.sup.2 and R.sup.3 together form a cyclopropyl
or cyclobutyl group, optionally substituted by one or more (e.g. 1
or 2) halogens; [0112] R.sup.4, R.sup.5 and R.sup.6 are each
independently selected from hydrogen, halogen, nitro, hydroxy,
thio, amino, C.sub.1-6-alkyl, and C.sub.1-6-alk.sub.yloxy, wherein
said alkyl or alkyloxy is optionally substituted by one or more
(e.g. 1, 2 or 3) groups selected from halogen, hydroxy, cyano, and
C.sub.1-6-alkyloxy; and [0113] A is a 5- or 6-membered aryl or
heteroaryl group (e.g., phenyl or thiazolyl), optionally
substituted with 1, 2 or 3 groups independently selected from
halogen, hydroxy, thio, amino, nitro, C.sub.1-6alkoxy and
C.sub.1-6alkyl.
[0114] In further embodiments of the any aspects of the present
disclosure, the present disclosure further relates to Compounds as
follows: [0115] 1.1 Compound 1, wherein R.sup.1 is selected from
hydrogen, halogen, cyano, nitro, hydroxy, thio, amino,
C.sub.1-6-alkyl, C.sub.1-6-alkyloxy, wherein said alkyl or alkyloxy
is optionally substituted with one or more (e.g., 1, 2 or 3) groups
selected from halogen, cyano, nitro, hydroxy, thio or amino; [0116]
1.2 Compound 1, wherein R.sup.1 is selected from hydrogen, halogen,
C.sub.1-6-alkyl, C.sub.1-6-alkyloxy, wherein said alkyl or alkyloxy
is optionally substituted with one or more (e.g., 1, 2 or 3) groups
selected from halogen, cyano, nitro, hydroxy, thio or amino; [0117]
1.3 Compound 1, wherein R.sup.1 is selected from hydrogen, halogen,
C.sub.1-4-alkyl, C.sub.1-4-alkyloxy, wherein said alkyl or alkyloxy
is optionally substituted with one or more (e.g., 1, 2 or 3) groups
selected from halogen, cyano, nitro, hydroxy, thio or amino; [0118]
1.4 Compound 1, wherein R.sup.1 is selected from hydrogen, halogen,
C.sub.1-4-alkyl, C.sub.1-4-alkyloxy, wherein said alkyl or alkyloxy
is optionally substituted with one or more (e.g., 1, 2 or 3, or 1
or 2) groups selected from cyano, nitro, hydroxy, thio or amino;
[0119] 1.5 Compound 1, wherein R.sup.1 is selected from hydrogen,
halogen, and C.sub.1-4-alkyl, wherein said alkyl is optionally
substituted with one or more (e.g., 1 or 2) groups selected from
halogen, hydroxy, thio or amino; [0120] 1.6 Compound 1, wherein
R.sup.1 is selected from hydrogen, fluorine, methyl and ethyl,
wherein said methyl or ethyl is optionally substituted with 1 or 2
groups selected from halogen, hydroxy, thio or amino; [0121] 1.7
Compound 1, wherein R.sup.1 is selected from hydrogen and methyl,
wherein said methyl is optionally substituted with 1 or 2 halogens;
[0122] 1.8 Compound 1, wherein R.sup.1 is hydrogen; [0123] 1.9
Compound 1, or any of 1.1-1.8, wherein R.sup.1 is not attached to
the nitrogen atom of the quinuclidine moiety; [0124] 1.10 Compound
1, or any of 1.1-1.9, wherein R.sup.2 and R.sup.3 are each
independently C.sub.1-3-alkyl, optionally substituted by one or
more (e.g. 1, 2 or 3) halogens; [0125] 1.11 Compound 1.11, wherein
R.sup.2 and R.sup.3 are each independently methyl or ethyl,
optionally substituted by 1 or 2 halogens; [0126] 1.12 Compound
1.11, wherein R.sup.2 and R.sup.3 are each independently selected
from methyl and ethyl, optionally substituted by one or more
fluorines, e.g., 1, 2 or 4 fluorines; [0127] 1.13 Compound 1.11,
wherein R.sup.2 and R.sup.3 are each independently methyl
substituted with 0, 1, 2 or 3 fluorines; [0128] 1.14 Compound 1.11,
wherein R.sup.2 and R.sup.3 are each methyl or trifluoromethyl;
[0129] 1.15 Compound 1.11, R.sup.2 and R.sup.3 are each methyl;
[0130] 1.16 Compound 1, or any of 1.1-1.9, wherein R.sup.2 and
R.sup.3 together form a cyclopropyl or cyclobutyl group, optionally
substituted by one or more (e.g. 1 or 2) halogens; [0131] 1.17
Compound 1.16, wherein R.sup.2 and R.sup.3 together form a
cyclopropyl group; 1.18 Compound 1 or any of 1.1-1.9, wherein
R.sup.2 and R.sup.3 are each methyl or R.sup.2 and R.sup.3 together
form a cyclopropyl group; [0132] 1.19 Compound 1, or any of
1.1-1.9, wherein R.sup.4, R.sup.5 and R.sup.6 are each
independently selected from hydrogen, halogen, C.sub.1-6-alkyl, and
C.sub.1-6-alkyloxy, wherein said alkyl or alkyloxy is optionally
substituted by one or more (e.g. 1, 2 or 3) groups selected from
halogen, hydroxy, cyano, and C.sub.1-6-alkyloxy; [0133] 1.20
Compound 1, or any of 1.1-1.9, wherein R.sup.4, R.sup.5 and R.sup.6
are each independently selected from hydrogen, halogen,
C.sub.1-3-alkyl, and C.sub.1-3-alkyloxy, wherein said alkyl or
alkyloxy is optionally substituted by one or more (e.g. 1, 2 or 3)
groups selected from halogen, hydroxy, cyano, and
C.sub.1-3-alkyloxy; [0134] 1.21 Compound 1.19, wherein R.sup.4,
R.sup.5 and R.sup.6 are each independently selected from hydrogen,
halogen, C.sub.1-3-alkyl, and C.sub.1-3-alkyloxy, wherein said
alkyl or alkyloxy is optionally substituted by one or more (e.g. 1,
2 or 3) groups selected from halogen, cyano, and
C.sub.1-3-alkyloxy; [0135] 1.22 Compound 1.19, wherein R.sup.4,
R.sup.5 and R.sup.6 are each independently selected from hydrogen,
halogen, C.sub.1-3-alkyl, and C.sub.1-3-alkyloxy, wherein said
alkyl or alkyloxy is optionally substituted by one or more (e.g. 1,
2 or 3) groups selected from halogen and C.sub.1-3-alkyloxy; [0136]
1.23 Compound 1.19, wherein R.sup.4, R.sup.5 and R.sup.6 are each
independently selected from halogen, C.sub.1-3-alkyl, and
C.sub.1-3-alkyloxy, wherein said alkyl or alkyloxy is optionally
substituted by one or more (e.g. 1, 2 or 3) groups selected from
halogen and C.sub.1-3-alkyloxy [0137] 1.24 Compound 1, or any of
1.19-1.23, R.sup.4 is selected from hydrogen, halogen,
C.sub.1-3-alkyl, and C.sub.1-3-alkyloxy, wherein said alkyl or
alkyloxy is optionally substituted by one or more (e.g. 1, 2 or 3)
groups selected from halogen and C.sub.1-3-alkyloxy; [0138] 1.25
Compound 1.24, R.sup.4 is selected from halogen (e.g., fluorine),
C.sub.1-3-alkyl (e.g., methyl), and C.sub.1-3-alkyloxy (e.g.,
methoxy or ethoxy), wherein said alkyl or alkyloxy is optionally
substituted by one or more (e.g. 1, 2 or 3) groups selected from
halogen and C.sub.1-3-alkyloxy (e.g., methoxy or ethoxy); [0139]
1.26 Compound 1.26, R.sup.4 is selected from halogen (e.g.,
fluorine) and C.sub.1-3-alkyloxy (e.g., methoxy or ethoxy), wherein
said alkyloxy is optionally substituted by one or more (e.g. 1, 2
or 3) groups selected from halogen and C.sub.1-3-alkyloxy (e.g.,
methoxy or ethoxy); [0140] 1.27 Compound 1.26, R.sup.4 is fluorine
or C.sub.1-3-alkyloxy (e.g., ethoxy), optionally substituted by one
or more (e.g. 1, 2 or 3) groups selected from halogen and
C.sub.1-3-alkyloxy (e.g., methoxy); [0141] 1.28 Compound 1.26,
wherein R.sup.4 is fluorine or ethoxy optionally substituted by one
or more (e.g. 1, 2 or 3) C.sub.1-3-alkyloxy (e.g., methoxy); [0142]
1.29 Compound 1, or any of 1.19-1.28, wherein R.sup.6 is hydrogen;
[0143] 1.30 Compound 1, or any of 1.19-1.28, wherein R.sup.5 and
R.sup.6 are each hydrogen; [0144] 1.31 Compound 1, or any of
1.19-1.28, R.sup.5 and R.sup.6 are each hydrogen, and R.sup.4 is
fluorine or C.sub.1-3-alkyloxy (e.g., ethoxy), optionally
substituted by one or more (e.g. 1, 2 or 3) groups selected from
halogen and C.sub.1-3-alkyloxy (e.g., methoxy); [0145] 1.32
Compound 1.31, wherein R.sup.5 and R.sup.6 are each hydrogen, and
R.sup.4 is fluorine or ethoxy optionally substituted by one or more
(e.g. 1, 2 or 3) C.sub.1-3-alkyloxy (e.g., methoxy); [0146] 1.33
Compound 1.32, wherein R.sup.5 and R.sup.6 are each hydrogen, and
R.sup.4 is fluorine or ethoxy substituted with methoxy (e.g.,
2-methoxyethoxy); [0147] 1.34 Compound 1.32, wherein R.sup.4 is
fluorine or 2-methoxyethoxy; [0148] 1.35 Compound 1, or any of
1.1-1.34, wherein at least one of R.sup.4, R.sup.5 and R.sup.6 is
not hydrogen; [0149] 1.36 Compound 1, or any of 1.1-1.35, wherein
R.sup.6 is hydrogen, and R.sup.4 and R.sup.5 are positioned at the
2, 4 or 6 positions of the phenyl ring to which they are attached
(i.e., ortho or para to the A substituent); [0150] 1.37 Compound 1,
or any of 1.1-1.35, wherein R.sup.6 is hydrogen, and R.sup.4 and
R.sup.5 are positioned independently at the 2 and 3 (i.e., adjacent
ortho and meta), 3 and 4 (i.e. adjacent meta and para), or 3 and 5
positions (i.e., meta) of the phenyl ring to which they are
attached (with respect to the A substituent); [0151] 1.38 Compound
1, or any of 1.1-1.35, wherein R.sup.6 is hydrogen, and R.sup.4 and
R.sup.5 are positioned at the 3 and 5 positions (i.e., meta) of the
phenyl ring to which they are attached (with respect to the A
substituent); [0152] 1.39 Compound 1, or any of 1.1-1.35, wherein
R.sup.5 and R.sup.6 are hydrogen, and R.sup.4 is positioned at the
2, 3 or 4 position of the phenyl ring to which it is attached
(e.g., ortho, meta or para or to the A substituent); [0153] 1.40
Compound 1, or any of 1.1-1.35, wherein R.sup.5 and R.sup.6 are
hydrogen, and R.sup.4 is positioned at the 2 or 4 position of the
phenyl ring to which it is attached (e.g., ortho or para to the A
substituent); [0154] 1.41 Compound 1, or any of 1.1-1.35, wherein
R.sup.5 and R.sup.6 are hydrogen, and R.sup.4 is positioned at the
4 position of the phenyl ring to which it is attached (e.g., para
to the A substituent); [0155] 1.42 Compound 1, or any of 1.1-1.35,
wherein none of R.sup.4, R.sup.5 and R.sup.6 are hydrogen, and each
of R.sup.4, R.sup.5 and R.sup.6 are independently positioned at the
2, 4 or 6 positions of the phenyl ring to which they are attached
(i.e., ortho or para to the A substituent); [0156] 1.43 Compound 1,
or any of 1.1-1.42, wherein R.sup.4 is positioned at the 4-position
of the phenyl ring to which it is attached (i.e., para to the A
substituent); [0157] 1.44 Compound 1, or any of 1.1-1.43, wherein A
is a 6-membered aryl group, a 5-membered heteroaryl group (e.g.,
containing 1, 2 or 3 heteroatoms in the heteroaryl ring selected
from N, O and S), or a 6-membered heteroaryl group (e.g.,
containing 1, 2 or 3 nitrogen atoms in the heteroaryl ring); [0158]
1.45 Compound 1.44, wherein A is a 6-membered aryl group or a
5-membered heteroaryl group (e.g., containing 1, 2 or 3 heteroatoms
in the heteroaryl ring selected from N, O and S), optionally
wherein the 5-membered heteroaryl group contains 1 or 2 heteroatoms
selected from N and S (e.g., one N and/or one S); [0159] 1.46
Compound 1.44 or 1.45, wherein A is selected from the group
consisting of phenyl, furyl, thienyl, thiazolyl, pyrazolyl,
isothiazolyl, oxazolyl, isoxazolyl, pyrrolyl, triazolyl,
imidazolyl, oxadiazolyl and thiadiazolyl; [0160] 1.47 Compound
1.46, wherein A is selected from the group consisting of phenyl,
thienyl, thiazolyl, pyrrolyl, and imidazolyl; [0161] 1.48 Compound
1.46, wherein A is selected from the group consisting of phenyl and
thiazolyl, e.g., 2-thiazol-4-yl or 4-thiazol-2-yl; [0162] 1.49
Compound 1, or any of 1.1-1.48, wherein A is unsubstituted [0163]
1.50 Compound 1, or any of 1.1-1.48, wherein A is substituted with
one or more (e.g., 1, 2 or 3) groups independently selected from a
halogen, hydroxy, thio, amino, nitro, C.sub.1-6alkoxy and
C.sub.1-6alkyl (e.g., methyl); [0164] 1.51 Compound 1.50, wherein A
is thiazolyl substituted with one halogen (e.g., fluorine), or
C.sub.1-6alkyl (e.g., methyl); [0165] 1.52 Compound 1.50, wherein A
is phenyl substituted with 1, 2 or 3 groups independently selected
from halogen (e.g., fluorine) and C.sub.1-6alkyl (e.g., methyl);
[0166] 1.53 Compound 1.52, wherein A is phenyl substituted with 1
or 2 fluorines or methyl groups; [0167] 1.54 Compound 1, or any of
1.1-1.53 wherein the two groups attached to the A substituent
(i.e., the phenyl ring (--(C.sub.6H.sub.2R.sup.4R.sup.5R.sup.6))
and the --C(R.sup.2R.sup.3)-- group) are positioned in a 1,2-, 1,3-
or 1,4-relationship to each other (i.e., ortho, meta, or para);
[0168] 1.55 Compound 1.54, wherein the two groups attached to the A
substituent are positioned in a 1,3-relationship to each other
(i.e., meta); [0169] 1.56 Compound 1.54, wherein the two groups
attached to the A substituent are positioned in a 1,4-relationship
to each other (i.e., para); [0170] 1.57 Any of Compounds 1.54 to
1.56, wherein the A substituent is a 5-membered heteroaryl group
and at least one of the two groups attached to the A substituent
(i.e., the phenyl ring (--(C.sub.6H.sub.2R.sup.4R.sup.5R.sup.6)) or
the --C(R.sup.2R.sup.3)-- group) is attached to a carbon atom of
the heteroaryl ring, optionally wherein both of such groups are
attached to carbon atoms of the heteroaryl ring; [0171] 1.58
Compound 1, or any of 1.1-1.57, wherein the Compound of Formula I
can be represented by any one or more of the following
substructures:
[0171] ##STR00003## ##STR00004## [0172] 1.59 Compound 1, or any of
1.1-1.58, wherein the compound of Formula I, or any of Formulas II
to XII, has the (S) configuration; [0173] 1.60 Compound 1, or any
of 1.1-1.58, wherein the compound of Formula I, or any of Formulas
II to XII, has the (R) configuration; [0174] 1.61 Compound 1, or
any of 1.1-1.60, wherein the compound of Formula I, or any of
Formulas II to XII, has an enantiomeric excess (e.g., of the (S)
configuration) of at least 90%, e.g., at least 92%, 94%, 95%, 96%,
97%, 98%, 99%, 99.5% or 99.9%; [0175] 1.62 Compound 1, or any of
1.1-1.58, wherein the compound of Formula I, or any of Formulas II
to XII, is racemic (i.e., approximately a 50:50 ratio of
enantiomers), or is a mixture of enantiomers of some other ratio
(e.g., less than 50:50 or greater than 50:50); [0176] 1.63 Compound
1, or any of 1.1-1.62, wherein the Compound of Formula I is
selected from the group consisting of:
TABLE-US-00001 [0176] Compound No. Compound 1 Quinuclidin-3-yl
(2-(4'-fluoro-[1,1'-biphenyl]-3-yl)propan-2- yl)carbamate 2
(S)-quinuclidin-3-yl (2-(2-(4-fluorophenyl)thiazol-4-
yl)propan-2-yl)carbamate 3 (S)-quinuclidin-3-yl
(2-(4'-(2-methoxyethoxy)-[1,1'-
biphenyl]-4-yl)propan-2-yl)carbamate 4 1-azabicyclo[2.2.2]oct-3-yl
[2-(biphenyl-3-yl)propan-2- yl]carbamate 5 (S)-quinuclidin-3-yl
2-(biphenyl-4-yl)propan-2-ylcarbamate 6 Quinuclidin-3-yl
1-(biphenyl-4-yl)cyclopropylcarbamate 7 (S)-quinuclidin-3-yl
1-(4'-fluorobiphenyl-4- yl)cyclopropylcarbamate 8
(S)-1-azabicyclo[2.2.2]oct-3-yl [1-(2',4'-difluorobiphenyl-4-
yl)cyclopropyl]carbamate 9 1-azabicyclo[2.2.2]oct-3-yl
[1-(4'-methoxybiphenyl-4- yl)cyclopropyl]carbamate 10
Quinuclidin-3-yl 2-(5-(4-fluorophenyl)thiophen-3-yl)propan-
2-ylcarbamate 11 (S)-quinuclidin-3-yl
2-(3-(4-fluorophenyl)isothiazol-5- yl)propan-2-ylcarbamate 12
(S)-quinuclidin-3-yl 2-(4-(4-fluorophenyl)thiazol-2-
yl)propan-2-ylcarbamate 13 Quinuclidin-3-yl
(2-(4'-(2-methoxyethoxy)-[1,1'-biphenyl]-4-
yl)propan-2-yl)carbamate 14 (S)-quinuclidin-3-yl
(2-(3'-(2-methoxyethoxy)-[1,1'-
biphenyl]-4-yl)propan-2-yl)carbamate 15 Quinuclidin-3-yl
(2-(4'-(2-methoxyethoxy)- [1,1'-biphenyl]-
3-yl)propan-2-yl)carbamate 16 Quinuclidin-3-yl
(2-(4'-(3-methoxypropoxy)-[1,1'-biphenyl]-
4-yl)propan-2-yl)carbamate 17 Quinuclidin-3-yl
(2-(4'-(hydroxymethyl)-[1,1'-biphenyl]-4- yl)propan-2-yl)carbamate
18 Quinuclidin-3-yl (2-(4'-(2-hydroxyethyl)-[1,1'-biphenyl]-4-
yl)propan-2-yl)carbamate 19 Quinuclidin-3-yl (2-(2-(4-(3-
methoxypropoxy)phenyl)thiazol-4-yl)propan-2-yl)carbamate 20
Quinuclidin-3-yl (2-(2-(4-(2-methoxyethoxy)phenyl)thiazol-
4-yl)propan-2-yl)carbamate 21 Quinuclidin-3-yl
2-(5-(4-(2-methoxyethoxy)phenyl)pyridin- 2-yl)propan-2-ylcarbamate
22 Quinuclidin-3-yl (2-(4'-(3-cyanopropoxy)-[1,1'-biphenyl]-4-
yl)propan-2-yl)carbamate 23 Quinuclidin-3-yl
(2-(4'-(cyanomethoxy)-[1,1'-biphenyl]-4-
yl)propan-2-yl)carbamate
[0177] 1.64 Compound 1, or any of 1.1-1.63, wherein the compound is
selected from quinuclidin-3-yl
(2-(4'-fluoro-[1,1'-biphenyl]-3-yl)propan-2-yl)carbamate,
(S)-quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate, and
(S)-quinuclidin-3-yl
(2-(4'-(2-methoxyethoxy)-[1,1'-biphenyl]-4-yl)propan-2-yl)carbamate;
[0178] 1.65 Compound 1, or any of 1.1-1.63, wherein the compound is
quinuclidin-3-yl
(2-(4'-fluoro-[1,1'-biphenyl]-3-yl)propan-2-yl)carbamate; [0179]
1.66 Compound 1 or any of 1.1-1.63, wherein the compound is
quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate, e.g.,
(S)-quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate; [0180]
1.67 Compound 1, or any of 1.1-1.66, wherein the Compound of
Formula I, or any of II to XII, is in free base form; [0181] 1.68
Compound 1, or any of 1.1-1.66, wherein the Compound of Formula I,
or any of II to XII, is in pharmaceutically acceptable salt form;
[0182] 1.69 Compound 1.68, wherein said salt form is an acid
addition salt form; [0183] 1.70 Compound 1.69, wherein said acid
addition salt form is a salt selected from the hydrochloride,
hydrobromide, hydroiodide, nitrate, sulfate, bisulfate, phosphate,
acid phosphate, acetate, lactate, citrate, acid citrate, tartrate,
bitartrate, succinate, hydroxysuccincate, malate, maleate,
fumarate, gluconate, saccharate, benzoate, methanesulfonate, and
pamoate; [0184] 1.71 Compound 1.70, wherein the acid addition salt
form is selected from hydrochloride, hydroxysuccinate (e.g.,
2-hydroxysuccinate), and malate; [0185] 1.72 Compound 1.68, wherein
said salt form is a base addition salt form; [0186] 1.73 Compound
1, or any of 1.1-1.72, wherein the compound is (S)-quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate in malate
salt form; [0187] 1.74 Compound 1, or any of 1.1-1.73, wherein the
Compound of Formula I, or any of II to XII, is in the form of a
prodrug, as described herein; [0188] 1.75 Compound 1, or any of
1.1-1.74, wherein the Compound of Formula I, or any of II to XII,
is in the form of a hydrate, solvate and/or polymorph.
[0189] Salts
[0190] Presently disclosed compounds, e.g., any of Compounds 1 or
1.1-1.75, that are basic in nature are generally capable of forming
a wide variety of different salts with various inorganic and/or
organic acids. Although such salts are generally pharmaceutically
acceptable for administration to animals and humans, it is often
desirable in practice to initially isolate a compound from the
reaction mixture as a pharmaceutically unacceptable salt and then
simply convert the latter back to the free base compound by
treatment with an alkaline reagent, and subsequently convert the
free base to a pharmaceutically acceptable acid addition salt. The
acid addition salts of the base compounds can be readily prepared
using conventional techniques, e.g. by treating the base compound
with a substantially equivalent amount of the chosen mineral or
organic acid in an aqueous solvent medium or in a suitable organic
solvent such as, for example, methanol or ethanol. Upon careful
evaporation of the solvent, the desired solid salt is obtained.
Presently disclosed compounds that are positively charged, e.g.
containing a quaternary ammonium, may also form salts with the
anionic component of various inorganic and/or organic acids.
[0191] Acids which can be used to prepare pharmaceutically
acceptable salts of quinuclidine compounds are those which can form
non-toxic acid addition salts, e.g. salts containing
pharmacologically acceptable anions, such as chloride, bromide,
iodide, nitrate, sulfate or bisulfate, phosphate or acid phosphate,
acetate, lactate, citrate or acid citrate, tartrate or bitartrate,
succinate, malate, maleate, fumarate, gluconate, saccharate,
benzoate, methanesulfonate and pamoate [i.e.
1,1'-methylene-bis-(2-hydroxy-3-naphthoate)] salts.
[0192] Presently disclosed compounds that are acidic in nature,
e.g. compounds containing a thiol moiety, are generally capable of
forming a wide variety of different salts with various inorganic
and/or organic bases. Although such salts are generally
pharmaceutically acceptable for administration to animals and
humans, it is often desirable in practice to initially isolate a
compound from the reaction mixture as a pharmaceutically
unacceptable salt and then simply convert the latter back to the
free acid compound by treatment with an acidic reagent, and
subsequently convert the free acid to a pharmaceutically acceptable
base addition salt. These base addition salts can be readily
prepared using conventional techniques, e.g. by treating the
corresponding acidic compounds with an aqueous solution containing
the desired pharmacologically acceptable cations, and then
evaporating the resulting solution to dryness, e.g. under reduced
pressure. Alternatively, they also can be prepared by mixing lower
alkanolic solutions of the acidic compounds and the desired alkali
metal alkoxide together, and then evaporating the resulting
solution to dryness in the same manner as before. In either case,
stoichiometric quantities of reagents may be employed in order to
ensure completeness of reaction and maximum product yields of the
desired solid salt.
[0193] Bases which can be used to prepare the pharmaceutically
acceptable base addition salts of quinuclidine compounds are those
which can form non-toxic base addition salts, e.g. salts containing
pharmacologically acceptable cations, such as, alkali metal cations
(e.g. potassium and sodium), alkaline earth metal cations (e.g.
calcium and magnesium), ammonium or other water-soluble amine
addition salts such as N-methylglucamine (meglumine), lower
alkanolammonium, and other such bases of organic amines.
[0194] In one embodiment, the pharmaceutically acceptable salt is a
succinate salt. In another embodiment, the pharmaceutically
acceptable salt is a 2-hydroxysuccinate salt, e.g. an
(S)-2-hydroxysuccinate salt. In another embodiment, the
pharmaceutically acceptable salt is a hydrochloride salt (i.e. a
salt with HCl). In another embodiment, the pharmaceutically
acceptable salt is a malate salt.
[0195] Prodrugs
[0196] The present disclosure further embraces prodrugs of the
compounds 1 and 1.1-1.75. The pharmaceutically acceptable prodrugs
disclosed herein are derivatives of quinuclidine compounds which
can be converted in vivo into the quinuclidine compounds described
herein. The prodrugs, which may themselves have some activity,
become pharmaceutically active in vivo when they undergo, for
example, solvolysis under physiological conditions or enzymatic
degradation. Methods for preparing prodrugs of compounds as
described herein would be apparent to one of skill in the art based
on the present disclosure.
[0197] In one embodiment, the carbamate moiety of the quinuclidine
compound is modified. For example, the carbamate moiety of the
quinuclidine compound may be modified by the addition of water
and/or one or two aliphatic alcohols. In this case, the
carbon-oxygen double bond of the carbamate moiety adopts what could
be considered a hemiacetal or acetal functionality. In one
embodiment, the carbamate moiety of the quinuclidine compound may
be modified by the addition of an aliphatic diol such as
1,2-ethanediol.
[0198] In one embodiment, one or more of the hydroxy, thio or amino
groups on the quinuclidine compound are modified. For example, one
or more of the hydroxy, thio and/or amino groups on the
quinuclidine compound may be modified to form acid derivatives,
e.g. esters, thioesters (or thiolesters) and/or amides. The acid
derivatives can be formed, for example, by reacting a quinuclidine
compound which comprises one or more hydroxy, thio or amino groups
with an acetylating agent. Examples of acetylating agents include
anhydrides such as acetic anhydride, acid chlorides such as benzyl
chloride, and dicarbonates such as di-tert-butyl dicarbonate.
[0199] Stereochemistry
[0200] The present disclosure further embraces stereoisomers and
mixture of stereoisomers of compounds 1 and 1.1-1.75. Stereoisomers
(e.g. cis and trans isomers) and all optical isomers of a presently
disclosed compound (e.g. R- and S-enantiomers), as well as racemic,
diastereomeric and other mixtures of such isomers are within the
scope of the present disclosure.
[0201] In one embodiment, the quinuclidin-3-yl group of a
quinuclidine compound as defined herein has the R-configuration.
Accordingly, the quinuclidine compound may be selected from the
group consisting of compounds of formulae (Ia) to (XIIa):
##STR00005## ##STR00006##
[0202] and the pharmaceutically acceptable salts and prodrugs
thereof.
[0203] In another embodiment, the quinuclidin-3-yl group of the
quinuclidine compound as defined herein has the S-configuration.
Accordingly, the quinuclidine compound may be selected from the
group consisting of compounds of formulae (Ib) to (XIIb):
##STR00007## ##STR00008##
[0204] and the pharmaceutically acceptable salts and prodrugs
thereof.
[0205] In one embodiment the quinuclidine compound is a compound of
formula (Xb) or a pharmaceutically acceptable salt or prodrug
thereof. In another embodiment the quinuclidine compound is a
compound of formula (XIIb) or a pharmaceutically acceptable salt or
prodrug thereof.
[0206] In one embodiment, the quinuclidin-3-yl group of the
quinuclidine compound as defined herein exists in a mixture of
isomers having the R- and S-configurations. For example, the
quinuclidine compound may be a mixture of compounds selected from
the group consisting of compounds of formulae (Ia) and (Ib), (IIa)
and (IIb), (IIIa) and (IIIb), (IVa) and (IVb), (Va) and (Vb), (VIa)
and (VIb), (VIIa) and (VIIb), (VIIIa) and (VIIIb)), (IXa) and
(IXb), (Xa) and (Xb), (XIa) and (XIb), and (XIIa) and (XIIb), and
the pharmaceutically acceptable salts and prodrugs thereof. In one
embodiment the quinuclidine compound is present as a racemic
mixture, e.g. the R- and S-isomers of the quinuclidin-3-yl group
are present in about equal amounts. In another embodiment the
quinuclidine compound is present as a mixture of isomers having the
R- and S-configurations, wherein the R- and S-isomers are present
in different amounts. In one embodiment the S-isomer is present in
an enantiomeric excess of at least about 5%, 10%, 25%, 40%, 70%,
80%, 90%, 95%, 97%, 98% or 99%, e.g. about 100%. In another
embodiment, the R-isomer is present in an enantiomeric excess of at
least about 5%, 10%, 25%, 40%, 70%, 80%, 90%, 95%, 97%, 98% or 99%,
e.g. about 100%.
[0207] Methods for preparing enantioenriched and/or enantiopure
quinuclidine compounds would be apparent to the person of skill in
the art based on the present disclosure.
[0208] The compounds presently disclosed can exist in several
tautomeric forms, including the enol and imine form, and the keto
and enamine form and geometric isomers and mixtures thereof.
Tautomers exist as mixtures of a tautomeric set in solution. In
solid form, usually one tautomer predominates. Even though one
tautomer may be described, all tautomers are within the scope of
the present disclosure.
[0209] Atropisomers are also within the scope of the present
disclosure. Atropisomers refer to compounds that can be separated
into rotationally restricted isomers.
[0210] Other Forms
[0211] The present disclosure further embraces hydrates, solvates
and polymorphs of Compound 1 and 1.1-1.75. Pharmaceutically
acceptable hydrates, solvates, and polymorphs, of the quinuclidine
compounds described herein are within the scope of the present
disclosure. Quinuclidine compounds as described herein may be in an
amorphous form and/or in one or more crystalline forms.
[0212] Isotopically-labeled compounds are also within the scope of
the present disclosure. As used herein, an "isotopically-labeled
compound" refers to a presently disclosed compound including
pharmaceutical salts and prodrugs thereof, each as described
herein, in which one or more atoms are replaced by an atom having
an atomic mass or mass number different from the atomic mass or
mass number usually found in nature. Examples of isotopes that can
be incorporated into compounds presently disclosed include isotopes
of hydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine and
chlorine, such as .sup.2H, .sup.3H, .sup.13C, .sup.14C, .sup.15N,
.sup.18O, .sup.17O, .sup.31P, .sup.32P, .sup.35S, .sup.18F, and
.sup.36Cl, respectively.
[0213] Medical Indications
[0214] The quinuclidine compounds, and pharmaceutical compositions
containing them, described herein are useful in therapy, in
particular in the therapeutic treatment of supranuclear gaze
palsies, including horizontal and vertical saccadic gaze palsies,
neurological deficits, including dementia and gait disorders in a
patient having a disease such as Gaucher disease. Subjects to be
treated according to the methods described herein include
vertebrates, such as mammals. In particular embodiments the mammal
is a human patient.
[0215] In a first aspect, the present invention provides a method
(Method 1) for treating or preventing supranuclear gaze palsies,
including horizontal and vertical saccadic gaze palsies in a
subject in need thereof, the method comprising administering to the
subject an effective amount of a quinuclidine compound as described
herein, e.g., a compound according to Formula I or any of II-XII,
Ia-XIIa or Ib-XIIb, or any of Compounds 1 or 1.1 to 1.75. Also
provided is a quinuclidine compound as described herein, e.g., a
compound according to Formula I or any of II-XII, Ia-XIIa or
Ib-XIIb, or any of Compounds 1 or 1.1 to 1.75, for use in a method
for treating or preventing supranuclear gaze palsies, including
horizontal and vertical saccadic gaze palsies in a subject in need
thereof, e.g., for use in Method 1 or any of 1.1-1.62. Further
provided is the use of a quinuclidine compound as described herein,
e.g., a compound according to Formula I or any of II-XII, Ia-XIIa
or Ib-XIIb, or any of Compounds 1 or 1.1 to 1.75, in the
manufacture of a medicament for use in a method of treating or
preventing supranuclear gaze palsies, including horizontal and
vertical saccadic gaze palsies in a subject in need thereof, e.g.,
in the manufacture of a medicament for use in Method 1 or any of
1.1-1.62.
[0216] In particular further embodiments of Method 1, the present
disclosure provides: [0217] 1.1 Method 1, wherein the method
comprises administering to the subject an effective amount of a
compound according to Formula I or any of II-XII, Ia-XIIa or
Ib-XIIb, or any of Compounds 1 or any of 1.1 to 1.75; [0218] 1.2
Method 1, wherein the method comprises administering to the subject
an effective amount of Compound 1 or any one or more of Compounds
1.1 to 1.75; [0219] 1.3 Method 1 or any of 1.1-1.2, wherein the
method comprises administering to the subject an effective amount
of a pharmaceutical composition comprising the compound according
to Formula I or any of II-XII, Ia-XIIa or Ib-XIIb, or any of
Compounds 1 or any of 1.1 to 1.75; [0220] 1.4 Method 1 or any of
1.1-1.2, wherein the method comprises administering to the subject
an effective amount of a pharmaceutical composition comprising the
Compound 1 or any one or more of Compounds 1.1 to 1.75; [0221] 1.5
Method 1.3 or 1.4, wherein the pharmaceutical composition further
comprises at least one pharmaceutically acceptable excipient, as
described herein; [0222] 1.6 Method 1 or any of 1.1-1.5, wherein
the method comprising administering a pharmaceutical dosage form
comprising an effective amount of the compound or an effective
amount of the pharmaceutical composition; [0223] 1.7 Method 1.6,
wherein the dosage form is an oral dosage form (e.g., a pill,
capsule, caplet, tablet, dragee, powder, granule, film, lozenge, or
liquid); [0224] 1.8 Method 1.7, wherein the dosage form is a
chewable tablet; [0225] 1.9 Method 1.6, wherein the dosage form is
a parenteral dosage form (e.g., wherein the pharmaceutical
composition is formulated for injection); [0226] 1.10 Method 1.9,
wherein the injection is intravenous, intramuscular, intrathecal or
subcutaneous injection, optionally a sterile injection; [0227] 1.11
Method 1.6, wherein the dosage form is a topical or rectal dosage
form; [0228] 1.12 Method 1.6, wherein the dosage form is an
intranasal dosage form (e.g., an aerosol); [0229] 1.13 Method 1 or
any of 1.1 to 1.12, wherein the method further comprises
concurrently administering a second active agent, e.g., a second
compound capable of treating or preventing supranuclear gaze
palsies in a patient in need thereof, as described herein; [0230]
1.14 Method 1.13, wherein the second active agent is administrated
in the same pharmaceutical composition or dosage form as the
quinuclidine compound; [0231] 1.15 Method 1.13 or 1.14, wherein the
second active agent is a GCS inhibitor (e.g., miglustat or
eliglustat); [0232] 1.16 Method 1, or any of 1.1-1.15, wherein the
subject is a mammalian animal; [0233] 1.17 Method 1.16, wherein the
subject is a primate animal; [0234] 1.18 Method 1.17, wherein the
subject is a human; [0235] 1.19 Method 1 or any of 1.1-1.18,
wherein the supranuclear gaze palsy is a conjugate gaze palsy;
[0236] 1.20 Method 1 or any of 1.1-1.19, wherein the supranuclear
gaze palsy comprises horizontal saccadic palsy, vertical saccadic
palsy, vestibulo-ocular reflex impairment, gaze-holding impairment,
deterioration of horizontal smooth pursuit and/or deterioration of
vertical smooth pursuit; [0237] 1.21 Method 1, or any of 1.1-1.20,
wherein the supranuclear gaze palsy comprises horizontal saccadic
gaze palsy; [0238] 1.22 Method 1, or any of 1.1-1.21, wherein the
supranuclear gaze palsy comprises vertical saccadic gaze palsy;
[0239] 1.23 Method 1, or any of 1.1-1.22, wherein the supranuclear
gaze palsy is a complete palsy (e.g., paralysis); [0240] 1.24
Method 1, or any of 1.1-1.23, wherein the subject has Gaucher
disease Type 3; [0241] 1.25 Method 1, or any of 1.1-1.24, wherein
the subject has Niemann-Pick disease Type C; [0242] 1.26 Method 1,
or any of 1.1-1.24, wherein the subject has a GM2-gangliosidosis
(e.g., Tay-Sachs disease, Sandhoff disease, or GM2 gangliosidosis
AB variant); [0243] 1.27 Method 1, or any of 1.1-1.24, wherein the
subject is diagnosed with a mutation in the gene GBA1; [0244] 1.28
Method 1, or any of 1.1-1.24, wherein the subject is diagnosed with
a mutation in the genes NPC1 and/or NPC2; [0245] 1.29 Method 1, or
any of 1.1-1.24, wherein the subject is diagnosed with a mutation
in the gene HEXA (encoding hexosaminidase A) and/or a mutation in
the gene HEXB (encoding hexosaminidase B) and/or a mutation in the
gene GM2A (encoding the GM2 ganglioside activator protein); [0246]
1.30 Method 1, or any of 1.1-1.29, wherein the subject is diagnosed
with Parkinson's disease; [0247] 1.31 Method 1, or any of 1.1-1.30,
wherein the subject undergoes concurrent treatment with enzyme
replacement therapy (ERT), e.g., using a glucocerebrosidase (e.g.,
imiglucerase, velaglucerase, or taliglucerase), optionally wherein
in each of such enzyme is a recombinant enzyme; [0248] 1.32 Method
1.31, wherein the subject undergoes concurrent treatment with one
or more of imiglucerase, velaglucerase (e.g., velaglucerase alfa),
and taliglucerase (e.g., taliglucerase alfa); [0249] 1.33 Method
1.32, wherein the subject under goes concurrent treatment with
imiglucerase; [0250] 1.34 Method 1.33, wherein the subject
undergoes concurrent treatment with imiglucerase at a dosage of
from 2.5 units/kg body weight to 80 units/kg body weight every 1 to
3 weeks, e.g., 40 to 60 units/kg body weight every 2 weeks (1 unit
of imiglucerase is the amount of enzyme that catalyzes the
hydrolysis of 1 micromole of the synthetic substrate
p-nitrophenyl-.beta.-D-glucopyranoside per minute at 37.degree.
C.); [0251] 1.35 Method 1.34, wherein the subject's dosage of
imiglucerase at each administration (e.g., every 1 to 3 weeks,
e.g., every 2 weeks) is administered as an intravenous (IV)
infusion over a period of 1-3 hours (e.g., 1-2 hours); [0252] 1.36
Method 1 or any of 1.1-1.35, wherein the subject has been
administered enzyme replacement therapy (e.g., imiglucerase,
velaglucerase and/or taliglucerase) prior to the initiation of
treatment with the compound according to Formula I (or any of
II-XII, Ia-XIIa or Ib-XIIb, or any of Compounds 1 or 1.1 to 1.75);
[0253] 1.37 Method 1.36, wherein the subject has been administered
imiglucerase therapy for at least 6 months prior to beginning
therapy with the compound according to Formula I (or any of II-XII,
Ia-XIIa or Ib-XIIb, or any of Compounds 1 or 1.1 to 1.75), for
example, at least 12 months (1 year), or at least 18 months, or at
least 2 years, or at least 3 years. [0254] 1.38 Method 1.36 or
1.37, wherein the subject has been administered imiglucerase
therapy for at least 6 months at a stable dose prior to beginning
therapy with the compound according to Formula I (or any of II-XII,
Ia-XIIa or Ib-XIIb, or any of Compounds 1 or 1.1 to 1.75); [0255]
1.39 Method 1 or any of 1.1-1.38, wherein the method further
comprises the step of transitioning the subject from ERT therapy
(e.g., imiglucerase, velaglucerase or taliglucerase) to treatment
with the compound according to Formula I (or any of II-XII, Ia-XIIa
or Ib-XIIb, or any of Compounds 1 or 1.1 to 1.75); [0256] 1.40
Method 1 or any of 1.1-1.39, wherein the subject has a hemoglobin
level of at least 11 g/dL for females and at least 12 g/dL for
males; [0257] 1.41 Method 1, or any of 1.1-1.40, wherein the
subject has a platelet count of at least 100,000/cubic millimeter;
[0258] 1.42 Method 1, or any of 1.1-1.41, wherein the subject has a
splenic volume of less than 10 multiples of normal (MN) and/or a
hepatic volume of less than 1.5 MN; [0259] 1.43 Method 1, or any of
1.1-1.42, wherein the subject is diagnosed with an oculomotor
apraxia, for example, an oculomotor apraxia characterized by
horizontal saccade abnormality; [0260] 1.44 Method 1, or any of
1.1-1.43, wherein the subject is at least 18 years of age (e.g.,
18-30 years of age) at the start of treatment with the compound
according to Formula I (or any of II-XII, Ia-XIIa or Ib-XIIb, or
any of Compounds 1 or 1.1 to 1.75); [0261] 1.45 Method 1, or any of
1.1-1.44, wherein the subject has a glucosylceramide (GL1)
concentration of 4.4-11.1 ng/mL in cerebrospinal fluid (CSF) and
4.9-8.3 .mu.g/mL in plasma; [0262] 1.46 Method 1, or any of
1.1-1.45, wherein the subject has a glucosylsphingosine (lyso-GL1)
concentration of 20.1-67.6 pg/mL in CSF and 8.8-159.0 ng/mL in
plasma; [0263] 1.47 Method 1, or any of 1.1-1.46, wherein the
subject is administered a daily dose of about 1 mg to about 150 mg
of the compound according to Formula I (or any of II-XII, Ia-XIIa
or Ib-XIIb, or any of Compounds 1 or 1.1 to 1.75), e.g., from 5 to
50 mg, or from 10 to 40 mg, or from 10 to 30 mg, or from 10 to 20
mg, or from 20 to 30 mg, or from 30 to 40 mg, or from 40 to 50 mg,
or from 5 to 25 mg, or from 20 to 50 mg, or from 5 to 15 mg, or
from 15 to 30 mg, or about 15 mg, or selected from 2, 5, 15, 25,
50, 100, or 150 mg; [0264] 1.48 Method 1, or any of 1.1-1.47,
wherein the subject is a human adult patient, e.g., of an age from
18 to 80 years old, e.g., from 18 to 60 years old, or from 18 to 40
years old, or from 18 to 30 years old, or from 18 to 25 years old;
[0265] 1.49 Method 1, or any of 1.1-1.47, wherein the subject is a
human pediatric patient, e.g., of an age from 0 to 18 years old,
e.g., from 1 to 15 years old, or from 1 to 5 years old, or from 5
to 10 years old, or from 10 to 15 years old, or from 10 to 18 years
old; [0266] 1.50 Method 1, or any of 1.1-1.49, wherein the method
is effective to stabilize the progression of the supranuclear gaze
palsy, e.g., for at least 6 months, or at least 9 months, or at
least 12 months; [0267] 1.51 Method 1, or any of 1.1-1.49, wherein
the method is effective to reverse the progression of the
supranuclear gaze palsy, e.g., for at least 6 months, or at least 9
months, or at least 12 months; [0268] 1.52 Method 1, or any of
1.1-1.51, wherein the method results in reduction in
glucosylceramide concentration in CSF and/or in plasma of at least
30% after 6 months of treatment, e.g., at least 40%, at least 50%,
at least 60% or at least 70%; [0269] 1.53 Method 1, or any of
1.1-1.52, wherein the method results in an increase in
glucosylsphingosine concentration in CSF and/or in plasma of at
least 30% after 6 months of treatment, e.g., at least 40%, at least
50%, at least 60% or at least 70%; [0270] 1.54 Method 1, or any of
1.1-1.53, wherein the method results in a statistically or
clinically unchanged Modified Severity Scoring Tool (mSST) value
for neurological disease after 6 months of treatment; [0271] 1.55
Method 1, or any of 1.1-1.54, wherein the compound according to
Formula I (or any of II-XII, Ia-XIIa or Ib-XIIb, or any of
Compounds 1 or 1.1 to 1.75), or pharmaceutically acceptable salt or
prodrug thereof, is administered by systemic administration, e.g.,
via a parenteral route or a non-parenteral route; [0272] 1.56
Method 1.55, wherein the route of administration is oral (enteral);
[0273] 1.57 Method 1.55, wherein the route of administration is
parenteral, e.g., by injection, such as, by intravenous injection;
[0274] 1.58 Method 1, or any of 1.1-1.57, wherein the compound
according to Formula I (or any of II-XII, Ia-XIIa or Ib-XIIb, or
any of Compounds 1 or 1.1 to 1.75), or pharmaceutically acceptable
salt or prodrug thereof, is administered by local administration,
e.g., by topical administration; [0275] 1.59 Method 1, or any of
1.1-1.58, wherein the compound is (S)-quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate or
quinuclidin-3-yl
(2-(4'-fluoro-[1,1'-biphenyl]-3-yl)propan-2-yl)carbamate; [0276]
1.60 Method 1.59, wherein the dosage of the compound is 15 mg/day
orally administered; [0277] 1.61 Method 1.60, wherein the dosage of
the compound is 15 mg/day in a single oral dose; [0278] 1.62 Method
1, or any of 1.1-1.61, wherein the subject is administered a single
daily dose of 5 mg, 10 mg, 15 mg, or 20 mg of the compound, e.g.,
of (S)-quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate,
optionally in malate salt acid addition salt form.
[0279] Disease and disorders such as those giving rise to
supranuclear gaze palsies are often associated with one or more
genetic mutations. In some embodiments of the present disclosure, a
subject or subject is diagnosed with having a particular disease or
disorder and is also diagnosed to have a particular genetic
mutation, for example, one that is known to be a cause of the
disease or disorder in question, although it often cannot be proven
that a particular patient's disease or disorder is caused by the
particular mutation that a person has been diagnosed with having.
As used in this manner, the term "diagnosed to have a particular
genetic mutation" means that a subject or patient has been tested,
e.g., by DNA or RNA sequencing, protein profiling, or other
suitable means, and found to have the mutation in question.
However, as discussed further below, many genetic diseases and
disorders can have multiple genetic causes (e.g., mutations), and
patients may have multiple mutations each of which may, under some
circumstances, be sufficient to cause the disease or disorder,
without it being subject to proof that a particular mutation causes
a particular disease or disorder in a particular patient.
[0280] The methods according to Method 1 et seq. may be beneficial
for subjects who have been diagnosed with a lysosomal storage
disease, such as Gaucher Type 3 or Niemann-Pick Type C, but who are
not yet experiencing the ocular symptoms associated with the
disease state. The methods according to Method 1 et seq. may also
be beneficial for subjects who are at risk of developing a
lysosomal storage disease, such as Gaucher Type 3 or Niemann-Pick
Type C, due to, for example, a mutation in the subject or the
subject's family lineage known to cause such disease. Therefore, in
some embodiments of the methods described herein, the subject has
been diagnosed as being at risk of developing said disease or
disorder, and the method prevents or delays the onset and/or
development of the ocular symptoms of the disease or disorder
(e.g., the supranuclear gaze palsy) in the subject. In some
embodiments, the subject has been diagnosed as being at risk of
developing said disease or disorder by virtue of having a mutation
in a gene as described herein.
[0281] In a second aspect, the present invention provides a method
(Method 4) for treating or preventing cognitive dysfunction and/or
gait abnormalities, including ataxia, associated with a lysosome
storage disease, in a subject in need thereof, the method
comprising administering to the subject an effective amount of a
quinuclidine compound as described herein, e.g., a compound
according to Formula I or any of II-XII, Ia-XIIa or Ib-XIIb, or any
of Compounds 1 or 1.1 to 1.75. Also provided is a quinuclidine
compound as described herein, e.g., a compound according to Formula
I or any of II-XII, Ia-XIIa or Ib-XIIb, or any of Compounds 1 or
1.1 to 1.75, for use in a method for treating or preventing
cognitive dysfunction and/or gait abnormalities, including ataxia,
associated with a lysosome storage disease, in a subject in need
thereof, e.g., for use in Method 4 or any of 4.1-4.62. Further
provided is the use of a quinuclidine compound as described herein,
e.g., a compound according to Formula I or any of II-XII, Ia-XIIa
or Ib-XIIb, or any of Compounds 1 or 1.1 to 1.75, in the
manufacture of a medicament for use in a method of treating or
preventing cognitive dysfunction and/or gait abnormalities,
including ataxia, associated with a lysosome storage disease, in a
subject in need thereof, e.g., in the manufacture of a medicament
for use in Method 4 or any of 4.1-4.62.
[0282] In particular further embodiments of Method 4, the present
disclosure provides: [0283] 4.1.Method 4, wherein the method
comprises administering to the subject an effective amount of a
compound according to Formula I or any of II-XII, Ia-XIIa or
Ib-XIIb, or any of Compounds 1 or any of 1.1 to 1.75; [0284]
4.2.Method 4, wherein the method comprises administering to the
subject an effective amount of Compound 1 or any one or more of
Compounds 1.1 to 1.75; [0285] 4.3.Method 4 or any of 4.1-4.2,
wherein the method comprises administering to the subject an
effective amount of a pharmaceutical composition comprising the
compound according to Formula I or any of II-XII, Ia-XIIa or
Ib-XIIb, or any of Compounds 1 or any of 1.1 to 1.75; [0286]
4.4.Method 4 or any of 4.1-4.2, wherein the method comprises
administering to the subject an effective amount of a
pharmaceutical composition comprising the Compound 1 or any one or
more of Compounds 1.1 to 1.75; [0287] 4.5.Method 4.3 or 4.4,
wherein the pharmaceutical composition further comprises at least
one pharmaceutically acceptable excipient, as described herein;
[0288] 4.6.Method 4 or any of 4.1-4.5, wherein the method
comprising administering a pharmaceutical dosage form comprising an
effective amount of the compound or an effective amount of the
pharmaceutical composition; [0289] 4.7.Method 4.6, wherein the
dosage form is an oral dosage form (e.g., a pill, capsule, caplet,
tablet, dragee, powder, granule, film, lozenge, or liquid); [0290]
4.8.Method 4.7, wherein the dosage form is a chewable tablet;
[0291] 4.9.Method 4.6, wherein the dosage form is a parenteral
dosage form (e.g., wherein the pharmaceutical composition is
formulated for injection); [0292] 4.10. Method 4.9, wherein the
injection is intravenous, intramuscular, intrathecal or
subcutaneous injection, optionally a sterile injection; [0293]
4.11. Method 4.6, wherein the dosage form is a topical or rectal
dosage form; [0294] 4.12. Method 4.6, wherein the dosage form is an
intranasal dosage form (e.g., an aerosol); [0295] 4.13. Method 4 or
any of 4.1 to 4.12, wherein the method further comprises
concurrently administering a second active agent, e.g., a second
compound capable of treating or preventing cognitive dysfunction
and/or gait abnormalities in a patient in need thereof, as
described herein; [0296] 4.14. Method 4.13, wherein the second
active agent is administrated in the same pharmaceutical
composition or dosage form as the quinuclidine compound; [0297]
4.15. Method 4.13 or 4.14, wherein the second active agent is a GCS
inhibitor (e.g., miglustat or eliglustat); [0298] 4.16. Method 4,
or any of 4.1-4.15, wherein the subject is a mammalian animal;
[0299] 4.17. Method 4.16, wherein the subject is a primate animal;
[0300] 4.18. Method 4.17, wherein the subject is a human; [0301]
4.19. Method 4 or any of 4.1-4.18, wherein the ataxia is a
cerebellar ataxia; [0302] 4.20. Method 4.19, wherein the ataxia
shows symptoms selected from gait instability, asthenia, asynergy,
delayed reaction time, dyschronometria, dysarthria, dysphagia,
hypotonia, dysmetria, hypometria, hypermetria, dysdiadochokinesia,
speech slurring, voice tremor, ataxic respiration, postural
instability, and combinations thereof, for example, wherein the
primary ataxic deficit is a gait instability; [0303] 4.21. Method
4.19 or 4.20, wherein the subject has a baseline ataxia of at least
0.5 on the Scale for Assessment and Rating of Ataxia (SARA) scale
at the initiation of therapy according to the method, e.g., a
baseline SARA score of at least 1, or at least 2, or at least 3, or
at least 4, or at least 5, or at least 10 or at least 20; [0304]
4.22. Method 4, or any of 4.1-4.21, wherein the cognitive
dysfunction is a dementia; [0305] 4.23. Method 4.22, wherein the
dementia shows signs of defects in visual search speed, scanning
speed of processing, mental flexibility and/or executive
functioning, e.g., as evidence by a TMT-A of greater than 30
seconds, or greater than 45 seconds, or greater than 60 seconds
and/or a TMT-B of greater than 70 seconds, or greater than 90
seconds, or greater than 120 seconds, or greater than 150 seconds,
or greater than 180 seconds and/or wherein TMT-B minus TMT-A is
greater than 40 seconds, or greater than 60 seconds, or greater
than 90 seconds, or greater than 120 seconds; [0306] 4.24. Method
4, or any of 4.4-4.23, wherein the subject has Gaucher disease Type
3; [0307] 4.25. Method 4, or any of 4.1-4.24, wherein the subject
has Niemann-Pick disease Type C; [0308] 4.26. Method 4, or any of
4.1-4.24, wherein the subject has a GM2-gangliosidosis (e.g.,
Tay-Sachs disease, Sandhoff disease, or GM2 gangliosidosis AB
variant); [0309] 4.27. Method 4, or any of 4.1-4.24, wherein the
subject is diagnosed with a mutation in the gene GBA1; [0310] 4.28.
Method 4, or any of 4.1-4.24, wherein the subject is diagnosed with
a mutation in the genes NPC1 and/or NPC2; [0311] 4.29. Method 4, or
any of 4.1-4.24, wherein the subject is diagnosed with a mutation
in the gene HEXA (encoding hexosaminidase A) and/or a mutation in
the gene HEXB (encoding hexosaminidase B) and/or a mutation in the
gene GM2A (encoding the GM2 ganglioside activator protein); [0312]
4.30. Method 4, or any of 4.1-4.29, wherein the subject is
diagnosed with Parkinson's disease; [0313] 4.31. Method 4, or any
of 4.1-4.30, wherein the subject undergoes concurrent treatment
with enzyme replacement therapy (ERT), e.g., using a
glucocerebrosidase (e.g., imiglucerase, velaglucerase, or
taliglucerase), optionally wherein in each of such enzyme is a
recombinant enzyme; [0314] 4.32. Method 4.31, wherein the subject
undergoes concurrent treatment with one or more of imiglucerase,
velaglucerase (e.g., velaglucerase alfa), and taliglucerase (e.g.,
taliglucerase alfa); [0315] 4.33. Method 4.32, wherein the subject
under goes concurrent treatment with imiglucerase; [0316] 4.34.
Method 4.33, wherein the subject undergoes concurrent treatment
with imiglucerase at a dosage of from 2.5 units/kg body weight to
80 units/kg body weight every 1 to 3 weeks, e.g., 40 to 60 units/kg
body weight every 2 weeks (1 unit of imiglucerase is the amount of
enzyme that catalyzes the hydrolysis of 1 micromole of the
synthetic substrate p-nitrophenyl-.beta.-D-glucopyranoside per
minute at 37.degree. C.); [0317] 4.35. Method 4.34, wherein the
subject's dosage of imiglucerase at each administration (e.g.,
every 1 to 3 weeks, e.g., every 2 weeks) is administered as an
intravenous (IV) infusion over a period of 1-3 hours (e.g., 1-2
hours); [0318] 4.36. Method 4 or any of 4.1-4.35, wherein the
subject has been administered enzyme replacement therapy (e.g.,
imiglucerase, velaglucerase and/or taliglucerase) prior to the
initiation of treatment with the compound according to Formula I
(or any of II-XII, Ia-XIIa or Ib-XIIb, or any of Compounds 1 or 1.1
to 1.75); [0319] 4.37. Method 4.36, wherein the subject has been
administered imiglucerase therapy for at least 6 months prior to
beginning therapy with the compound according to Formula I (or any
of II-XII, Ia-XIIa or Ib-XIIb, or any of Compounds 1 or 1.1 to
1.75), for example, at least 12 months (1 year), or at least 18
months, or at least 2 years, or at least 3 years. [0320] 4.38.
Method 4.36 or 4.37, wherein the subject has been administered
imiglucerase therapy for at least 6 months at a stable dose prior
to beginning therapy with the compound according to Formula I (or
any of II-XII, Ia-XIIa or Ib-XIIb, or any of Compounds 1 or 1.1 to
1.75); [0321] 4.39. Method 4 or any of 4.1-4.38, wherein the method
further comprises the step of transitioning the subject from ERT
therapy (e.g., imiglucerase, velaglucerase or taliglucerase) to
treatment with the compound according to Formula I (or any of
II-XII, Ia-XIIa or Ib-XIIb, or any of Compounds 1 or 1.1 to 1.75);
[0322] 4.40. Method 4 or any of 4.1-4.39, wherein the subject has a
hemoglobin level of at least 11 g/dL for females and at least 12
g/dL for males; [0323] 4.41. Method 4, or any of 4.1-4.40, wherein
the subject has a platelet count of at least 100,000/cubic
millimeter; [0324] 4.42. Method 4, or any of 4.1-4.41, wherein the
subject has a splenic volume of less than 10 multiples of normal
(MN) and/or a hepatic volume of less than 1.5 MN; [0325] 4.43.
Method 4, or any of 4.1-4.42, wherein the subject is diagnosed with
a concurrent dementia, e.g., Alzheimer's disease or Parkinson's
disease; [0326] 4.44. Method 4, or any of 4.1-4.43, wherein the
subject is at least 18 years of age (e.g., 18-30 years of age) at
the start of treatment with the compound according to Formula I (or
any of II-XII, Ia-XIIa or Ib-XIIb, or any of Compounds 1 or 1.1 to
1.75); [0327] 4.45. Method 4, or any of 4.1-4.44, wherein the
subject has a glucosylceramide (GL1) concentration of 4.4-11.1
ng/mL in cerebrospinal fluid (CSF) and 4.9-8.3 .mu.g/mL in plasma;
[0328] 4.46. Method 4, or any of 4.1-4.45, wherein the subject has
a glucosylsphingosine (lyso-GL1) concentration of 20.1-67.6 pg/mL
in CSF and 8.8-159.0 ng/mL in plasma; [0329] 4.47. Method 4, or any
of 4.1-4.46, wherein the subject is administered a daily dose of
about 1 mg to about 150 mg of the compound according to Formula I
(or any of II-XII, Ia-XIIa or Ib-XIIb, or any of Compounds 1 or 1.1
to 1.75), e.g., from 5 to 50 mg, or from 10 to 40 mg, or from 10 to
30 mg, or from 10 to 20 mg, or from 20 to 30 mg, or from 30 to 40
mg, or from 40 to 50 mg, or from 5 to 25 mg, or from 20 to 50 mg,
or from 5 to 15 mg, or from 15 to 30 mg, or about 15 mg, or
selected from 2, 5, 15, 25, 50, 100, or 150 mg; [0330] 4.48. Method
4, or any of 4.1-4.47, wherein the subject is a human adult
patient, e.g., of an age from 18 to 80 years old, e.g., from 18 to
60 years old, or from 18 to 40 years old, or from 18 to 30 years
old, or from 18 to 25 years old; [0331] 4.49. Method 4, or any of
4.1-4.47, wherein the subject is a human pediatric patient, e.g.,
of an age from 0 to 18 years old, e.g., from 1 to 15 years old, or
from 1 to 5 years old, or from 5 to 10 years old, or from 10 to 15
years old, or from 10 to 18 years old; [0332] 4.50. Method 4, or
any of 4.1-4.49, wherein the method is effective to provide a
reduction on the SARA ataxia scale of at least 0.5, e.g., a SARA
score reduction of at least 1, or at least 2, or at least 3, or at
least 5, or at least 10; or wherein the method is effective to
reduce the SARA score to between 0.00 and 3.00, or between 0.00 and
2.00, or between 0.00 and 1.50, or between 0.00 and 1.00, or
between 0.00 and 0.50. [0333] 4.51. Method 4, or any of 4.1-4.50,
wherein the method is effective to improve cognitive ability or
reduce cognitive deficits, e.g., as measured by a reduction in the
time taken to complete the trail-making test (TMT), TMT-A and/or
TMT-B, a reduction in the difference between TMT-A time and TMT-B
time (TMT-A-TMT-B), for example, a reduction of at least 10%, or at
least 20%, or at least 30%, or at least 40%, or at least 50% (e.g.,
wherein TMT-A decreases by 5-20%, and/or TMT-B decreases by 25-30%,
and/or [TMT-A-TMT-B] decreases by 25-30%); [0334] 4.52. Method 4,
or any of 4.1-4.51, wherein the method results in reduction in
glucosylceramide concentration in CSF and/or in plasma of at least
30% after 6 months of treatment, e.g., at least 40%, at least 50%,
at least 60% or at least 70%; [0335] 4.53. Method 4, or any of
4.1-4.52, wherein the method results in an increase in
glucosylsphingosine concentration in CSF and/or in plasma of at
least 30% after 6 months of treatment, e.g., at least 40%, at least
50%, at least 60% or at least 70%; [0336] 4.54. Method 4, or any of
4.1-4.53, wherein the method results in a statistically or
clinically unchanged Modified Severity Scoring Tool (mSST) value
for neurological disease after 6 months of treatment; [0337] 4.55.
Method 4, or any of 4.1-4.54, wherein the compound according to
Formula I (or any of II-XII, Ia-XIIa or Ib-XIIb, or any of
Compounds 1 or 1.1 to 1.75), or pharmaceutically acceptable salt or
prodrug thereof, is administered by systemic administration, e.g.,
via a parenteral route or a non-parenteral route; [0338] 4.56.
Method 4.55, wherein the route of administration is oral (enteral);
[0339] 4.57. Method 4.55, wherein the route of administration is
parenteral, e.g., by injection, such as, by intravenous injection;
[0340] 4.58. Method 4, or any of 4.1-4.57, wherein the compound
according to Formula I (or any of II-XII, Ia-XIIa or Ib-XIIb, or
any of Compounds 1 or 1.1 to 1.75), or pharmaceutically acceptable
salt or prodrug thereof, is administered by local administration,
e.g., by topical administration; [0341] 4.59. Method 4, or any of
4.1-4.58, wherein the compound is (S)-quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate or
quinuclidin-3-yl
(2-(4'-fluoro-[1,1'-biphenyl]-3-yl)propan-2-yl)carbamate; [0342]
4.60. Method 4.59, wherein the dosage of the compound is 15 mg/day
orally administered; [0343] 4.61. Method 4.60, wherein the dosage
of the compound is 15 mg/day in a single oral dose; [0344] 4.62.
Method 4, or any of 4.1-4.61, wherein the subject is administered a
single daily dose of 5 mg, 10 mg, 15 mg, or 20 mg of the compound,
e.g., of (S)-quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate,
optionally in malate salt acid addition salt form.
[0345] In some embodiments of the present disclosure, a subject or
subject is diagnosed with having a particular disease or disorder
and is also diagnosed to have a particular genetic mutation, for
example, one that is known to be a cause of the disease or disorder
in question, although it often cannot be proven that a particular
patient's disease or disorder is caused by the particular mutation
that a person has been diagnosed with having. As used in this
manner, the term "diagnosed to have a particular genetic mutation"
means that a subject or patient has been tested, e.g., by DNA or
RNA sequencing, protein profiling, or other suitable means, and
found to have the mutation in question. However, as discussed
further below, many genetic diseases and disorders can have
multiple genetic causes (e.g., mutations), and patients may have
multiple mutations each of which may, under some circumstances, be
sufficient to cause the disease or disorder, without it being
subject to proof that a particular mutation causes a particular
disease or disorder in a particular patient.
[0346] The methods according to Method 4 et seq. may be beneficial
for subjects who have been diagnosed with a lysosomal storage
disease, such as Gaucher Type 3 or Niemann-Pick Type C, but who are
not yet experiencing the cognitive and/or ataxic symptoms
associated with the disease state. The methods according to Method
4 et seq. may also be beneficial for subjects who are at risk of
developing a lysosomal storage disease, such as Gaucher Type 3 or
Niemann-Pick Type C, due to, for example, a mutation in the subject
or the subject's family lineage known to cause such disease.
Therefore, in some embodiments of the methods described herein, the
subject has been diagnosed as being at risk of developing said
disease or disorder, and the method prevents or delays the onset
and/or development of the cognitive and/or ataxic symptoms of the
disease or disorder (e.g., the supranuclear gaze palsy) in the
subject. In some embodiments, the subject has been diagnosed as
being at risk of developing said disease or disorder by virtue of
having a mutation in a gene as described herein.
[0347] Pharmaceutical Compositions
[0348] The present disclosure also provides pharmaceutical
compositions comprising at least one quinuclidine compound as
described herein and at least one pharmaceutically acceptable
excipient, e.g. for use according to the methods disclosed herein.
The pharmaceutically acceptable excipient can be any such excipient
known in the art including those described in, for example,
Remington's Pharmaceutical Sciences, Mack Publishing Co. (A. R.
Gennaro edit. 1985). Pharmaceutical compositions of the compounds
presently disclosed may be prepared by conventional means known in
the art including, for example, mixing at least one presently
disclosed compound with a pharmaceutically acceptable
excipient.
[0349] Thus, in one aspect the present disclosure provides a
pharmaceutical dosage form comprising a quinuclidine compound as
described herein and a pharmaceutically acceptable excipient,
wherein the dosage form is formulated to provide, when administered
(e.g. when administered orally), an amount of said compound
sufficient to treat a disease or disorder described herein (e.g.,
in any of Method 1 et seq., or Method 4 et seq.).
[0350] A pharmaceutical composition or dosage form of the invention
can include an agent and another carrier, e.g. compound or
composition, inert or active, such as a detectable agent, label,
adjuvant, diluent, binder, stabilizer, buffers, salts, lipophilic
solvents, preservative, adjuvant or the like. Carriers also include
pharmaceutical excipients and additives, for example, proteins,
peptides, amino acids, lipids, and carbohydrates (e.g. sugars,
including monosaccharides, di-, tri-, tetra-, and oligosaccharides;
derivatized sugars such as alditols, aldonic acids, esterified
sugars and the like; and polysaccharides or sugar polymers), which
can be present singly or in combination, comprising alone or in
combination 1 to 99.99% by weight or volume. Exemplary protein
excipients include serum albumin such as human serum albumin (HSA),
recombinant human albumin (rHA), gelatin, casein, and the like.
Representative amino acid/antibody components, which can also
function in a buffering capacity, include alanine, glycine,
arginine, betaine, histidine, glutamic acid, aspartic acid,
cysteine, lysine, leucine, isoleucine, valine, methionine,
phenylalanine, aspartame, and the like. Carbohydrate excipients are
also intended within the scope of this invention, examples of which
include but are not limited to monosaccharides such as fructose,
maltose, galactose, glucose, D-mannose, sorbose, and the like;
disaccharides, such as lactose, sucrose, trehalose, cellobiose, and
the like; polysaccharides, such as raffinose, melezitose,
maltodextrins, dextrans, starches, and the like; and alditols, such
as mannitol, xylitol, maltitol, lactitol, xylitol sorbitol
(glucitol) and myoinositol.
[0351] Carriers which may be used include a buffer or a pH
adjusting agent; typically, the buffer is a salt prepared from an
organic acid or base. Representative buffers include organic acid
salts such as salts of citric acid, ascorbic acid, gluconic acid,
carbonic acid, tartaric acid, succinic acid, acetic acid, or
phthalic acid; Tris, tromethamine hydrochloride, or phosphate
buffers. Additional carriers include polymeric excipients/additives
such as polyvinylpyrrolidones, ficolls (a polymeric sugar),
dextrates (e.g. cyclodextrins, such as
2-hydroxypropyl-.beta.-cyclodextrin), polyethylene glycols,
flavoring agents, antimicrobial agents, sweeteners, antioxidants,
antistatic agents, surfactants (e.g. polysorbates such as "TWEEN
20" and "TWEEN 80"), lipids (e.g. phospholipids, fatty acids),
steroids (e.g. cholesterol), and chelating agents (e.g. EDTA).
[0352] The present disclosure also provides pharmaceutical
compositions, and kits comprising said compositions, which contain
at least one quinuclidine compound as described herein and at least
one further pharmaceutically-active agent. These pharmaceutical
compositions and kits may be adapted to allow simultaneous,
subsequent and/or separate administration of the quinuclidine
compound and the further active agent. For example, the
quinuclidine compound and the further active agent may be
formulated in separate dosage forms, e.g. in separate tablets,
capsules, lyophilizates or liquids, or they may be formulated in
the same dosage form, e.g. in the same tablet, capsule,
lyophilizate or liquid. Where the quinuclidine compound and the
further active agent are formulated in the same dosage form, the
quinuclidine compound and the further active agent may be present
substantially in admixture, e.g. within the core of a tablet, or
they may be present substantially in discrete regions of the dosage
form, e.g. in separate layers of the same tablet. In one
embodiment, the pharmaceutical dosage form comprises a further
agent which is capable of treating or preventing a supranuclear
gaze palsy, e.g., in a patient having, diagnosed with or
predisposed to a lysosomal storage disease, such as Gaucher Type 3
or Niemann-Pick Type C, or pain, e.g., in a patient having,
diagnosed with or predisposed to a lysosomal storage disease, such
as Fabry disease, as described herein.
[0353] In a further aspect the present disclosure provides a
pharmaceutical composition comprising: (i) a quinuclidine compound
as described herein; (ii) a further active agent; and (iii) a
pharmaceutically acceptable excipient. In one embodiment, the
further active agent is an agent which is capable of treating or
preventing a supranuclear gaze palsy, a gait disorder or a
cognitive dysfunction (e.g., dementia), e.g., in a patient having,
diagnosed with or predisposed to a lysosomal storage disease, such
as Gaucher Type 3 or Niemann-Pick Type C, as described herein.
[0354] The presently disclosed quinuclidine compounds and
pharmaceutical compositions can be used in an animal or human.
Thus, a presently disclosed compound can be formulated as a
pharmaceutical composition for oral, buccal, parenteral (e.g.
intravenous, intramuscular or subcutaneous), topical, rectal or
intranasal administration or in a form suitable for administration
by inhalation or insufflation. In particular embodiments, the
quinuclidine compound or pharmaceutical composition is formulated
for systemic administration, e.g. via a non-parenteral route. In
one embodiment, the quinuclidine compound or pharmaceutical
composition is formulated for oral administration, e.g. in solid
form. Such modes of administration and the methods for preparing
appropriate pharmaceutical compositions are described, for example,
in Gibaldi's Drug Delivery Systems in Pharmaceutical Care (1st ed.,
American Society of Health-System Pharmacists 2007).
[0355] The pharmaceutical compositions can be formulated so as to
provide slow, extended, or controlled release of the active
ingredient therein using, for example, hydroxypropyl methyl
cellulose in varying proportions to provide the desired release
profile, other polymer matrices, liposomes and/or microspheres. The
pharmaceutical compositions can also optionally contain opacifying
agents and may be of a composition that releases the active
ingredient(s) only, or preferentially, in a certain portion of the
gastrointestinal tract, optionally, in a delayed manner, e.g. by
using an enteric coating. Examples of embedding compositions
include polymeric substances and waxes. The active ingredient can
also be in micro-encapsulated form, if appropriate, with one or
more pharmaceutically acceptable carriers, excipients, or diluents
well known in the art (see, e.g., Remington's). The compounds
presently disclosed may be formulated for sustained delivery
according to methods well known to those of ordinary skill in the
art. Examples of such formulations can be found in U.S. Pat. Nos.
3,119,742; 3,492,397; 3,538,214; 4,060,598; and 4,173,626.
[0356] In solid dosage forms for oral administration (e.g.
capsules, tablets, pills, dragees, powders, granules and the like),
the active ingredient is mixed with one or more pharmaceutically
acceptable carriers, excipients, or diluents, such as sodium
citrate or dicalcium phosphate, and/or any of the following: (1)
fillers or extenders, such as starches, lactose, sucrose, glucose,
mannitol, microcrystalline cellulose, calcium phosphate and/or
silicic acid; (2) binders, such as, for example,
carboxymethylcellulose, alginates, gelatin, pregelatinized maize
starch, polyvinyl pyrrolidone, hydroxypropyl methylcellulose,
sucrose and/or acacia; (3) humectants, such as glycerol; (4)
disintegrating agents, such as agar-agar, calcium carbonate, sodium
starch glycolate, potato or tapioca starch, alginic acid, certain
silicates, and sodium carbonate; (5) solution retarding agents,
such as paraffin; (6) absorption accelerators, such as quaternary
ammonium compounds; (7) wetting agents, such as, for example,
sodium lauryl sulphate, acetyl alcohol and glycerol monostearate;
(8) absorbents, such as kaolin and bentonite clay; (9) lubricants,
such as talc, silica, calcium stearate, magnesium stearate, solid
polyethylene glycols, sodium lauryl sulfate, and mixtures thereof;
and (10) coloring agents. In the case of capsules, tablets, and
pills, the pharmaceutical compositions can also comprise buffering
agents. Solid compositions of a similar type can also be prepared
using fillers in soft and hard-filled gelatin capsules, and
excipients such as lactose or milk sugars, as well as high
molecular weight polyethylene glycols and the like.
[0357] A tablet can be made by compression or molding, optionally
with one or more accessory ingredients. Compressed tablets can be
prepared using binders (for example, gelatin or hydroxypropyl
methyl cellulose), lubricants, inert diluents, preservatives,
disintegrants (for example, sodium starch glycolate or cross-linked
sodium carboxymethyl cellulose), surface-actives, and/or dispersing
agents. Molded tablets can be made by molding in a suitable machine
a mixture of the powdered active ingredient moistened with an inert
liquid diluent. The tablets and other solid dosage forms, such as
dragees, capsules, pills, and granules, can optionally be scored or
prepared with coatings and shells, such as enteric coatings and
other coatings well known in the art.
[0358] In embodiments, the pharmaceutical compositions are
administered orally in a liquid form. Liquid dosage forms for oral
administration of an active ingredient include pharmaceutically
acceptable emulsions, microemulsions, solutions, suspensions,
syrups and elixirs. Liquid preparations for oral administration may
be presented as a dry product for constitution with water or other
suitable vehicle before use. In addition to the active ingredient,
the liquid dosage forms can contain inert diluents commonly used in
the art, such as, for example, water or other solvents,
solubilizing agents and emulsifiers, such as ethyl alcohol,
isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol,
benzyl benzoate, propylene glycol, 1,3-butylene glycol, oils (e.g.
cottonseed, groundnut, corn, germ, olive, castor and sesame oils),
glycerol, tetrahydrofuryl alcohol, polyethylene glycols and fatty
acid esters of sorbitan, and mixtures thereof. In addition to inert
diluents, the liquid pharmaceutical compositions can include
adjuvants such as wetting agents, emulsifying and suspending
agents, sweetening, flavoring, coloring, perfuming and preservative
agents, and the like. Suspensions, in addition to the active
ingredient(s) can contain suspending agents such as, but not
limited to, ethoxylated isostearyl alcohols, polyoxyethylene
sorbitol and sorbitan esters, microcrystalline cellulose, aluminum
metahydroxide, bentonite, agar-agar and tragacanth, and mixtures
thereof. Suitable liquid preparations may be prepared by
conventional means with a pharmaceutically acceptable additive(s)
such as a suspending agent (e.g. sorbitol syrup, methyl cellulose
or hydrogenated edible fats); emulsifying agent (e.g. lecithin or
acacia); non-aqueous vehicle (e.g. almond oil, oily esters or ethyl
alcohol); and/or preservative (e.g. methyl or propyl
p-hydroxybenzoates or sorbic acid). The active ingredient(s) can
also be administered as a bolus, electuary, or paste.
[0359] For buccal administration, the composition may take the form
of tablets or lozenges formulated in a conventional manner.
[0360] In embodiments, the pharmaceutical compositions are
administered by non-oral means such as by topical application,
transdermal application, injection, and the like. In related
embodiments, the pharmaceutical compositions are administered
parenterally by injection, infusion, or implantation (e.g.
intravenous, intramuscular, intra-arterial, subcutaneous, and the
like).
[0361] Presently disclosed compounds may be formulated for
parenteral administration by injection, including using
conventional catheterization techniques or infusion. Formulations
for injection may be presented in unit dosage form, e.g. in ampules
or in multi-dose containers, with an added preservative. The
compositions may take such forms as suspensions, solutions or
emulsions in oily or aqueous vehicles, and may contain a
formulating agent such as a suspending, stabilizing and/or
dispersing agent recognized by those of skill in the art.
Alternatively, the active ingredient may be in powder form for
reconstitution with a suitable vehicle, e.g. sterile pyrogen-free
water, before use.
[0362] The pharmaceutical compositions may be administered directly
to the central nervous system. Accordingly, in certain embodiments
the compositions are administered directly to the central nervous
system so as to avoid the blood brain barrier. In some embodiments,
the composition can be administered via direct spinal cord
injection. In embodiments, the composition is administered by
intrathecal injection. In some embodiments, the composition is
administered via intracerebroventricular injection. In embodiments,
the composition is administered into a cerebral lateral ventricle.
In embodiments, the composition is administered into both cerebral
lateral ventricles. In additional embodiments, the composition is
administered via intrahippocampal injection. The compositions may
be administered in one injection or in multiple injections. In
other embodiments, the composition is administered to more than one
location (e.g. to two sites in the central nervous system).
[0363] The pharmaceutical compositions can be in the form of
sterile injections. The pharmaceutical compositions can be
sterilized by, for example, filtration through a bacteria-retaining
filter, or by incorporating sterilizing agents in the form of
sterile solid compositions which can be dissolved in sterile water,
or some other sterile injectable medium immediately before use. To
prepare such a composition, the active ingredient is dissolved or
suspended in a parenterally acceptable liquid vehicle. Exemplary
vehicles and solvents include, but are not limited to, water, water
adjusted to a suitable pH by addition of an appropriate amount of
hydrochloric acid, sodium hydroxide or a suitable buffer,
1,3-butanediol, Ringer's solution and isotonic sodium chloride
solution. The pharmaceutical composition can also contain one or
more preservatives, for example, methyl, ethyl or n-propyl
p-hydroxybenzoate. To improve solubility, a dissolution enhancing
or solubilizing agent can be added or the solvent can contain
10-60% w/w of propylene glycol or the like.
[0364] The pharmaceutical compositions can contain one or more
pharmaceutically acceptable sterile isotonic aqueous or nonaqueous
solutions, dispersions, suspensions or emulsions, or sterile
powders, which can be reconstituted into sterile injectable
solutions or dispersions just prior to use. Such pharmaceutical
compositions can contain antioxidants; buffers; bacteriostats;
solutes, which render the formulation isotonic with the blood of
the intended recipient; suspending agents; thickening agents;
preservatives; and the like.
[0365] Examples of suitable aqueous and nonaqueous carriers, which
can be employed in the pharmaceutical compositions of the invention
include water, ethanol, polyols (such as glycerol, propylene
glycol, polyethylene glycol, and the like), and suitable mixtures
thereof, vegetable oils, such as olive oil, and injectable organic
esters, such as ethyl oleate. Proper fluidity can be maintained,
for example, by the use of coating materials, such as lecithin, by
the maintenance of the required particle size in the case of
dispersions, and by the use of surfactants. In some embodiments, in
order to prolong the effect of an active ingredient, it is
desirable to slow the absorption of the compound from subcutaneous
or intramuscular injection. This can be accomplished by the use of
a liquid suspension of crystalline or amorphous material having
poor water solubility. The rate of absorption of the active
ingredient then depends upon its rate of dissolution which, in
turn, can depend upon crystal size and crystalline form.
Alternatively, delayed absorption of a parenterally-administered
active ingredient is accomplished by dissolving or suspending the
compound in an oil vehicle. In addition, prolonged absorption of
the injectable pharmaceutical form can be brought about by the
inclusion of agents that delay absorption such as aluminum
monostearate and gelatin.
[0366] Controlled release parenteral compositions can be in form of
aqueous suspensions, microspheres, microcapsules, magnetic
microspheres, oil solutions, oil suspensions, emulsions, or the
active ingredient can be incorporated in biocompatible carrier(s),
liposomes, nanoparticles, implants or infusion devices. Materials
for use in the preparation of microspheres and/or microcapsules
include, but are not limited to, biodegradable/bioerodible polymers
such as polyglactin, poly-(isobutyl cyanoacrylate),
poly(2-hydroxyethyl-L-glutamine) and poly(lactic acid).
Biocompatible carriers which can be used when formulating a
controlled release parenteral formulation include carbohydrates
such as dextrans, proteins such as albumin, lipoproteins or
antibodies. Materials for use in implants can be non-biodegradable,
e.g. polydimethylsiloxane, or biodegradable such as, e.g.,
poly(caprolactone), poly(lactic acid), poly(glycolic acid) or
poly(ortho esters).
[0367] For topical administration, a presently disclosed compound
may be formulated as an ointment or cream. Presently disclosed
compounds may also be formulated in rectal compositions such as
suppositories or retention enemas, e.g. containing conventional
suppository bases such as cocoa butter or other glycerides.
[0368] For intranasal administration or administration by
inhalation, presently disclosed compounds may be conveniently
delivered in the form of a solution or suspension from a pump spray
container that is squeezed or pumped by the patient or as an
aerosol spray presentation from a pressurized container or a
nebulizer, with the use of a suitable propellant, e.g.
dichlorodifluoromethane, trichlorofluoromethane,
dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In
the case of a pressurized aerosol, the dosage unit may be
determined by providing a valve to deliver a metered amount. The
pressurized container or nebulizer may contain a solution or
suspension of the presently disclosed compound. Capsules and
cartridges (made, for example, from gelatin) for use in an inhaler
or insufflator may be formulated containing a powder mix of a
presently disclosed compound and a suitable powder base such as
lactose or starch.
[0369] Generally, the agents and compositions described herein are
administered in an effective amount or quantity sufficient to treat
or prevent a supranuclear gaze palsy in a subject in need thereof.
Typically, the dose can be adjusted within this range based on,
e.g., age, physical condition, body weight, sex, diet, time of
administration, and other clinical factors. Determination of an
effective amount is well within the capability of those skilled in
the art.
[0370] Having been generally described herein, the follow
non-limiting examples are provided to further illustrate this
invention.
EXAMPLES
[0371] General Procedures for Chemical Synthesis
[0372] General Procedure A: Carbamate Formation with
Triphosgene
[0373] To a suspension of amine hydrochloride (1 equivalent) and
triethylamine (3-4 equivalents) in a THF (concentration
.about.0.2M) at room temperature was added triphosgene (0.35
equivalents). The reaction mixture was stirred for 10 min and small
amount of ether (1-2 mL) was added. The triethylammonium salt was
filtered off to afford a clear solution of isocyanate in
THF/ether.
[0374] To a solution of alcohol (1.5 equivalents) in THF
(concentration .about.0.2M) at room temperature was added NaH [60%,
oil] (1.5 equivalents). The reaction mixture was stirred for 15 min
and the above solution (isocyanate in THF/ether) was added
dropwise. In a standard workup, the reaction was quenched with
brine. The solution was extracted with EtOAc and the organic layer
was dried over Na.sub.2SO.sub.4, filtered and concentrated. The
crude material was purified on combiflash (SiO.sub.2 cartridge,
CHCl.sub.3 and 2N NH.sub.3 in MeOH) to afford the corresponding
carbamate.
[0375] General Procedure B: Alkylation with Organocerium
[0376] A suspension of CeCl.sub.3 (4 equivalents) in THF
(concentration .about.0.2M) was stirred at room temperature for 1
h. The suspension was cooled to -78.degree. C. and MeLi/Ether
[1.6M] (4 equivalents) was added dropwise. The organocerium complex
was allowed to form for a period of 1 h and a solution of nitrile
(1 equivalent) in THF (concentration 2.0M) was added dropwise. The
reaction mixture was warmed up to room temperature and stirred for
18 h. The solution was cooled to 0.degree. C. and quenched with
water (.about.1 mL) followed by addition of 50% aqueous solution of
ammonium hydroxide (.about.3 mL) until precipitated formed and
settled to the bottom of the flask. The mixture was filtered
through a pad of celite and concentrated. The crude material was
treated with a solution of HCl/dioxane [4.0M]. The intermediate
arylpropan-2-amine hydrochloride was triturated in ether and used
as is for the next step. Alternatively, the crude free base amine
was purified on combiflash (SiO.sub.2 cartridge, CHCl.sub.3 and 2N
NH.sub.3 in MeOH) to afford the corresponding arylpropylamine.
[0377] General Procedure C: Suzuki Coupling
[0378] To a solution of aryl halide (1 equivalent) in a mixture of
DME/water [4:1] (concentration .about.0.2M) was added boronic acid
(2 equivalents), palladium catalyst (0.1-0.25 equivalent) and
sodium carbonate (2 equivalents). The reaction mixture was
microwaved 25 min at 150.degree. C. After filtering through a
celite plug and concentrating, the crude product was purified on
combiflash (SiO.sub.2 cartridge, CHCl.sub.3 and 2N NH.sub.3 in
MeOH) to afford the corresponding coupling adduct.
[0379] Alternatively: To a solution of aryl halide (1 equivalent)
in a mixture of toluene/water [20:1] (concentration .about.0.2 M)
was added boronic acid (1.3-2.5 equivalents), palladium catalyst
(0.05-0.15 equivalent), tricyclohexylphosphine (0.15-0.45
equivalent) and potassium phosphate (5 equivalents). The reaction
mixture was microwaved 25 min at 150.degree. C. After filtering
through a celite plug and concentrating, the crude product was
purified on combiflash (SiO.sub.2 cartridge, CHCl.sub.3 and 2N
NH.sub.3 in MeOH) to afford the corresponding coupling adduct.
[0380] General Procedure D: Cyclopropanation
[0381] To a mixture of aryl nitrile (1 equivalent) and
Ti(Oi-Pr).sub.4 (1.7 equivalents) stirring at -70.degree. C., was
added dropwise EtMgBr [3.0 M in ether] (1.1 equivalents). The
reaction mixture was allowed to warm to 25.degree. C. and stirred
for 1 h. To the above mixture was added BF.sub.3Et.sub.20 (3
equivalents) dropwise at 25.degree. C. After the addition, the
mixture was stirred for another 2 h, and then quenched with aqueous
HCl [2M]. The resulting solution was then basified by adding
aqueous NaOH [2M]. The organic material was extracted with ethyl
ether. The organic layers were combined, dried over
Na.sub.2SO.sub.4, filtered and concentrated. The crude material was
purified by silica gel column chromatography (eluting with
petroleum ether/EtOAc: 10/1 to 1/1) to give the corresponding
1-aryl-cyclopropanamine.
[0382] General Procedure E: Biaryl Coupling using Suzuki
Conditions
[0383] To a stirred solution of the aryl halide component (1
equivalent) in 5:1 (v/v) dioxane/water (.about.0.15 M) or 5:1 (v/v)
N,N-dimethylformamide (.about.0.15 M), was added the arylboronate
or arylboronic acid component (1-1.5 equivalents), sodium carbonate
(2-3 equivalents) and
[1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) (0.05
equivalents). The mixture was heated (90.degree. C.) overnight and
then filtered through a plug of Celite. The Celite was rinsed with
ethyl acetate and the combined filtrate was washed with brine,
dried (Na.sub.2SO.sub.4) and concentrated. The residue was purified
by flash chromatography over silica.
[0384] General Procedure F: Carbamate Formation using an Isocyanate
Generated via a Mixed Anhydride/Curtius Rearrangement Route
[0385] To a stirred solution of the carboxylic acid component (1
equivalent) in tetrahydrofuran (.about.0.1 M) was added
triethylamine (2 equivalents). The reaction was cooled (0.degree.
C.) and treated with isobutyl chloroformate (1.5 equivalents).
After 1 hour at 0.degree. C., a solution of sodium azide (2
equivalents) in water (.about.1 M) was added and the reaction was
allowed to warm to room temperature. After overnight stirring, the
reaction was diluted with water and extracted with ethyl acetate.
The combined extracts were washed with aqueous sodium bicarbonate
solution and brine, dried (Na.sub.2SO.sub.4) and concentrated. The
crude acyl azide was further dried via coevaporation with toluene
and then taken up in toluene (.about.0.1 M). The stirred solution
was refluxed for 2-2.5 hours, cooled and treated with an alcohol
component (1.25-2 equivalents). The reaction was heated at reflux
overnight and then concentrated. The residue was taken up in either
ethyl acetate or chloroform and washed with aqueous sodium
carbonate, (Na.sub.2SO.sub.4) and concentrated. The crude product
was purified by flash chromatography over silica using
chloroform/methanol (less polar carbamates) or
chloroform/methanol/ammonia (more polar carbamates) solvent
gradients.
Example 1: Synthesis of Quinuclidine Compounds
1-azabicyclo[2.2.2]oct-3-yl[2-(4'-fluorobiphenyl-3-yl)propan-2-yl]carbamat-
e (Compound 1)
[0386] Using General Procedure C, 1-azabicyclo[2.2.2]oct-3-yl
[2-(3-bromophenyl)propan-2-yl]carbamate (600 mg, 1.63 mmol),
4-fluorophenyl boronic acid (457 mg, 3.27 mmol) and palladium (II)
acetate gave the title compound as a white solid (373 mg; 60%).
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 7.56 (s, 1H), 7.52 (dd,
J=5.4, 8.4 Hz, 2H), 7.42-7.38 (m, 3H), 7.12 (m, 2H), 5.18 (5, 1H),
4.62 (s, 1H), 2.66 (m, 6H), 1.72 (s, 6H), 2.01-0.83 (m, 5H) ppm.
.sup.13C NMR (100 MHz, CDCl.sub.3) .delta. 125.0, 124.0, 123.8,
116.0, 116.0, 71.3, 55.9, 55.5, 47.6, 46.7, 29.6, 25.6, 24.8, 19.8
ppm. Purity: 98.0% UPLCMS (210 nm); retention time 0.95 min; (M+1)
382.9. Anal. Calcd. for
C.sub.23H.sub.27FN.sub.2O.sub.2.0.37(CHCl.sub.3): C, 65.86; H,
6.47; N, 6.57. Found: C, 65.85; H, 6.69; N, 6.49.
(S)-quinuclidin-3-yl
2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-ylcarbamate (Compound
2)
[0387] To a stirred solution of 4-fluorothiobenzamide (8.94 g, 57.6
mmol) in ethanol (70 mL) was added ethyl 4-chloroacetoacetate (7.8
mL, 58 mmol). The reaction was heated at reflux for 4 hours,
treated with an addition aliquot of ethyl 4-chloroacetoacetate (1.0
mL, 7.4 mmol) and refluxed for an additional 3.5 hours. The
reaction was then concentrated and the residue was partitioned
between ethyl acetate (200 mL) and aqueous NaHCO.sub.3 (200 mL).
The organic layer was combined with a back-extract of the aqueous
layer (ethyl acetate, 1.times.75 mL), dried (Na.sub.2SO.sub.4) and
concentrated. The resulting amber oil was purified by flash
chromatography using a hexane/ethyl acetate gradient to afford
ethyl 2-(2-(4-fluorophenyl)thiazol-4-yl)acetate as a low melting,
nearly colourless solid (13.58 g, 89%).
[0388] To a stirred solution of ethyl
2-(2-(4-fluorophenyl)thiazol-4-yl)acetate (6.28 g, 23.7 mmol) in
DMF (50 mL) was added sodium hydride [60% dispersion in mineral
oil] (2.84 g, 71.0 mmol). The frothy mixture was stirred for 15
minutes before cooling in an ice bath and adding iodomethane (4.4
mL, 71 mmol). The reaction was stirred overnight, allowing the
cooling bath to slowly warm to room temperature. The mixture was
then concentrated and the residue partitioned between ethyl acetate
(80 mL) and water (200 mL). The organic layer was washed with a
second portion of water (1.times.200 mL), dried (Na.sub.2SO.sub.4)
and concentrated. The resulting amber oil was purified by flash
chromatography using a hexane/ethyl acetate gradient to afford
ethyl 2-(2-(4-fluorophenyl)thiazol-4-yl)-2-methylpropanoate as a
colourless oil (4.57 g, 66%).
[0389] To a stirred solution of ethyl
2-(2-(4-fluorophenyl)thiazol-4-yl)-2-methylpropanoate (4.56 g, 15.5
mmol) in 1:1:1 THF/ethanol/water (45 mL) was added lithium
hydroxide monohydrate (2.93 g, 69.8 mmol). The reaction was stirred
overnight, concentrated and redissolved in water (175 mL). The
solution was washed with ether (1.times.100 mL), acidified by the
addition of 1.0 N HCl (80 mL) and extracted with ethyl acetate
(2.times.70 mL). The combined extracts were dried
(Na.sub.2SO.sub.4) and concentrated to afford
2-(2-(4-fluorophenyl)thiazol-4-yl)-2-methylpropanoic acid as a
white solid (4.04 g, 98%). This material was used in the next step
without purification.
[0390] To a stirred and cooled (0.degree. c) solution of
2-(2-(4-fluorophenyl)thiazol-4-yl)-2-methylpropanoic acid (4.02 g,
15.2 mmol) in THF (100 mL) was added trimethylamine (4.2 mL, 30
mmol) followed by isobutyl chloroformate (3.0 mL, 23 mmol). The
reaction was stirred cold for another 1 hour before adding a
solution of sodium azide (1.98 g, 30.5 mmol) in water (20 mL). The
reaction was stirred overnight, allowing the cooling bath to slowly
warm to room temperature. The mixture was then diluted with water
(100 mL) and extracted with ethyl acetate (2.times.60 mL). The
combined extracts were washed with aqueous NaHCO.sub.3 (1.times.150
mL) and brine (1.times.100 mL), dried (Na.sub.2SO.sub.4) and
concentrated. After coevaporating with toluene (2.times.50 mL), the
resulting white solid was taken up in toluene (100 mL) and refluxed
for 4 hours. (S)-3-quinuclidinol (3.87 g, 30.4 mmol) was then added
and reflux was continued overnight. The reaction was concentrated
and the residue partitioned between ethyl acetate (100 mL) and
aqueous NaHCO.sub.3 (150 mL). The organic layer was washed with
water (1.times.150 mL), dried (Na.sub.2SO.sub.4) and concentrated.
The resulting off-white solid was purified by flash chromatography
using a chloroform/methanol/ammonia gradient to afford the title
compound as a white solid (4.34 g, 73%). .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 7.96-7.88 (m, 2H), 7.16-7.04 (m, 3H), 5.55 (br
s, 1H), 4.69-4.62 (m, 1H), 3.24-3.11 (m, 1H), 3.00-2.50 (m, 5H),
2.01-1.26 (m, 11H) ppm. .sup.13C NMR (400 MHz, CDCl.sub.3)
.delta.166.4, 165.1, 163.8 (d, J=250.3 Hz), 162.9, 155.0, 130.1 (d,
J=3.3 Hz), 128.4 (d, J=8.5 Hz), 115.9 (d, J=22.3 Hz), 112.5, 71.2,
55.7, 54.2, 47.5, 46.5, 28.0, 25.5, 24.7, 19.6 ppm. Purity: 100%
UPLCMS (210 nm & 254 nm); retention time 0.83 min; (M+1)
390.
(S)-quinuclidin-3-yl(2-(4'-(2-methoxyethoxy)-[1,1'-biphenyl]-4-yl)propan-2-
-yl)carbamate (Compound 3)
[0391] Using General Procedure E and the reaction inputs ethyl
2-(4-bromophenyl)-2-methylpropanoate and
4-(2-methoxyethoxy)phenylboronic acid, ethyl
2-(4'-(2-methoxyethoxy)-[1,1'-biphenyl]-4-yl)-2-methylpropanoate
was prepared as an off-white solid. To a stirred solution of this
compound (3.01 g, 8.78 mmol) in 1:1:1 (v/v/v)
tetrahydrofuran/ethanol/water (45 mL) was added lithium hydroxide
monohydrate (1.47 g, 61.4 mmol). The mixture was heated at reflux
overnight and then concentrated. The residue was dissolved in
water, treated with 1N hydrochloric acid (65 mL) and extracted with
ethyl acetate. The combined organic layers were washed with brine,
dried (Na.sub.2SO.sub.4) and concentrated to afford
2-(4'-(2-methoxyethoxy)-[1,1'-biphenyl]-4-yl)-2-methylpropanoic
acid as a white solid (2.75 g, 100%). This intermediate and
(S)-quinuclidin-3-ol were reacted according to General Procedure F
to generate the title compound as a colourless, glassy solid.
.sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 7.62-7.29 (m, 7H), 7.01
(d, J=8.9 Hz, 2H), 4.47-4.37 (m, 1H), 4.17-4.08 (m, 2H), 3.72-3.62
(m, 2H), 3.32 (s, 3H), 3.09-2.25 (m, 6H), 2.05-1.18 (m, 11H) ppm.
.sup.13C NMR (100 MHz, DMSO-d.sub.6) .delta. 157.9, 154.5, 146.7,
137.4, 132.5, 127.5, 125.7, 125.2, 114.8, 70.4, 70.0, 66.9, 58.2,
55.4, 54.2, 46.9, 45.9, 29.4, 25.3, 24.2, 19.2 ppm. Purity: 100%,
100% (210 & 254 nm) UPLCMS; retention time: 0.87 min;
(M+H.sup.+) 439.5.
1-azabicyclo[2.2.2]oct-3-yl[2-(biphenyl-3-yl)propan-2-yl]carbamate
(Compound 4)
[0392] Using General Procedure C, 1-azabicyclo[2.2.2]oct-3-yl
[2-(3-bromophenyl)propan-2-yl]carbamate (600 mg, 1.63 mmol),
phenylboronic acid (398 mg, 3.27 mmol) and palladium (II) acetate
gave the title compound as a white solid (379 mg, 64%). .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 7.61 (s, 1H), 7.56 (d, J=7.4 Hz, 2H),
7.50-7.38 (m, 4H), 7.34 (m, 2H), 5.16 (s, 1H), 4.63 (s, 1H),
3.39-2.09 (m, 6H), 1.72 (s, 6H), 2.02-0.73 (m, 5H) ppm. .sup.13C
NMR (100 MHz, CDCl.sub.3) .delta. 154.8, 147.8, 141.6, 129.0,
129.0, 128.6, 127.5, 125.8, 125.0, 124.0, 71.6, 71.3, 55.9, 55.5,
47.6, 46.8, 31.5, 30.2, 30.0, 29.5, 25.6, 24.8, 19.8 ppm. Purity:
99% UPLCMS (210 nm); retention time 0.84 min; (M+1) 365.0. Anal.
Calcd. for C.sub.23H.sub.28N.sub.2O.sub.2.0.29(CHCl.sub.3): C,
70.02; H, 7.14; N, 7.01. Found: C, 70.02; H, 7.37; N, 6.84.
(S)-quinuclidin-3-yl 2-(biphenyl-4-yl)propan-2-ylcarbamate
(Compound 5)
[0393] Using General Procedure B, bromobenzonitrile (2.00 g, 11.0
mmol) was converted to the corresponding
2-(4-bromophenyl)propan-2-amine (1.20 g, 51%) as a brown oil.
[0394] Using General Procedure A, 2-(4-bromophenyl)propan-2-amine
(1.0 g, 4.7 mmol) and (S)-quinuclidin-3-ol gave
(S)-quinuclidin-3-yl 2-(4-bromophenyl)propan-2-ylcarbamate (1.0 g,
58%) as a brown oil.
[0395] Using General Procedure C, the above bromide (200 mg, 0.540
mmol), phenylboronic acid (133 mg, 1.10 mmol) and
[PdCl.sub.2(pddf)]CH.sub.2Cl.sub.2 gave the title compound as a
white solid (70 mg, 35%). .sup.1H NMR (500 MHz, CDCl.sub.3) .delta.
7.60-7.53 (m, 4H), 7.47 (d, J=8.5 Hz, 2H), 7.42 (t, J=7.5 Hz, 2H),
7.33 (t, J=7.5 Hz, 1H), 5.26 (br s, 1H), 4.64 (m, 1H), 3.33-3.15
(m, 1H), 3.10-2.45 (m, 5H), 2.40-1.80 (m, 2H), 1.78-1.58 (m, 7H),
1.55-1.33 (m, 2H) ppm. .sup.13C NMR (125 MHz, CDCl.sub.3) .delta.
154.5, 146.1, 140.8, 139.5, 128.7, 127.2, 127.1, 127.1, 125.2,
70.9, 55.5, 55.1, 47.4, 46.4, 31.1, 29.5, 25.3, 24.5, 19.5 ppm.
Purity: 100% LCMS (214 nm & 254 nm); retention time 1.56 min;
(M+1) 365.
Quinuclidin-3-yl 1-(biphenyl-4-yl)cyclopropylcarbamate (Compound
6)
[0396] Using General Procedure D, bromobenzonitrile (3.00 g, 16.5
mmol) was converted to the corresponding
1-(4-bromophenyl)cyclopropanamine (1.80 g, 51%) as a yellow
solid.
[0397] Using General Procedure A, 1-(4-bromophenyl)cyclopropanamine
(1.0 g, 4.7 mmol) and quinuclidin-3-ol gave quinuclidin-3-yl
1-(4-bromophenyl)cyclopropyl-carbamate (1.3 g, 75%) as a white
semi-solid.
[0398] Using General Procedure C, the above carbamate (400 mg, 1.12
mmol), phenylboronic acid (267 mg, 2.22 mmol) and
[PdCl.sub.2(pddf)]CH.sub.2Cl.sub.2 the title compound as a viscous
oil (100 mg, 25%). .sup.1H NMR (500 MHz, CDCl.sub.3) .delta. 7.47
(d, J=7.5 Hz, 2H), 7.43 (d, J=8.0 Hz, 2H), 7.33 (t, J=7.5 Hz, 2H),
7.26-7.15 (m, 3H), 5.93 (br s, 0.6H), 5.89 (br s, 0.4H), 4.67 (m,
1H), 3.20-3.06 (m, 1H), 2.88-2.42 (m, 5H), 1.98-1.08 (m, 9H) ppm.
.sup.13C NMR (125 MHz, CDCl.sub.3) .delta. 155.0, 141.0, 139.7,
138.2, 127.7, 126.1, 126.0, 124.8, 124.1, 70.0, 54.5, 46.3, 45.4,
34.1, 24.3, 23.2, 18.3, 17.0 ppm. Purity: 100% LCMC (214 nm &
254 nm); retention time 1.52 min; (M+1) 363.
(S)-quinuclidin-3-yl 1-(4'-fluorobiphenyl-4-yl)cyclopropylcarbamate
(Compound 7)
[0399] Using General Procedure C, (S)-quinuclidin-3-yl
1-(4-bromophenyl)cyclopropyl carbamate, 4-F-phenylboronic acid and
[PdCl.sub.2(pddf)]CH.sub.2Cl.sub.2 gave the title compound as a
white solid (45%). .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta.
8.06-7.83 (d, 1H), 7.69-7.66 (m, 2H), 7.59-7.55 (m, 2H), 7.29-7.22
(m, 4H), 4.56-4.54 (m, 1H), 3.13-2.32 (m, 6H), 1.91-1.19 (m, 9H)
ppm. .sup.13C NMR (125 MHz, DMSO-d.sub.6) .delta. 163.2, 161.2,
156.4, 143.7, 136.9, 128.9, 128.8, 126.8, 125.6, 116.2, 116.0,
70.7, 55.8, 47.4, 46.4, 34.8, 25.7, 24.6, 19.6, 18.7, 18.6 ppm.
Purity: >97% LCMS (214 nm & 254 nm); retention time 1.96
min; (M+1) 381.2.
(S)-1-azabicyclo[2.2.2]oct-3-yl[1-(2',4'-difluorobiphenyl-4-yl)cyclopropyl-
]carbamate (Compound 8)
[0400] Using General Procedure C, (S)-quinuclidin-3-yl
1-(4-bromophenyl)cyclopropylcarbamate (0.446 g, 1.22 mmol),
2,4-difluorophenyl boronic acid (0.386 g, 2.44 mmol) and
Pd(OAc).sub.2 (0.015 g, 0.067 mmol) gave the title compound as a
tan solid (0.111 g, 23%). .sup.1H NMR (CDCl.sub.3) .delta. 7.43
(dd, J=8.4, 1.6 Hz, 2H), 7.40-7.33 (m, 1H), 7.31 (d, J=7.7 Hz, 2H),
6.99-6.81 (m, 2H), 5.54 (d, J=48.0 Hz, 1H), 4.82-4.65 (m, 1H),
3.30-3.07 (m, 1H), 2.98-2.44 (m, 5H), 1.97 (d, J=32.7 Hz, 1H), 1.83
(d, J=10.3 Hz, 1H), 1.64 (s, 1H), 1.52 (s, 1H), 1.39 (s, 1H), 1.31
(d, J=6.8 Hz, 4H) ppm. .sup.13C NMR major rotomer (CDCl.sub.3)
.delta. 162.2 (dd, J=12.8, 249.1 Hz), 159.8 (dd, J=11.8, 251.0 Hz),
156.9, 156.0, 142.6, 133.1, 131.3 (m), 128.9, 125.6, 124.9, 111.5
(dd, J=3.9, 21.2 Hz) 104.4 (dd, J=25.2, 29.4 Hz), 72.1, 71.6, 55.7,
47.4, 46.5, 35.7, 35.3, 25.5, 24.6, 24.4, 19.5, 18.1 ppm. Purity:
LCMS>99.3% (214 nm & 254 nm); retention time 0.90 min; (M+1)
399.0.
1-azabicyclo[2.2.2]oct-3-yl[1-(4'-methoxybiphenyl-4-yl)cyclopropyl]carbama-
te (Compound 9)
[0401] Using General Procedure C, quinuclidin-3-yl
1-(4-bromophenyl)cyclopropylcarbamate (0.485 g, 1.33 mmol),
4-methoxyphenyl boronic acid (0.404 g, 2.66 mmol) and Pd(OAc).sub.2
(0.016 g, 0.071 mmol) gave the title compound as a grey solid
(0.337 mg, 65%). .sup.1H NMR (CDCl.sub.3) .delta. 7.48 (dd, J=8.6,
5.5 Hz, 4H), 7.29 (d, J=7.6 Hz, 2H), 6.96 (d, J=8.8 Hz, 2H), 5.58
(d, J=48.7 Hz, 1H), 4.83-4.63 (m, 1H), 3.84 (s, 3H), 3.20 (dd,
J=24.0, 15.5 Hz, 1H), 2.97-2.42 (m, 5H), 1.97 (d, J=30.9 Hz, 1H),
1.81 (s, 1H), 1.75-1.33 (m, 3H), 1.28 (d, J=6.8 Hz, 4H) ppm.
.sup.13C NMR major rotomer (CDCl.sub.3) .delta. 159.1, 156.0,
141.4, 139.0, 133.4, 128.0, 126.7, 125.9, 114.2, 71.5, 55.7, 55.3,
47.4, 46.5, 35.3, 25.5, 24.6, 19.6, 17.8 ppm. Purity: LCMS>97.1%
(214 nm & 254 nm); retention time 0.88 min; (M+1) 393.4.
Quinuclidin-3-yl
2-(5-(4-fluorophenyl)thiophen-3-yl)propan-2-ylcarbamate (Compound
10)
[0402] To a stirred and cooled (0.degree. C.) solution of ethyl
5-bromothiophene-3-carboxylate (13.30 g, 56.57 mmol) in THF (100
mL) was added a solution of methylmagnesium bromide in diethyl
ether [3.0 M] (55.0 mL, 165 mmol), dropwise over 20 minutes. After
2 hours, the reaction solution was concentrated. The residue was
taken up in aqueous NH.sub.4Cl (200 mL) and extracted with ethyl
acetate (2.times.100 mL). The combined extracts were dried
(Na.sub.2SO.sub.4) and concentrated. The resulting amber oil was
purified by flash chromatography using a hexane/ethyl acetate
gradient to afford 2-(5-bromothiophen-3-yl)propan-2-ol as a pale
amber oil (8.05 g, 64%).
[0403] To a stirred solution of 2-(5-bromothiophen-3-yl)propan-2-ol
(8.03 g, 36.3 mmol) in methylene chloride (80 mL) was added sodium
azide (7.08 g, 109 mmol) followed by trifluoroacetic acid (8.0 mL;
dropwise over 5-6 minutes). The thickening suspension was stirred
for 1.5 hour before diluting with water (350 mL) and extracting
with ethyl acetate (1.times.200 mL). The organic layer was washed
with aqueous NaHCO.sub.3 (1.times.250 mL), dried (Na.sub.2SO.sub.4)
and concentrated to afford the crude azide product. To a stirred
solution of this material in THF (160 mL) was added water (11 mL)
followed by triphenylphosphine (23.8 g, 90.7 mmol). The reaction
was stirred for 2 days before concentrating. The resulting residue
was dissolved in ethyl acetate (250 mL) and extracted with 1 N
aqueous HCl (4.times.75 mL). The combined extracts were basified
with concentrated NH.sub.4OH and extracted with ethyl acetate
(2.times.100 mL). These extracts were, in turn, dried
(Na.sub.2SO.sub.4) and concentrated. The resulting amber oil was
purified by flash chromatography using a methylene
chloride/methanol/ammonia gradient to afford a mixture of
2-(5-bromothiophen-3-yl)propan-2-amine and triphenylphosphine oxide
(.about.70/30 ratio) as a viscous amber oil (1.32 g, 17%).
[0404] To a stirred solution of 3-quinuclidinol (3.00 g, 23.6 mmol)
in THF (100 mL) was added 4-nitrophenyl chloroformate (5.94 g,
29.5). After stirring for 4 hours, the precipitate was filtered
off, rinsed with THF and air dried on the frit under house vacuum.
The filter cake was dissolved in ethyl acetate (150 mL) and washed
with aqueous NaHCO.sub.3 (1.times.150 mL) and water (2.times.150
mL). The organic layer was dried (Na.sub.2SO.sub.4) and
concentrated to afford crude 4-nitrophenyl quinuclidin-3-yl
carbonate product, which was used in the next step without
purification.
[0405] To a stirred solution of
2-(5-bromothiophen-3-yl)propan-2-amine (0.366 g, 1.66 mmol) in THF
(10 mL) was added 4-nitrophenyl quinuclidin-3-yl carbonate (0.571
g, 1.95 mmol) and a few granules of 4-(dimethylamino)pyridine. The
mixture was refluxed overnight, concentrated and partitioned
between ethyl acetate (50 mL) and aqueous NaHCO.sub.3 (50 mL). The
organic layer was washed again with aqueous NaHCO.sub.3 (1.times.50
mL), dried (Na.sub.2SO.sub.4) and concentrated. The resulting dirty
yellow gum was purified by flash chromatography using a
chloroform/methanol/ammonia gradient to afford quinuclidin-3-yl
(1-(5-bromothiophen-3-yl)cyclopropyl)carbamate as an off-white
solid (0.305 g, 49%).
[0406] Using General Procedure C, quinuclidin-3-yl
(1-(5-bromothiophen-3-yl)cyclopropyl)carbamate (0.227 g, 0.742
mmol), 4-fluorophenyl boronic acid (0.208 g, 1.49 mmol),
tricyclohexylphosphine (0.021 g, 0.075 mmol), potassium phosphate
(0.866, 4.08 mmol) and palladium acetate (8.0 mg, 36 .mu.mol) gave
the title compound as a grey solid (0.142 g, 49%). .sup.1H NMR (400
MHz, CDCl.sub.3) .delta. 7.60-7.45 (m, 2H), 7.24-7.19 (m, 1H),
7.10-6.97 (m, 3H), 5.23 (br s, 1H), 4.72-4.61 (m, 1H), 3.30-3.04
(m, 1H), 3.03-2.25 (m, 5H), 2.09-1.02 (m, 11H) ppm. .sup.13C NMR
(400 MHz, CDCl.sub.3) .delta. 162.3 (d, J=247.1 Hz), 154.5, 149.8,
143.6, 130.7, 127.4 (d, J=8.1 Hz), 121.8, 118.9, 115.8 (d, J=21.6
Hz), 70.8, 55.5, 53.4, 47.3, 46.4, 29.0, 25.4, 24.4, 19.4 ppm.
Purity: 95.8% UPLCMS (210 nm & 254 nm); retention time 0.90
min; (M+1) 389.
(S)-quinuclidin-3-yl
2-(3-(4-fluorophenyl)isothiazol-5-yl)propan-2-ylcarbamate (Compound
11)
[0407] To stirred solution of
2-(3-(4-fluorophenyl)isothiazol-5-yl)propan-2-amine (1.21 g, 5.12
mmol) in toluene was added a solution of phosgene in toluene
[.about.1.9 M] (10.8 mL, 20.5 mmol). The reaction was heated at
reflux for two hours and then concentrated. The residue was
co-evaporated with toluene (2.times.15 mL) to afford the crude
isocyanate intermediate as golden oil. This material was taken up
in toluene (10 mL) and treated with (S)-3-quinuclidinol (0.749 g,
5.89 mmol). The reaction was heated at reflux overnight and
concentrated. The residue was purified by flash chromatography
using a chloroform/methanol/ammonia gradient to afford the title
compound as a white solid (0.971 g, 49%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 8.09-8.00 (m, 2H), 7.87 (br s, 1H), 7.75 (s,
1H), 7.35-7.25 (m, 2H), 4.54-4.45 (m, 1H), 3.14-2.92 (m, 1H),
2.87-2.17 (m, 5H), 1.98-0.98 (m, 11H) ppm. .sup.13C NMR (400 MHz,
DMSO-d.sub.6) .delta. 180.1, 165.6, 162.6 (d, J=246.4 Hz), 154.7,
131.2 (d, J=3.0 Hz), 128.7 (d, J=8.4 Hz), 118.2, 115.7 (d, J=21.8
Hz), 70.6, 55.3, 52.8, 46.9, 45.9, 29.9, 25.2, 24.2, 19.2 ppm.
Purity: 100% UPLCMS (210 nm & 254 nm); retention time 0.82 min;
(M+1) 390.
(S)-quinuclidin-3-yl
2-(4-(4-fluorophenyl)thiazol-2-yl)propan-2-ylcarbamate (Compound
12)
[0408] To a stirred solution of ethyl 3-amino-3-thioxopropanoate
(20.00g, 135.9 mmol) in ethanol (120 mL) was added
2-bromo-4'-fluoroacetophenone (29.49 g, 135.9 mmol). The mixture
was refluxed for 1 hour, concentrated and partitioned between ethyl
acetate (300 mL) and aqueous NaHCO.sub.3 (400 mL). The organic
layer was combined with a back-extract of the aqueous layer (ethyl
acetate, 1.times.100 mL), dried (Na.sub.2SO.sub.4) and
concentrated. The resulting light brown solid was purified by flash
chromatography using a hexane/ethyl acetate gradient to afford
ethyl 2-(4-(4-fluorophenyl)thiazol-2-yl)acetate as an off-white
solid (29.92 g, 83%).
[0409] To a stirred and cooled (-78.degree. C.) solution of ethyl
2-(4-(4-fluorophenyl)thiazol-2-yl)acetate (10.00 g, 37.69 mmol) in
THF (250 mL) was added a solution of potassium t-butoxide in THF
[1.0 M] (136 mL, 136 mmol), dropwise over 15 minutes, followed by
18-crown-6 (1.6 mL, 7.5 mmol). After an additional 30 minutes at
-78.degree. C., iodomethane (8.5 mL) was added, dropwise over 5
minutes. The reaction was stirred cold for another 2 hours before
pouring into water (450 mL) and extracting with ethyl acetate
(2.times.150 mL). The combined extracts were washed with brine
(1.times.200 mL), dried (Na.sub.2SO.sub.4) and concentrated. The
resulting brown oil was purified by flash chromatography using a
hexane/ethyl acetate gradient to afford ethyl
2-(4-(4-fluorophenyl)thiazol-2-yl)-2-methylpropanoate as a pale
amber oil (8.64 g, 78%).
[0410] To a stirred solution of ethyl
2-(4-(4-fluorophenyl)thiazol-2-yl)-2-methylpropanoate (0.900 g,
3.07 mmol) in 1:1:1 THF/ethanol/water (15 mL) was added lithium
hydroxide monohydrate (0.451 g, 10.7 mmol). After overnight
stirring, the reaction was concentrated and redissolved in water
(80 mL). The solution was washed with ether (1.times.50 mL),
acidified with the addition of 1N HCl (15 mL) and extracted with
ethyl acetate (2.times.50 mL). The combined extracts were dried
(Na.sub.2SO.sub.4) and concentrated to afford
2-(4-(4-fluorophenyl)thiazol-2-yl)-2-methylpropanoic acid as a pale
golden solid (0.808 g, 99%).
[0411] To stirred and cooled (0.degree. C.) solution of
2-(4-(4-fluorophenyl)thiazol-2-yl)-2-methylpropanoic acid (0.784 g,
2.96 mmol) in THF (25 mL) was added triethylamine (0.82 mL, 5.9
mmol) followed by isobutyl chloroformate (0.58 mL, 4.4 mmol). The
reaction was stirred cold for another 1 hour before adding a
solution of sodium azide (0.385 g, 5.92 mmol) in water (7 mL). The
reaction was stirred overnight, allowing the cooling bath to slowly
warm to room temperature. The mixture was then diluted with water
(100 mL) and extracted with ethyl acetate (2.times.60 mL). The
combined extracts were washed with aqueous NaHCO.sub.3 (1.times.150
mL) and brine (1.times.100 mL), dried (Na.sub.2SO.sub.4) and
concentrated. After co-evaporating with toluene (2.times.30 mL),
the resulting off-white solid was taken up in toluene (25 mL) and
refluxed for 4 hours. (S)-3-quinuclidinol (0.753 g, 5.92 mmol) was
then added and reflux was continued for 3 hours. The reaction was
concentrated and the residue was purified by flash chromatography
using a chloroform/methanol/ammonia gradient to afford the title
compound as a white solid (0.793 g, 69%). .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 7.90-7.81 (m, 2H), 7.32 (s, 1H), 7.14-7.05 (m,
2H), 5.76 (br s, 1H), 4.72-4.65 (m, 1H), 3.26-3.10 (m, 1H),
3.03-2.37 (m, 5H), 2.05-1.23 (m, 11H) ppm. .sup.13C NMR (400 MHz,
CDCl.sub.3) .delta. 177.6, 162.6 (d, J=248.4 Hz), 154.8, 153.6,
130.8 (d, J=3.2 Hz), 128.1 (d, J=8.1 Hz), 115.9 (d, J=21.7 Hz),
112.2, 71.6, 55.7, 47.4, 46.5, 29.1, 25.4, 24.7, 19.6 ppm. Purity:
100% UPLCMS (210 nm & 254 nm); retention time 0.82 min; (M+1)
390.
Quinuclidin-3-yl(2-(4'-(2-methoxyethoxy)-[1,1'-biphenyl]-4-yl)propan-2-yl)-
carbamate (Compound 13)
[0412] Using General Procedure F and the reaction inputs
2-(4'-(2-methoxyethoxy)-[1,1'-biphenyl]-4-yl)-2-methylpropanoic
acid (prepared as described in Example 3) and quinuclidin-3-ol, the
title compound was generated as a colourless, glassy solid (23%).
NMR data matched that of Example 3. Purity: 100%, 99.1% (210 &
254 nm) UPLCMS; retention time: 0.87 min; (M+H.sup.+) 439.0.
(S)-quinuclidin-3-yl(2-(3'-(2-methoxyethoxy)-[1,1'-biphenyl]-4-yl)propan-2-
-yl)carbamate (Compound 14)
[0413] Exchanging 4-(2-methoxyethoxy)phenylboronic acid for
3-(2-methoxyethoxy)phenylboronic acid, the reaction sequence
outlined in Example 3 was used to prepare
2-(3'-(2-methoxyethoxy)-[1,1'-biphenyl]-4-yl)-2-methylpropanoic
acid. This intermediate and quinuclidin-3-ol were reacted according
to General Procedure F to generate the title compound as a glassy,
colourless solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
7.63-7.31 (m, 6H), 7.24-7.10 (m, 2H), 6.92 (dd, J=8.2, 1.9 Hz, 1H),
4.51-4.34 (m, 1H), 4.21-4.08 (m, 2H), 3.72-3.64 (m, 2H), 3.32 (s,
3H), 3.09-2.26 (m, 5H), 2.04-1.22 (m, 9H) ppm. .sup.13C NMR (100
MHz, DMSO-d.sub.6) .delta. 158.9, 154.6, 147.6, 141.5, 137.6,
129.9, 126.3, 125.2, 118.9, 113.2, 112.5, 70.4, 70.0, 66.9, 58.2,
55.4, 54.2, 46.9, 45.9, 29.4, 25.3, 24.2, 19.2 ppm. Purity: 100%,
100% (210 & 254 nm) UPLCMS; retention time: 0.91 min; 15
(M+H.sup.+) 439.4.
Quinuclidin-3-yl(2-(4'-(2-methoxyethoxy)-[1,1'-biphenyl]-3-yl)propan-2-yl)-
carbamate (Compound 15)
[0414] Exchanging ethyl 2-(4-bromophenyl)-2-methylpropanoate for
ethyl 2-(3-bromophenyl)-2-methylpropanoate, the reaction sequence
outlined in Example 3 was used to prepare
2-(4'-(2-methoxyethoxy)-[1,1'-biphenyl]-3-yl)-2-methylpropanoic
acid. This intermediate and quinuclidin-3-ol were reacted according
to General Procedure F to generate the title compound as a yellow
solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 7.62-7.20 (m,
7H), 7.03 (d, J=8.7 Hz, 2H), 4.48-4.35 (m, 2H), 4.18-4.08 (m, 2H),
3.72-3.62 (m, 2H), 3.32 (s, 3H), 3.10-2.19 (m, 6H), 2.10-1.10 (m,
11H) ppm. .sup.13C NMR (100 MHz, DMSO-d.sub.6) .delta. 158.0,
154.6, 148.8, 139.5, 133.1, 128.5, 127.7, 123.8, 123.2, 122.7,
114.8, 70.4, 69.9, 67.0, 58.2, 55.3, 54.5, 47.0, 45.9, 29.4, 25.3,
24.2, 19.2 ppm. Purity: 97.4%, 94.6% (210 & 254 nm) UPLCMS;
retention time: 0.88 min; (M+H.sup.+) 439.3.
Quinuclidin-3-yl(2-(4'-(3-methoxypropoxy)-[1,1'-biphenyl]-4-yl)propan-2-yl-
)carbamate (Compound 16)
[0415] To a stirred solution of 4-iodophenol (10.05 g, 45.68 mmol)
in acetonitrile (100 mL) was added potassium carbonate (6.95 g,
50.2 mmol) and 1-chloro-3-methoxypropane (6.4 mL, 57.1 mmol). The
mixture was heated at reflux overnight and then concentrated. The
residue was taken up in water and extracted with ethyl acetate. The
combined extracts were washed with aqueous sodium bicarbonate
solution, dried (Na.sub.2SO.sub.4) and concentrated. The crude
material was purified by flash chromatography over silica using a
hexane/ethyl acetate eluent to afford
1-iodo-4-(3-methoxypropoxy)benzene as a colourless oil (4.39 g,
33%). This intermediate and ethyl
2-methyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)propano-
ate were reacted according to General Procedure E to generate ethyl
2-(4'-(3-methoxypropoxy)-[1,1'-biphenyl]-4-yl)-2-methylpropanoate.
To a stirred solution of this compound (0.693 g, 1.94 mmol) in
1:1:1 (v/v/v) tetrahydrofuran/ethanol/water (10 mL) was added
lithium hydroxide monohydrate (0.326 g, 7.77 mmol). The mixture was
heated at reflux overnight and then concentrated. The residue was
dissolved in water, treated with 1N hydrochloric acid (10 mL) and
extracted with ethyl acetate. The combined organic layers were
washed with brine, dried (Na.sub.2SO.sub.4) and concentrated to
afford
2-(4'-(3-methoxypropoxy)-[1,1'-biphenyl]-4-yl)-2-methylpropanoic
acid as a waxy, off-white solid (0.630 g, 99%). This intermediate
and quinuclidin-3-ol were reacted according to General Procedure F
to generate the title compound as a glassy, colourless solid (62%).
.sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 7.61-7.29 (m, 7H), 7.00
(d, J=8.8 Hz, 2H), 4.47-4.36 (m, 1H), 4.05 (t, J=6.4 Hz, 2H), 3.48
(t, J=6.3 Hz, 2H), 3.26 (s, 3H), 3.10-2.25 (m, 6H), 2.04-1.74 (m,
4H), 1.65-1.23 (m, 9H) ppm..sup.13C NMR (100 MHz, DMSO-d.sub.6)
.delta. 158.0, 154.5, 146.7, 137.4, 132.4, 127.5, 125.7, 125.2,
114.8, 69.9, 68.5, 64.6, 57.9, 55.4, 54.2, 46.9, 46.0, 29.4, 29.0,
25.2, 24.1, 19.2 ppm. Purity: 97.7%, 98.2% (210 & 254 nm)
UPLCMS; retention time: 0.96 min; (M+H.sup.+) 453.5.
Quinuclidin-3-yl(2-(4'-(hydroxymethyl)-[1,1'-biphenyl]-4-yl)propan-2-yl)ca-
rbamate (Compound 17)
[0416] Using General Procedure E and the reaction inputs ethyl
2-(4-bromophenyl)-2-methylpropanoate and 4-formylphenylboronic
acid, ethyl 2-(4'-formyl-[1,1'-biphenyl]-4-yl)-2-methylpropanoate
was prepared as a pale amber solid. This intermediate and
quinuclidin-3-ol were reacted according to General Procedure F to
generate quinuclidin-3-yl
(2-(4'-formyl-[1,1'-biphenyl]-4-yl)propan-2-yl)carbamate as foamy,
yellow solid. To a stirred solution of this material (0.755 g, 1.92
mmol) in 2:1 (v/v) tetrahydrofuran/ethanol (15 mL) was added sodium
borohydride (0.073 g, 1.93 mmol). After 45 minutes, the reaction
was diluted with water and extracted with chloroform. The combined
extracts were dried (Na.sub.2SO.sub.4) and concentrated onto
silica. Flash chromatography over silica using a
chloroform/methanol/ammonia eluent provided the title compound as a
white solid (0.323 g, 43%). .sup.1H NMR (400 MHz, DMSO-d.sub.6)
.delta. 7.66-7.29 (m, 9H), 5.18 (t, J=5.7 Hz, 1H), 4.53 (d, J=5.7
Hz, 2H), 4.46-4.37 (m, 1H), 3.11-2.19 (m, 6H), 2.11-1.10 (m, 11H)
ppm. .sup.13C NMR (100 MHz, DMSO-d.sub.6) .delta. 154.7, 147.3,
141.5, 138.4, 137.7, 127.0, 126.2, 126.1, 125.3, 70.0, 62.6, 55.4,
54.2, 46.9, 45.9, 29.4, 25.3, 24.2, 19.2 ppm. Purity: 97.5%, 99.1%
(210 & 254 nm) UPLCMS; retention time: 0.73 min; (M+H.sup.+)
395.
Quinuclidin-3-yl(2-(4'-(2-hydroxyethyl)-[1,1'-biphenyl]-4-yl)propan-2-yl)c-
arbamate (Compound 18)
[0417] Using General Procedure E and the reaction inputs
1-(2-(benzyloxy)ethyl)-4-bromobenzene and ethyl
2-methyl-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)propano-
ate, ethyl
2-(4'-(2-(benzyloxy)ethyl)-[1,1'-biphenyl]-4-yl)-2-methylpropan-
oate was prepared as a colourless gum. To a stirred solution of
this compound (1.34 g, 3.33 mmol) in 1:1:1 (v/v/v)
tetrahydrofuran/ethanol/water (18 mL) was added lithium hydroxide
monohydrate (0.698 g, 16.6 mmol). After heating at reflux
overnight, the reaction was concentrated and partitioned between
water and diethyl ether. The resulting emulsion was extracted
repeatedly with 0.2 N aqueous sodium hydroxide solution (5.times.50
mL). The clear portion of the aqueous layer was removed each time.
The combined aqueous layers were then treated with 1.0 N
hydrochloric acid (80 mL) and the resulting suspension of white
solid was extracted with ethyl acetate. The combined organic layers
were dried (Na.sub.2SO.sub.4) and concentrated to afford
2-(4'-(2-(benzyloxy)ethyl)-[1,1'-biphenyl]-4-yl)-2-methylpropanoic
acid as a white solid (1.20 g, 96%). This compound and
quinuclidin-3-ol were reacted according to General Procedure F to
generate quinuclidin-3-yl
(2-(4'-(2-benzyloxyethyl)-[1,1'-biphenyl]-4-yl)propan-2-yl)carbamate.
To a stirred solution of this material (0.435 g, 0.806 mmol) in
methanol was added 1.0 N hydrochloric acid (1 mL) and 10% palladium
on carbon (50% water; 0.087 g). The mixture was cycled between
vacuum and a nitrogen purge several times, refilling with hydrogen
after the last evacuation. After 1.25 hours the reaction was
filtered through Celite and concentrated. The residue was taken up
in aqueous sodium carbonate solution and extracted with 4:1 (v/v)
chloroform/isopropanol. The combined extracts were dried
(Na.sub.2SO.sub.4) and concentrated onto silica. Flash
chromatography over silica using a chloroform/methanol/ammonia
gradient provided the purified title compound as a colourless
solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 7.85-7.63 (m,
1H), 7.63-7.19 (m, 8H), 4.78-4.62 (m, 2H), 3.71-2.78 (m, 8H), 2.76
(t, J=6.8 Hz, 2H), 2.26-1.96 (m, 2H), 1.96-1.40 (m, 9H) ppm.
.sup.13C NMR (100 MHz, DMSO-d.sub.6) .delta. 153.8, 146.8, 138.7,
137.9, 137.6, 129.4, 126.3, 126.1, 125.3, 66.2, 62.1, 54.4, 52.8,
45.4, 44.5, 38.6, 29.5, 29.2, 24.0, 19.9, 16.6 ppm. Purity: 100%,
100% (210 & 254 nm) UPLCMS; retention time: 0.75 min;
(M+H.sup.+) 409.
Quinuclidin-3-yl(2-(2-(4-(3-methoxypropoxy)phenyl)thiazol-4-yl)propan-2-yl-
)carbamate (Compound 19)
[0418] To a stirred suspension of 4-methoxythiobenzamide (9.99 g,
59.7 mmol) in ethanol (75 mL) was added ethyl 4-chloroacetoacetate
(8.1 mL, 60 mmol). The mixture was heated at reflux for 4 hours
before cooling, adding additional ethyl 4-chloroacetoacetate (0.81
mL, 6.0 mmol) and returning to reflux. After 4 more hours of
heating the reaction was concentrated and partitioned between ethyl
acetate and aqueous sodium bicarbonate solution. The organic layer
was combined with additional ethyl acetate extracts, dried
(Na.sub.2SO.sub.4) and concentrated. The crude product was purified
by flash chromatography over silica using a hexane/ethyl acetate
gradient to afford ethyl 2-(2-(4-methoxyphenyl)thiazol-4-yl)acetate
as a pale amber oil (14.51 g, 87%). To a stirred solution of this
compound (14.48 g, 52.2 mmol) in N,N-dimethylformamide (125 mL) was
added sodium hydride (60% dispersion in mineral oil; 6.27 g, 157
mmol), portion wise over 15 minutes. The resulting red suspension
was cooled (0.degree. C.) and treated, dropwise over 10 minutes,
with iodomethane (9.80 mL, 157 mmol). The cooling bath was removed
and the reaction was allowed to stir 4 hours before concentrating
and partitioning the residue between ethyl acetate and water. The
organic layer was washed twice more with water, dried
(Na.sub.2SO.sub.4) and concentrated. The residue was purified by
flash chromatography over silica using a hexane/ethyl acetate
gradient to afford ethyl
2-(2-(4-methoxyphenyl)thiazol-4-yl)-2-methylpropanoate as a pale
amber oil (14.12 g, 89%). To a stirred solution of this
intermediate (14.12 g, 46.24 mmol) in methylene chloride (250 mL)
was added boron tribromide (11.0 mL, 116 mmol), dropwise over 5
minutes. After stirring overnight, the reaction was quenched by the
slow addition of methanol (.about.20 mL) and then concentrated. The
residue was taken up in methanol (250 mL) and concentrated sulfuric
acid (7.0 mL). The stirred solution was heated at reflux for 2
hours, concentrated and partitioned between ethyl acetate and
aqueous sodium bicarbonate solution. The organic layer was combined
with a second ethyl acetate extract of the aqueous layer, dried
(Na.sub.2SO.sub.4) and concentrated to afford methyl
2-(2-(4-hydroxyphenyl)thiazol-4-yl)-2-methylpropanoate as a white
solid (12.56 g, 98%). To a stirred solution of
1-bromo-3-methoxypropane (1.66 g, 10.8 mmol) in acetone (30 mL) was
added the phenol intermediate (2.00 g, 7.21 mmol) and potassium
carbonate (1.25 g, 9.04 mmol). The mixture was heated overnight at
reflux, filtered and concentrated. The residue was purified by
flash chromatography over silica using a hexane/ethyl acetate
gradient to afford methyl
2-(2-(4-(3-methoxypropoxy)phenyl)thiazol-4-yl)-2-methylpropanoate
as a faint amber gum (2.47 g, 98%). To a stirred solution of this
compound (2.45 g, 7.01 mmol) in 1:1:1 (v/v/v)
tetrahydrofuran/ethanol/water (45 mL) was added lithium hydroxide
monohydrate (1.47 g, 35.0 mmol). After overnight stirring, the
reaction was concentrated and partitioned between water and diethyl
ether. The aqueous layer was treated with 1.0 N hydrochloric acid
(40 mL) and extracted with ethyl acetate. The combined extracts
were dried (Na.sub.2SO.sub.4) and concentrated to afford
2-(2-(4-(3-methoxypropoxy)phenyl)thiazol-4-yl)-2-methylpropanoic
acid as a white solid (2.19 g, 40 93%). This compound and
quinuclidin-3-ol were reacted according to General Procedure F to
generate the title compound as a soft, faint amber solid. .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. 7.82 (d, J=8.9 Hz, 2H), 7.36
(br s, 1H), 7.24 (br s, 1H), 7.03 (d, J=8.9 Hz, 2H), 4.49-4.41 (m,
1H), 4.07 (t, J=6.4 Hz, 2H), 3.48 (t, J=6.4 Hz, 2H), 3.26 (s, 3H),
3.09-2.26 (m, 6H), 2.02-1.91 (m, 2H), 1.91-1.03 (m, 11H) ppm.
.sup.13C NMR (100 MHz, DMSO-d.sub.6) .delta. 165.8, 162.4, 160.0,
154.6, 127.5, 126.1, 114.9, 112.1, 70.1, 68.4, 64.8, 57.9, 55.4,
53.5, 46.9, 45.9, 28.9, 28.3, 25.2, 24.2, 19.2 ppm. Purity: 100%,
100% (210 & 254 nm) UPLCMS; retention time: 0.87 min;
(M+H.sup.+) 460.
Quinuclidin-3-yl(2-(2-(4-(2-methoxyethoxy)phenyl)thiazol-4-yl)propan-2-yl)-
carbamate (Compound 20)
[0419] To a stirred solution of 2-bromoethyl methyl ether (1.88 g,
13.5 mmol) in acetone was added methyl
2-(2-(4-hydroxyphenyl)thiazol-4-yl)-2-methylpropanoate (prepared as
described in Example 19, 2.00 g, 7.21 mmol) and potassium carbonate
(1.56 g, 11.3 mmol). After heating at reflux overnight, the mixture
was treated with additional 2-bromo ethyl methyl ether (1.88 g,
13.5 mmol) and potassium carbonate (1.56 g, 11.3 mmol). The
reaction was heated at reflux for a second night, filtered and
concentrated. The residue was purified by flash chromatography over
silica using a hexane/ethyl acetate gradient to afford methyl
2-(2-(4-(2-methoxyethoxy)phenyl)thiazol-4-yl)-2-methylpropanoate as
a white solid (2.71 g, 90%). To a stirred solution of this compound
(2.71 g, 8.08 mmol) in 1:1:1 (v/v/v) tetrahydrofuran/ethanol/water
(50 mL) was added lithium hydroxide monohydrate (1.70 g, 40.5
mmol). After overnight stirring, the reaction was concentrated and
partitioned between water and diethyl ether. The aqueous layer was
treated with 1.0 N hydrochloric acid (41 mL) and extracted with
ethyl acetate. The combined extracts were dried (Na.sub.2SO.sub.4)
and concentrated to afford
2-(2-(4-(2-methoxyethoxy)phenyl)thiazol-4-yl)-2-methylpropanoic
acid as a white solid (2.57 g, 99%). This compound and
quinuclidin-3-ol were reacted according to General Procedure F to
generate the title compound as a pale amber solid. .sup.1H NMR (400
MHz, DMSO-d.sub.6) .delta. 7.82 (d, J=8.8 Hz, 2H), 7.36 (br s, 1H),
7.24 (br s, 1H), 7.04 (d, J=8.8 Hz, 2H), 4.49-4.41 (m, 1H),
4.19-4.12 (m, 2H), 3.71-3.65 (m, 2H), 3.32 (s, 3H), 3.11-2.87 (m,
1H), 2.86-2.19 (m, 5H), 1.92-1.16 (m, 11H) ppm. .sup.13C NMR (100
MHz, DMSO-d.sub.6) .delta. 165.7, 162.9, 159.9, 154.6, 127.5,
126.2, 114.9, 112.2, 70.3, 70.1, 67.1, 58.2, 55.4, 53.5, 46.9,
45.9, 28.3, 25.2, 24.3, 19.2 ppm. Purity: 100%, 100% (210 & 254
nm) UPLCMS; retention time: 0.85 min; (M+H.sup.+) 446.
Quinuclidin-3-yl
2-(5-(4-(2-methoxyethoxy)phenyl)pyridin-2-yl)propan-2-ylcarbamate
(Compound 21)
[0420] Using General Procedure E and the reaction inputs
5-bromopicolinonitrile and
2-(4-(2-methoxyethoxy)phenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane,
5-(4-(2-methoxyethoxy)phenyl)picolinonitrile was prepared. Cerium
trichloride (8.05 g, 21.6 mmol) was loaded into a flask and dried
by heating (170.degree. C.) under vacuum for 3 hours. The solid was
taken up in tetrahydrofuran (20 mL) and stirred vigorously for 30
minutes. The suspension was cooled to -78.degree. C. and treated,
dropwise, with a 3.0 M solution of methyllithium in diethyl ether
(7.2 mL, 21.6 mmol). Following addition, the reaction was stirred
at -78.degree. C. for 1 hour before adding a solution of the above
aryl borate (1.83 g, 7.20 mmol) in tetrahydrofuran (20 mL). The
mixture was maintained at -78.degree. C. for 2 hours and then
allowed to warm to room temperature. At this time, the reaction was
quenched by the addition of aqueous ammonium hydroxide (10 mL) and
filtered through a plug of Celite. The filtrate was extracted with
ethyl acetate and the combined extracts were washed with brine,
dried (Na.sub.2SO.sub.4) and concentrated. The residue was purified
by flash chromatography over silica using ethyl acetate eluent to
afford 2-(5-(4-(2-methoxyethoxy)phenyl)pyridin-2-yl)propan-2-amine
as a yellow solid (0.800 g, 39%). To a stirred suspension of this
intermediate (0.500 g, 1.75 mmol) in water (10 mL) and concentrated
hydrochloric acid (0.44 mL) was added toluene (10 mL). The mixture
was cooled (0.degree. C.) and treated with, simultaneously over 1
hour, solutions of triphosgene (0.776 g, 2.62 mmol) in toluene (10
mL) and sodium bicarbonate (2.2 g, 26 mmol) in water (20 mL).
Following the additions, the reaction was stirred for an additional
30 minutes before the upper toluene layer was removed and dried
(Na.sub.2SO.sub.4). At the same time, a stirred solution of
quinuclidin-3-ol (0.445 g, 3.64 mmol) in tetrahydrofuran (10 mL)
was treated with sodium hydride (60% dispersion in mineral oil;
0.154 g, 3.85 mmol). This mixture was stirred for 5 minutes and
then added to the solution of crude isocyanate in toluene. The
reaction was stirred for 10 minutes, quenched with the addition of
brine (5 mL) and extracted with ethyl acetate. The combined
extracts were dried (Na.sub.2SO.sub.4) and concentrated. The
residue was purified by flash chromatography over reversed phase
silica to afford the title compound as a light yellow solid (0.100
g, 13%). .sup.1H NMR (500 MHz, CDCl.sub.3) .delta. 8.70-8.70 (d,
J=2.0 Hz, 1H), 7.83-7.81 (m, 1H), 7.49-7.47 (d, J=9.0 Hz, 2H),
7.45-7.43 (d, J=8.0 Hz, 1H), 7.03-7.01 (d, J=8.5 Hz, 2H), 6.63 (br
s, 1H), 4.68-4.66 (m, 1H), 4.16 (t, J=5.0 Hz, 2H), 3.77 (t, J=5.0
Hz, 2H), 3.45 (s, 3H), 3.19-2.70 (m, 6H), 2.15-1.89 (m, 2H), 1.76
(s, 6H), 1.73-1.36 (m, 3H) ppm. .sup.13C NMR (125 MHz, CDCl.sub.3)
.delta. 162.7, 158.9, 154.9, 145.9, 134.8, 134.3, 130.1, 128.1,
119.2, 115.2, 71.0, 70.8, 67.4, 59.2, 55.9, 55.7, 47.4, 46.5, 46.4,
27.9, 25.4, 24.6, 19.5 ppm. Purity: >99% (214 & 254 nm)
LCMS; retention time: 1.32 min; (M+H.sup.+) 440.2.
Quinuclidin-3-yl(2-(4'-(3-cyanopropoxy)-[1,1'-biphenyl]-4-yl)propan-2-yl)c-
arbamate (Compound 22)
[0421] To a stirred solution of 4-bromophenol (17.1 g, 98.8 mmol)
in acetonitrile (150 mL) was added 1-bromobutylnitrile (12.3 mL,
124 mmol) and potassium carbonate (15.0 g, 109 mmol). The mixture
was heated to reflux overnight, cooled and concentrated. The
residue was taken up in water and extracted with ethyl acetate. The
combined extracts were dried (Na.sub.2SO.sub.4) and concentrated
and the crude material was purified by flash chromatography over
silica using a hexane/ethyl acetate eluent to afford
4-(4-bromophenoxy)butanenitrile as a white solid (20.8 g, 88%). To
a stirred solution of this product in N,N-dimethylformamide (100
mL), was added bis(pinacolato)diboron (4.60 g, 18.1 mmol),
potassium acetate (7.41 g, 75.5 mmol) and
[1,1'-bis(diphenylphosphino)ferrocene]-dichloropalladium(II)
complex with dichloromethane (0.616 g, 1.04 mmol). The mixture was
heated to reflux overnight and then concentrated. The residue was
taken up in ethyl acetate and washed with water and brine. The
organic layer was dried (Na.sub.2SO.sub.4) and concentrated and the
crude product was purified by flash chromatography over silica
using a hexane/ethyl acetate eluent to afford
4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenoxy)butaneni-
trile as a white solid (3.43 g, 79%). This product and
quinuclidin-3-yl (2-(4-bromophenyl)propan-2-yl)carbamate (prepared
by reacting quinuclidin-3-ol and 2-(4-bromophenyl)propan-2-amine
using General Procedure F) were reacted according to General
Procedure E to generate the title compound as a white solid.
.sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 7.67-7.26 (m, 7H), 7.02
(d, J=8.8 Hz, 2H), 4.50-4.33 (m, 1H), 4.08 (t, J=6.0 Hz, 2H),
3.14-2.18 (m, 8H), 2.04 (quin, J=6.7 Hz, 2H), 1.94-1.70 (m, 11H)
ppm. .sup.13C NMR (100 MHz, DMSO-d.sub.6) .delta. 157.7, 154.5,
146.8, 137.4, 132.7, 127.6, 125.7, 125.2, 120.2, 114.9, 70.0, 65.8,
55.4, 54.2, 46.9, 45.9, 29.4, 25.3, 24.7, 24.2, 19.2, 13.4 ppm.
Purity: 100%, 98.9% (210 & 254 nm) UPLCMS; retention time: 0.88
min; (M+H.sup.+) 448.6.
Quinuclidin-3-yl(2-(4'-cyanomethoxy)-[1,1'-biphenyl]-4-yl)propan-2-yl)carb-
amate (Compound 23)
[0422] Using General Procedure E and the reaction inputs
quinuclidin-3-yl (2-(4-bromophenyl)propan-2-yl)carbamate (prepared
by reacting quinuclidin-3-ol and 2-(4-bromophenyl)propan-2-amine
using General Procedure F) and 4-(cyanomethoxy)phenylboronic acid,
the title compound was prepared as a pale amber solid. .sup.1H NMR
(400 MHz, DMSO-d.sub.6) .delta. 7.65 (d, J=8.2 Hz, 2H), 7.60-7.31
(m, 5H), 7.15 (d, J=8.9 Hz, 2H), 5.21 (s, 2H), 4.53-4.30 (m, 1H),
3.18-2.19 (m, 6H), 2.05-1.18 (m, 11H) ppm. .sup.13C NMR (100 MHz,
DMSO-d.sub.6) .delta. 155.8, 154.6, 147.2, 137.2, 134.4, 127.8,
126.0, 125.3, 116.7, 115.3, 70.0, 55.4, 54.2, 53.5, 46.9, 45.9,
29.4, 25.2, 24.2, 19.2 ppm. Purity: 100%, 100% (210 & 254 nm)
UPLCMS; retention time: 0.85 min; (M+H.sup.+) 420.3.
Example 2: Preparation of
(S)-Quinuclidin-3-yl(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbam-
ate Free Base
[0423] Step 1: Dimethylation with Methyl Iodide
##STR00009##
[0424] A 3N RB flask was equipped with a thermometer, an addition
funnel and a nitrogen inlet. The flask was flushed with nitrogen
and potassium tert-butoxide (MW 112.21, 75.4 mmol, 8.46 g, 4.0
equiv., white powder) was weighed out and added to the flask via a
powder funnel followed by the addition of THF (60 mL). Most of the
potassium tert-butoxide dissolved to give a cloudy solution. This
mixture was cooled in an ice-water bath to 0-2.degree. C. (internal
temperature). In a separate flask, the starting ester (MW 265.3,
18.85 mmol, 5.0 g, 1.0 equiv.) was dissolved in THF (18 mL+2 mL as
rinse) and transferred to the addition funnel. This solution was
added dropwise to the cooled mixture over a period of 25-30 min,
keeping the internal temperature below 5.degree. C. during the
addition. The reaction mixture was cooled back to 0-2.degree. C. In
a separate flask, a solution of methyl iodide (MW 141.94, 47.13
mmol, 6.7 g, 2.5 equiv.) in THF (6 mL) was prepared and transferred
to the addition funnel. The flask containing the methyl iodide
solution was then rinsed with THF (1.5 mL) which was then
transferred to the addition funnel already containing the clear
colorless solution of methyl iodide in THF. This solution was added
carefully dropwise to the dark brown reaction mixture over a period
of 30-40 min, keeping the internal temperature below 10.degree. C.
at all times during the addition. After the addition was complete,
the slightly turbid mixture was stirred for an additional 1 h
during which time the internal temperature dropped to 0-5.degree.
C. After stirring for an hour at 0-5.degree. C., the reaction
mixture was quenched with the slow dropwise addition of 5.0M
aqueous HCl (8 mL) over a period of 5-7 min. The internal
temperature was maintained below 20.degree. C. during this
addition. After the addition, water (14 mL) was added and the
mixture was stirred for 2-3 min. The stirring was stopped and the
two layers were allowed to separate. The two layers were then
transferred to a 250 mL 1N RB flask and the THF was evaporated in
vacuo as much as possible to obtain a biphasic layer of THF/product
and water. The two layers were allowed to separate. A THF solution
of the Step 1 product was used in the next reaction.
[0425] Step 2: Hydrolysis of the Ethyl Ester with LiOH
Monohydrate
##STR00010##
[0426] The crude ester in THF was added to the reaction flask.
Separately, LiOH.H.sub.2O (MW 41.96, 75.0 mmol, 3.15 grams, 2.2
equiv.) was weighed out in a 100 mL beaker to which a stir bar was
added. Water (40 mL) was added and the mixture was stirred till all
the solid dissolved to give a clear colorless solution. This
aqueous solution was then added to the 250 mL RB flask containing
the solution of the ester in tetrahydrofuran (THF). A condenser was
attached to the neck of the flask and a nitrogen inlet was attached
at the top of the condenser. The mixture was heated at reflux for
16 hours. After 16 hours, the heating was stopped and the mixture
was cooled to room temperature. The THF was evaporated in vacuo to
obtain a brown solution. An aliquot of the brown aqueous solution
was analyzed by HPLC and LC/MS for complete hydrolysis of the ethyl
ester. Water (15 mL) was added and this aqueous basic solution was
extracted with TBME (2.times.40 mL) to remove the t-butyl ester.
The aqueous basic layer was cooled in an ice-water bath to
0-10.degree. C. and acidified with dropwise addition of
concentrated HCl to pH.about.1 with stirring. To this gummy solid
in the aqueous acidic solution was added TBME (60 mL) and the
mixture was shaken and then stirred vigorously to dissolve all the
acid into the TBME layer. The two layers were transferred to a
separatory funnel and the TBME layer was separated out. The pale
yellow aqueous acidic solution was re-extracted with TBME (40 mL)
and the TBME layer was separated and combined with the previous
TBME layer. The aqueous acidic layer was discarded. The combined
TBME layers are dried over anhydrous Na.sub.2SO.sub.4, filtered and
evaporated in vacuo to remove TBME and obtain the crude acid as an
orange/dark yellow oil that solidified under high vacuum to a dirty
yellow colored solid. The crude acid was weighed out and
crystallized by heating it in heptane/TBME (3:1, 5 mL/g of crude)
to give the acid as a yellow solid.
[0427] Step 3: Formation of Hydroxamic Acid with NH.sub.2OH.HCl
##STR00011##
[0428] The carboxylic acid (MW 265.3, 18.85 mmol, 5.0 g, 1.0
equiv.) was weighed and transferred to a 25 mL 1N RB flask under
nitrogen. THF (5.0 mL) was added and the acid readily dissolved to
give a clear dark yellow to brown solution. The solution was cooled
to 0-2.degree. C. (bath temperature) in an ice-bath and N,
N'-carbonyldiimidazole (CDI; MW 162.15, 20.74 mmol, 3.36 g, 1.1
equiv.) was added slowly in small portions over a period of 10-15
minutes. The ice-bath was removed and the solution was stirred at
room temperature for 1 h. After 1 h of stirring, the solution was
again cooled in an ice-water bath to 0-2.degree. C. (bath
temperature). Hydroxylamine hydrochloride (NH.sub.2OH.HCl; MW
69.49, 37.7 mmol, 2.62 g, 2.0 equiv.) was added slowly in small
portions as a solid over a period of 3-5 minutes as this addition
was exothermic. After the addition was complete, water (1.0 mL) was
added to the heterogeneous mixture dropwise over a period of 2
minutes and the reaction mixture was stirred at 0-10.degree. C. in
the ice-water bath for 5 minutes. The cooling bath was removed and
the reaction mixture was stirred under nitrogen at room temperature
overnight for 20-22 h. The solution became clear as all of the
NH.sub.2OH.HCl dissolved. After 20-22 h, an aliquot of the reaction
mixture was analyzed by High Pressure Liquid Chromatography (HPLC).
The THF was then evaporated in vacuo and the residue was taken up
in dichloromethane (120 mL) and water (60 mL). The mixture was
transferred to a separatory funnel where it was shaken and the two
layers allowed to separate. The water layer was discarded and the
dichloromethane layer was washed with 1N hydrochloride (HCl; 60
mL). The acid layer was discarded. The dichloromethane layer was
dried over anhydrous Na.sub.2SO.sub.4, filtered and the solvent
evaporated in vacuo to obtain the crude hydroxamic acid as a pale
yellow solid that was dried under high vacuum overnight.
[0429] Step 3 continued: Conversion of hydroxamic acid to cyclic
intermediate (not isolated)
##STR00012##
[0430] The crude hydroxamic acid (MW 280.32, 5.1 g) was transferred
to a 250 mL 1N RB flask with a nitrogen inlet. A stir bar was added
followed by the addition of acetonitrile (50 mL). The solid was
insoluble in acetonitrile. The yellow heterogeneous mixture was
stirred for 2-3 minutes under nitrogen and CDI (MW 162.15, 20.74
mmol, 3.36 g, 1.1 equiv.) was added in a single portion at room
temperature. No exotherm was observed. The solid immediately
dissolved and the clear yellow solution was stirred at room
temperature for 2-2.5 h. After 2-2.5 h, an aliquot was analyzed by
HPLC and LC/MS which showed conversion of the hydroxamic acid to
the desired cyclic intermediate.
[0431] The acetonitrile was then evaporated in vacuo to give the
crude cyclic intermediate as reddish thick oil. The oil was taken
up in toluene (60 mL) and the reddish mixture was heated to reflux
for 2 hours during which time, the cyclic intermediate released
CO.sub.2 and rearranged to the isocyanate (see below).
##STR00013##
[0432] Step 3 Continued: Conversion of the Isocyanate to the Free
Base
##STR00014##
[0433] The reaction mixture was cooled to 50-60.degree. C. and
(S)-(+)-quinuclidinol (MW 127.18, 28.28 mmol, 3.6 g, 1.5 equiv.)
was added to the mixture as a solid in a single portion. The
mixture was re-heated to reflux for 18 h. After 18 h, an aliquot
was analyzed by HPLC and LC/MS which showed complete conversion of
the isocyanate to the desired product. The reaction mixture was
transferred to a separatory funnel and toluene (25 mL) was added.
The mixture was washed with water (2.times.40 mL) and the water
layers were separated. The combined water layers were re-extracted
with toluene (30 mL) and the water layer was discarded. The
combined toluene layers were extracted with 1N HCl (2.times.60 mL)
and the toluene layer (containing the 0-acyl impurity) was
discarded. The combined HCl layers were transferred to a 500 mL
Erlenmeyer flask equipped with a stir bar. This stirring clear
yellow/reddish orange solution was basified to pH 10-12 by the
dropwise addition of 50% w/w aqueous NaOH. The desired free base
precipitated out of solution as a dirty yellow gummy solid which
could trap the stir bar. To this mixture was added isopropyl
acetate (100 mL) and the mixture was stirred vigorously for 5
minutes when the gummy solid went into isopropyl acetate. The
stirring was stopped and the two layers were allowed to separate.
The yellow isopropyl acetate layer was separated and the basic
aqueous layer was re-extracted with isopropyl acetate (30 mL). The
basic aqueous layer was discarded and the combined isopropyl
acetate layers were dried over anhydrous Na.sub.2SO.sub.4, filtered
into a pre-weighed RB flask and the solvent evaporated in vacuo to
obtain the crude free base as beige to tan solid that was dried
under high vacuum overnight.
[0434] Step 3 Continued: Recrystallization of the Crude Free
Base
[0435] The beige to tan colored crude free base was weighed and
re-crystallized from heptane/isopropyl acetate (3:1, 9.0 mL of
solvent/g of crude free base). The appropriate amount of
heptane/isopropyl acetate was added to the crude free base along
with a stir bar and the mixture was heated to reflux for 10 min
(free base was initially partially soluble but dissolved to give a
clear reddish orange solution when heated to reflux). The heat
source was removed and the mixture was allowed to cool to room
temperature with stirring when a white precipitate formed. After
stirring at room temperature for 3-4 h, the precipitate was
filtered off under hose vacuum using a Buchner funnel, washed with
heptane (20 mL) and dried under hose vacuum on the Buchner funnel
overnight. The precipitate was the transferred to a crystallizing
dish and dried at 55.degree. C. overnight in a vacuum oven. .sup.1H
NMR (400 MHz, CDCl.sub.3) .delta. 8.04-7.83 (m, 2H), 7.20-6.99 (m,
3H), 5.53 (s, 1H), 4.73-4.55 (m, 1H), 3.18 (dd, J=14.5, 8.4 Hz,
1H), 3.05-2.19 (m, 5H), 2.0-1.76 (m, 11H) ppm. .sup.13C NMR (100
MHz, CDCl.sub.3) .delta. 166.38, 165.02, 162.54, 162.8-155.0 (d,
C--F), 130.06, 128.43, 128.34, 116.01, 115.79, 112.46, 71.18,
55.70, 54.13, 47.42, 46.52, 27.94, 25.41, 24.67, 19.58 ppm.
Example 3: Preparation of Crystalline Forms of
(S)-Quinuclidin-3-yl(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbam-
ate Salts
[0436] Crystalline salts of (S)-Quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate may be
formed from the free base prepared as described in Example 23.
[0437] For example, the free base of (S)-Quinuclidin-3-yl
(2-(2-(4-fluorophenyl)thiazol-4-yl)propan-2-yl)carbamate (about 50
mmol) is dissolved IPA (140 ml) at room temperature and filtered.
The filtrate is added into a 1 L r.b. flask which is equipped with
an overhead stirrer and nitrogen in/outlet. L-malic acid (about 50
mmol) is dissolved in IPA (100+30 ml) at room temperature and
filtered. The filtrate is added into the above 1 Liter flask. The
resulting solution is stirred at room temperature (with or without
seeding) under nitrogen for 4 to 24 hours. During this period of
time crystals form. The product is collected by filtration and
washed with a small amount of IPA (30 ml). The crystalline solid is
dried in a vacuum oven at 55.degree. C. for 72 hours to yield the
desired malate salt.
[0438] Crystal forms of other salts, e.g. acid addition salts with
succinic acid or HCl, may be prepared in an analogous manner.
Example 4: In-Vitro GCS Inhibition (Compound 2 and Analogs)
[0439] Inhibition of glucosylceramide synthase activity can be
measured with one or more assays. A first assay is a microsomal
assay that directly measures the conversion of ceramide to
glucosylceramide by HPLC. Microsomes are a source of
glucosylceramide synthase activity in the microsomal assay. A
second assay is a cell based, phenotypic assay that monitors cell
surface expression of the downstream lipid GM3 by antibody mediated
immunofluorescence. Specific protocols are provided below.
[0440] Glucosylceramide Synthase Activity Microsomal Assay:
[0441] An enzyme assay using microsomes as a source of
glucosylceramide synthase activity. Fluorescent ceramide substrate
is delivered to membrane-bound enzyme as a complex with albumin.
After reaction, ceramide and glucosylceramide are separated and
quantitated by reverse-phase HPLC with fluorescence detection.
Enzymatic activity is assessed using a fluorescent labeled
substrate and microsomes as a source of glucosylceramide synthase.
C.sub.6-NBD-Ceramide is complexed with albumin for delivery to
microsomes that are isolated according to the procedure described
below. The final concentration of C.sub.6-NBD-Ceramide in the stock
solution is 0.5 mM; the final concentration of BSA is 0.5 mM.
Separation and quantitation of substrate and product
(glucosylceramide) are achieved by reverse-phase HPLC with
fluorescence detection.
[0442] Preparation of Microsomes from A375 Human Melanoma
Cells;
[0443] Microsomes are isolated from A375 human melanoma cells.
Eight to ten million cells are harvested by trypsinization and
washed with ice cold PBS. Cells are resuspended in ice-cold lysis
buffer containing protease inhibitors. Cell lysate is sonicated on
ice using a probe sonicator. After sonication, the cell lysate is
separated from debris by centrifugation at 10,000 g for 10 minutes
at 4.degree. C. The supernatant is removed and cleared by
additional centrifugation at 100,000 g for 1 hour at 4.degree. C.
The pellet is then resuspended in the lysis buffer, aliquoted and
stored at -80.degree. C. prior to use.
[0444] Glucosylceramide Synthase Assay
[0445] To determine glucosylceramide synthase inhibition,
substrates at 2.times. of their Km (fluorescent ceramide and
UDP-glucose, 3 .mu.M and 4 .mu.M respectively) and microsomes (1:50
dilution) are combined 1:1 and incubated at room temperature for 1
hour in the dark on a plate shaker. The reaction is stopped by the
addition of 150 .mu.L of 100 .mu.M C.sub.8-ceramide in 50% aq.
isopropanol; 10 .mu.L of the final mix is analyzed on HPLC (with
fluorescence detector). The mobile phase is 1% formic acid added to
81% methanol/19% water with flow rate 0.5 mL/min. Fluorescence is
detected with .lamda..sub.ex=470 nm and .lamda..sub.em=530 nm.
Under these conditions, NBD-C.sub.6-GluCer had a retention time of
about 1.7 min and NBD-C.sub.6-Cer elutes from the column after
about 2.1 min. Both peaks are separated from each other and the
baseline and were integrated automatically by the HPLC software.
The percent conversion of substrate to product is used as the
readout for inhibitor testing.
[0446] GM3 Fluorescent-Linked Immunosorbent Assay (FLISA):
[0447] This is a phenotypic assay that measures GM3 expression in
B16 mouse melanoma or C32 human melanoma cells following treatment
with test compounds. Cell surface GM3 expression is determined by
antibody mediated fluorescence.
[0448] Compounds are diluted in media and plated in 384 well plates
in DMSO. B16 and C32 cells are assayed at densities of 20,000
cells/ml and 62,500 cells/ml, respectively, per well. Each
titration curve contains 10 points that are assayed in duplicate on
each test run. The plates are incubated for 48 hours at 37.degree.
C., 5% CO2, and are then washed once with TBS. Anti-GM3 antibody is
added to each well and the plates are then incubated for an
additional one hour at room temperature. Plates are subsequently
washed twice and incubated for an additional hour with the labeled
secondary antibody. Following the final incubation, the plates are
washed twice and the fluorescence at .lamda..sub.ex=D640/20 nm and
.lamda..sub.em=657 nm is detected on a fluorescent reader.
[0449] Assay Results
[0450] Individual assay results of certain exemplified compounds in
these assays are presented in the Table below. The results of the
microsomal assays are expressed as "GCS IC.sub.50", which
represents the concentration of the compound causing 50% inhibition
of glucosylceramide synthase activity. The results of the
cell-based assays are expressed as "GM3 B16 IC.sub.50" or "GM3 C32
IC.sub.50" for the B16 assay and the C32 assay, respectively. These
values represent the concentration of the compound causing 50%
inhibition of GM3 expression on the cell surface.
TABLE-US-00002 Compound GCS IC.sub.50 GM3 B16 GM3 C32 No. (mM)
IC.sub.50 (mM) IC.sub.50 (mM) 1 0.0019 0.0156 0.0021 2 0.0601
0.1068 0.0096 3 0.00414 0.0437 0.00131 4 0.0015 0.0116 0.0008 5
0.0012 0.0193 0.0003 6 0.0028 0.0181 0.0006 7 0.0014 0.0081 0.0004
8 0.0010 0.0075 0.0004 9 0.0014 0.0168 0.0004 10 0.0064 0.0213
0.0022 11 0.0149 0.0819 0.0018 12 0.0203 0.0878 0.0037 13 0.0035
0.0386 0.0007 14 0.0104 0.1096 0.0053 15 0.0267 0.0295 0.0049 16
0.0024 0.0666 0.0016 17 0.4544 0.8786 0.0216 18 0.1480 0.6555
0.0223 19 0.1701 0.1972 0.0426 20 0.3601 0.1065 0.0198 21 0.0506
0.2658 0.0111 22 0.0096 0.0865 0.0032 23 0.0026 0.0477 0.0008
[0451] These comparative results demonstrate that compounds
according to the present disclosure have comparable in-vitro
activity as inhibitors of GCS, and as a result, are expected to
demonstrate similar in-vivo benefits.
Example 5: Clinical Study of Compound 2 in GD-3 Patients
[0452] A 156-week, multi-part, open-label, multinational study of
the safety, tolerability, pharmacokinetics, pharmacodynamics, and
exploratory efficacy of Compound 2 in combination with imiglucerase
in adult patients with Gaucher disease Type 3 stabilized with
imiglucerase was initiated.
[0453] Patients 18 years of age or older with a clinical diagnosis
of GD3 and documented deficiency of acid beta-glucosidase activity
having received treatment with ERT for at least 3 years and with
imiglucerase (Cerezyme) at a stable monthly dose for at least 6
months prior to enrollment were included in the study. Patients
must have reached the following GD therapeutic goals: hemoglobin
level of .gtoreq.11.0 g/dL for females and .gtoreq.12.0 g/dL for
males; platelet count .gtoreq.100 000/mm3; spleen volume <10
multiples of normal (MN), or total splenectomy (provided the
splenectomy occurred >3 years prior to randomization); liver
volume <1.5 MN; and no bone crisis and free of symptomatic bone
disease such as bone pain attributable to osteonecrosis and/or
pathological fractures within the last year. Patients must have GD3
featuring oculomotor apraxia (supranuclear gaze palsy)
characterized by a horizontal saccade abnormality.
[0454] (A) 26-Week Interim Analysis
[0455] An interim analysis was performed when 5 patients had
completed 26 weeks of concurrent treatment with (1) imiglucerase
(Cerezyme from Sanofi Genzyme) under each patient's established
regimen, and (2) Compound 2 administered orally at 15 mg/day in a
single dose. During the study patients were evaluated for safety
and tolerability, CSF and plasma biomarkers (glucosylceramide,
GL-1; glucosylsphingosine, lyso-GL1), pharmacokinetics, markers for
systemic disease (spleen and liver volume measured by magnetic
resonance imaging (MRI), platelet count, hemoglobin levels),
indicia of interstitial lung disease (high resolution pulmonary
computed tomography (CT)), and horizontal saccadic eye
movement.
[0456] At baseline, four patients had mild neurological involvement
and one had moderate neurological involvement, as measured using
the Modified Severity Scoring Tool (mSST; Davies, et al., 2011).
All patients showed evidence of interstitial lung disease based on
chest CT. One patient had anemia, as evidenced by a plasma
hemoglobin level of 10.6 g/dL.
[0457] All patients reported no serious or lasting
treatment-emergent adverse events. The most frequently reported
events were headache and backache, and these were only considered
mild to moderate and transient in duration (likely related to
lumbar puncture performed for CSF testing).
[0458] It was found that Compound 2 effectively cross the
blood-brain barrier in all patients, as demonstrated in the Table
below:
TABLE-US-00003 Compound 2 in plasma Day 1 (N = 5) AUC.sub.0-24, ng
h/mL (mean .+-. SD) 715 .+-. 225 C.sub.max, ng/mL (mean .+-. SD)
48.2 .+-. 19.2 t.sub.max, h (median) 2.00 Compound 2 in CSF Week 4
(N = 5) Week 26 (N = 4) Concentration 4 hours post dose, 4.17 .+-.
0.83 4.71 .+-. 2.50 ng/mL (mean .+-. SD)
[0459] The higher variability of CSF concentration of Compound 2
seen at 26 weeks is attributed to a reduction in exposure of one of
the patients (Patient 5) at weeks 12 to 26 for unknown reasons.
Patient 1 was excluded from the Week 26 CSF determination because
of an error in sample collection.
[0460] It was found that there were significant improvements in
plasma and CSF biomarkers for GD-3 at 26 weeks. At baseline, the
mean (.+-.SD) GL-1 concentration in CSF was 7.1.+-.2.8 ng/mL (range
4.4-11.1 ng/mL), while the mean (.+-.SD) lyso-GL-1 concentration in
CSF was 39.3.+-.22.9 pg/mL (range 20.1-67.6 pg/mL). For comparison,
the healthy GL-1 concentration in CSF is 4.5-5.9 ng/mL, and the
healthy level of lyso-GL-1 in CSF is less than 5.0 pg/mL. At 4
weeks and at 26 weeks, individual reductions in the CSF biomarkers
was found to be as follows (shown as percent reduction from the
baseline CSF concentration):
TABLE-US-00004 Patient Patient Patient Patient Patient Mean 1 2 3 4
5 (n = 4)* GL-1 at 4 -86% -55% -53% -62% -36% -64% weeks GL-1 at 26
-86% -78% -58% -78% +4% -75% weeks Lyso-GL1 -38% -16% -34% -39%
-11% -32% at 4 weeks Lyso-GL1 -52% -42% -41% -57% +94% -48% at 26
weeks *Excludes Patient 5, who showed reduced exposure on PK
analysis of week 12-26 results
[0461] Severity of interstitial lung disease was characterized by
the percent of lung volume affect by ILD as measured by
high-resolution CT in four lung regions (aortic arch, carina, lower
zone L3, lower zone L4). Patients were rated as having severe ILD
(51-100% of lung volume affected), moderate ILD (26-50% of lung
volume affected), mild ILD (1-25% of lung volume affected) or
normal (0% of lung volume showing ILD). All patients showed ILD at
baseline, and 4 out of 5 patients showed regression of ILD after 26
weeks of treatment (patient 5 showed slight progression of
ILD):
TABLE-US-00005 Aortic Arch Carina Zone L3 Zone L4 Left Right Left
Right Left Right Left Right Patient 1 Initial Mild Mild Mild Mild
Norm Norm Mild Mild Week 26 Norm Norm Norm Norm Norm Norm Norm Norm
Patient 2 Initial Mild Mild Mild Mild Mild Norm Mod Mod Week 26
Norm Norm Norm Mild Norm Norm Norm Norm Patient 3 Initial Mild Mild
Mod Mild Severe Mod Severe Severe Week 26 Norm Norm Norm Norm Mild
Mild Mod Mod Patient 4 Initial Norm Mild Norm Norm Norm Norm Norm
Norm Week 26 Norm Norm Norm Norm Norm Norm Norm Norm Patient 5
Initial Mild Mild Mild Mild Mild Mild Mild Mild Week 26 Mild Mild
Mild Mod Mild Mild Mod Mod
[0462] All patients showed an absence of systemic deterioration.
Two patients showed reductions in splenic volume of 10% or more.
There were no clinically meaningful changes in hemoglobin levels.
On average, platelet counts rose 17% from baseline to week 26, and
the three patients with lowest baseline platelet counts showed
individual increase at 26 weeks of 23-42%. Individual patient data
for platelet counts are shown in the table below (shown as 10.sup.9
platelets/L):
TABLE-US-00006 Patient 1 Patient 2 Patient 3 Patient 4 Patient 5
Screening 192 207 259* 313 149 Day 1 186* 192* n/a 307* 127* Week 4
203 243 247 239 154 Week 12 214 294 264 314 180 Week 26 264 236 248
267 173 % Change from +42% +23% -4.3% -13% +36% baseline to 26
weeks *indicates figure used as baseline
[0463] Quantification of horizontal and vertical saccadic eye
movements (HSEM and VSEM, respectively) was performed in all 5
patients. In the five patients, mean peak velocity (PV) of
horizontal rightward 15.degree. saccades was 50.8.degree./s
(+/-8.1.degree./s) at baseline and 47.5.degree./min
(+/-12.6.degree./s) at Week 26; and the mean PV of horizontal
leftward 15.degree. saccades was 44.7.degree. /s
(+/-17.9.degree./s) at baseline and 32.3.degree./s
(+/-15.9.degree./s) at Week 26. A slower velocity implies a more
significant degree of neurological impairment. The mean PV of
horizontal rightward 30.degree. saccades was 77.7.degree./s
(+/-16.4.degree./s) at baseline and 68.1.degree./min
(+/-24.7.degree./s) at Week 26; and the mean PV of horizontal
leftward 30.degree. saccades was 58.7.degree./s (+/-21.5.degree./s)
at baseline and 49.9.degree./s (+/-8.5.degree./s) at Week 26. The
normal range for 150 and 300 horizontal saccades has previously
been reported as >200.degree./s and >400.degree./s
(Bremova-Ertl et al, 2018). HSEM measurements in each of the five
patients are shown in FIGS. 1 and 2. In summary, no clinically
meaningful changes in HSEM were observed over the 26-week treatment
period. As with HSEM, VSEM measurements were stable between
baseline and week 26.
[0464] Four exploratory biomarkers were quantified in plasma, serum
and/or CSF of GD3 patients at week 4 and week 26: chitotriosidase
(CHITO; an enzyme known to be elevated in GD patients) was measured
in CSF and serum; GM3 (a glycosphingolipid marker known to be
elevated in GD patients) was measured in CSF and plasma; and
glycoprotein nonmetastatic melanoma protein B (GPNMB; reportedly a
biomarker of neuropathic GD3) was measured in CSF. The results are
shown in the table below as percent change from baseline at week 4
and week 26 for each parameter:
TABLE-US-00007 Patient Patient Patient Patient Patient 1 2 3 4 5
CHITO CSF- -5% +11% +9% -6% -6% Week 4 CSF- -10% -11% +20% -1% +68%
Week 26 Serum- -11% +8% -7% -12% -1% Week 4 Serum- -43% -55% -1%
+98% -11% Week 26 GM3 CSF- -51% 0% -59% -59% 0% Week 4 CSF- -51% 0%
-59% -59% 0% Week 26 Plasma- -70% -63% -65% -62% -47% Week 4
Plasma- -86% -69% -74% -72% +10% Week 26 GPNMB CSF- -63% +30% +9%
-6% -2% Week 4 CSF- -14% 0% +45% +10% -8% Week 26
[0465] (B) 52-Week Interim Analysis
[0466] A second interim analysis was performed when the first 6
patients had reached 52-weeks of treatment, as described above in
section (A). This analysis included Patients 1-5 as described in
section (A), as well as new Patient 6. All six patients had L444P
(1448T/C) homozygous Gaucher phenotype.
[0467] At 52 weeks, all patients remained enrolled in the study.
There were a total of 30 treatment-emergent adverse events reported
among the six patients, all of which were mild or moderate in
severity, and none of which were considered related to treatment
with Compound 2 or imiglucerase. The events were primarily headache
and back pain, likely related to lumbar puncture.
[0468] Analysis of plasma and CSF concentrations of Compound 2
shows largely comparable values to the values obtained at Week 26.
Patient 5, however, is found to have about 50% lower concentrations
of Compound 2 in plasma and CSF at Week 26, and undetectable
concentrations at Week 52. It is believed that this is due to
either compliance or dosing errors, and therefore, analysis is
repeated without Patient 5's Week 26 and 52 data included. The data
supports the conclusion that a steady state concentration of
Compound 2 is reached in plasma and CSF at or prior to Week 4:
TABLE-US-00008 Compound 2 in plasma Day 1 (N = 6) AUC.sub.0-24, ng
h/mL 729 .+-. 205 (mean .+-. SD) C.sub.max, ng/mL 49.1 .+-. 17.3
(mean .+-. SD) t.sub.max, h (median) 2.00 Compound 2 in Day 1 Week
4 Week 26 Week 52 Plasma (N = 6) (N = 6) (N = 6) (N = 6)
Concentration 2-4 39.7 .+-. 12.6 92.3 .+-. 36.4 102.0 .+-. 49.5
69.8 .+-. 58.3 hours post dose, ng/mL (mean .+-. SD) Excluding
Patient 5 112 84 Day 1 Week 4 Week 26 Week 52 Compound 2 in CSF (N
= 6) (N = 6) (N = 6) (N = 6) Concentration 2-4 <LLOQ 4.56 .+-.
1.20 5.26 .+-. 2.49 4.43 .+-. 3.23 hours post dose, ng/mL (mean
.+-. SD) Excluding Patient 5 6.13 5.32
[0469] At 52 weeks, the data further shows sustained significant
improvements in plasma and CSF biomarkers for GD-3. The results are
similar to those obtained at 26 weeks. Over all six GD3 patients,
plasma and CSF GL-1 and lyso-GL-1 concentrations were as
follows:
TABLE-US-00009 Lyso-GL-1 GL-1 Baseline 52-weeks Baseline 52-weeks
Plasma 29.3 ng/mL 15.2 ng/mL 6.21 .mu.g/mL 1.59 .mu.g/mL
(6.3-159.0) (2.5-46.8) (4.2-8.3) (0.9-2.7) CSF 34.0 pg/mL 17.3
pg/mL 6.36 ng/mL 2.48 ng/mL (20.1-67.6) (5.8-37.4) (4.4-11.1)
(1.0-6.1)
[0470] Thus, at 52-weeks compared to baseline, plasma and CSF
concentrations had changed as follows:
TABLE-US-00010 Lyso-GL-1 (% change) GL-1 (% change) Plasma
Concentration -56.7% -71.6% CSF Concentration -55.9% -55.4%
[0471] In addition, exploratory biomarkers were quantified in CSF
of GD3 patients: ceramide (the precursor of GL-1), chitotriosidase
(CHITO), GM3, and GPNMB. After 52 weeks of treatment, no
significant changes were observed in CSF concentrations of
ceramide, CHITO or GPNMB. Four of the six patients had measurable
concentrations of GM3 in CSF at baseline, and each of these
patients was found to have undetectable GM3 in CSF at 4 weeks, 26
weeks and 52 weeks.
[0472] At 52 weeks, quantification of horizontal and vertical
saccadic eye movements was performed in all six patients in a
similar manner as described in section (A). However, it was
determined that the methodology used to account for noise (e.g.,
caused by blinking or head movements) may have introduced bias into
the results. The method of accounting for noise was therefore
modified, and a set of control criteria were developed to assess
validity of the datasets obtained by the eye movement reader. On
reevaluation of the 26-week saccadic eye movement data, and on
evaluation of the 52-week data, it was found that the level noise
was too high to permit drawing any conclusions from the data.
[0473] In addition, at 52 weeks, 5 of 6 patients showed
improvements in ataxia. The degree of ataxia at baseline and
throughout the study was evaluated by the Scale for Assessment and
Rating of Ataxia (SARA; Schmitz-Hubsch et al. [2006]), which
assesses eight distinct attributes of cerebellar ataxia on a scale
of 0-40. The eight attributes are gait, stance, sitting, speech
disturbance, finger chase, nose-finger test, fast alternate hand
movement, and heel-shin slide. SARA ataxia scoring results for all
six patients is presented in the chart below:
TABLE-US-00011 SARA Cumulative Score At Screening Week 26 Week 52
Patient 1 3.0 1.0 0.0 Patient 2 3.0 2.0 1.5 Patient 3 3.5 0.0 0.0
Patient 4 3.0 5.0 7.5 Patient 5 0.5 0.0 0.0 Patient 6 4.0 2.0 3.0
Average Score 2.83 1.67 2.00 Average Score 2.80 1.00 0.90 excluding
Patient 4
[0474] As shown in the table, five of the six patients were mildly
ataxic at baseline, with the mean cumulative SARA score being 2.8
(SD=1.2). The most common deficits at baseline were gait disorders.
Excluding Patient 5 due to the low level of Compound 2 exposure in
this patient and the patient's substantially normal baseline ataxia
score (only 0.5), then 4 out of 5 patients exhibited an improvement
in ataxia at Week 52 (mean improvement=-0.9; SD=3.2). Patient 4
exhibited an increase in ataxia scoring, with the score at baseline
being 3 and at Week 52, 7.5. It should be noted that this apparent
deterioration was driven almost entirely by a change in the
`stance` scoring parameter (stance score at baseline and Week 26=1;
score at Week 52=5) and that the patient was complaining of left
knee pain at the time of the exam. Additionally, the subject had
injured his left great toe prior to the exam; this injury was
considered resolved 11 days after the exam. Excluding these outlier
effect of Patient 4, treatment with Compound 2 resulted in a
significantly decreased mean SARA Score by Week 26 which was
further slightly improved upon by Week 52.
[0475] The trail making test (TMT) was used to evaluate cognitive
function in the patients. The TMT is one of the most widely used
neuropsychological tests and is included in most test batteries.
The TMT is a diagnostic tool to assess general intelligence and
cognitive dysfunctions (Tombaugh et al. [2004]; Cavaco et al.
[2013]). In part A of the TMT, subjects are asked to connect a
cluster of numbers in ascending order. This task is a combination
of visual search and general visual and motor processing speed.
Part B presents a sequence which alternates between numbers and
letters. Subjects must actively switch between both categories when
connecting them in ascending, but alternating order. Hence, this
task is considered to include an executive function component since
the subject must actively switch between categories while
connecting the symbols (MacPherson et al. [2017]).
[0476] TMT-A evaluates mainly perceptual and psychomotor speed.
TMT-B assesses more specifically mental flexibility and shifting
abilities. TMT B minus TMT A score is used to remove the variance
attributable to the graphomotor and visual scanning components of
TMT A. This derived score reflects the unique task requirements of
TMT B.
[0477] In a study of normative data for TMT A and TMT B in
community-dwelling individuals aged 18-89 years (n=911), mean (SD)
values in the 18-24 years age group (n=155) were 22.9 s (6.9) for
TMT A and 49 s (12.7) for TMT B (Tombauch et al. [2004]). In
contrast, the mean times taken to complete Trail A and Trail B for
patients in the study were 67.8 s (SD=60.3 s) and 193.8 s
(SD=197.0), respectively. At baseline, the mean difference in time
taken to complete Trail B minus Trail A was 126.0 s (SD=142.9 s).
This shows that the GD-3 patients in this study demonstrated some
degree of cognitive dysfunction at baseline.
[0478] At Week 52, the mean time taken to complete Trail A was 56.5
s (SD=55.2 s) and Trail B was 122.7 s (SD=91.8 s). Four of six
patients exhibited reduction in time taken to complete Trail A and
six of six exhibited reduction in time taken to complete Trail B.
Excluding Patient 5 due to the low level of Compound 2 exposure in
this patient, four of five patients exhibited a TMT-A reduction and
five of five patients exhibited a TMT-B reduction.
[0479] At Week 52, 5 of 6 patients exhibited a reduction in the
(TMT B-TMT A) time. Individual results are shown in the table
below.
TABLE-US-00012 Change (%) TMT-A (s)- from Baseline TMT-B (s) At
Screening Week 26 Week 52 to Week 52 Patient 1 13 21 16 +23%
Patient 2 71 37 57 -20% Patient 3 72 56 20 -72% Patient 4 116 (no
data) 66 -43% Patient 5 74 60 72 -3% Patient 6 410 440 166 -60%
Average 126 (n = 6) 123 (n = 5) 66 (n = 6) -29%
[0480] At 52 weeks, the mean difference in time taken to complete
Trail B minus Trail A was 66.2 s (SD=54.3). Excluding Patient 5,
four of five patients exhibited an improvement in Trail B minus
Trial A at Week 52, with a mean improvement of -71.4 s (-31.6%) (SD
99.3 s (37.6%)).
[0481] Neurological function-was further evaluated using functional
magnetic resonance imaging (fMRI). Patient 2 was excluded because
no fMRI data was collected at the Week 52 session. Resting-state
fMRI screening sessions were performed at baseline screening, Week
26, and Week 52 visits. Connectivity estimates from four subjects
(Patients 1, 3, 4 and 5) were entered into second-level analyses as
a "compliant" group. Patient 5 was isolated due to likely
non-compliance with study medication, as described above. Analyses
were performed as described elsewhere (Smith et al. [2009]).
[0482] It was found that the compliant subjects demonstrate an
enhanced connectivity between a more broadly distributed set of
brain regions than the non-compliant subject, with increasing
strength between posterior and anterior aspects as the most
prominent feature. At the anatomic level, compliant subjects
demonstrate a widespread and robust strengthening of connections
between occipital-parietal structures and frontal, temporal and
limbic targets. Connectivity changes in Patient 5 were more modest
and restricted within spatially proximal structures. At the
functional level, enhanced connectivity between default mode and
medial frontal networks is seen in every subject except Patient 5.
This suggests signal within these disparate networks becomes more
coherent, such that brain activity can be more efficiently
transferred between cognitive reserve (posterior) and higher-order
executive functions (anterior). A consistent reciprocal mapping of
resting state networks (RSNs) 2 and 3
("cognition-language-orthography" and "cognition-space") to RSNs 8
and 9 (executive and left frontoparietal) is also evident. The
spatial distribution of connectivity changes is much more focal for
Patient 5, primarily reflecting overlap between medial-frontal and
frontoparietal networks. Both perspectives suggest that patient who
fully complied with the treatment protocol developed greater
coherence between posterior and anterior aspects of the brain, such
that the entire brain becomes amenable to efficient information
transfer. Where apparent, altered connectivity for Patient 5
appears within a narrower set of anterior brain regions and
represents less holistic evidence of therapeutic benefit.
[0483] The results are summarized in the table below. Spatial
analysis of the connectivity between different anatomic regions of
the brain is performed to define a correlation coefficient for
regressed voxelwise mean intensity. The results show that
connectivity between the default mode (resting) network and the
executive function network increased in Patients 1, 3, 4 and 6, but
decreased in Patient 5.
TABLE-US-00013 Patient 1 Patient 3 Patient 4 Patient 5 Patient 6
Change in +0.20 +0.20 +0.20 -0.13 +0.70 Correlation Coefficient
[0484] It was additionally found that two patients experience
spleen volume reductions at Week 52, and mean platelet
concentration increased by 9.3% on average (range -8.2% to +45.3%),
all patients maintaining the therapeutic goal of greater than
120.times.10.sup.9/L platelet count. The increase in mean platelet
concentration was primarily driven by increases in 3 of the 6
patients. There were no clinically meaningful changes in hemoglobin
levels.
Example 6: Pharmacokinetics of Compound 2 in Healthy Human
Volunteers
[0485] Two Phase 1 clinical studies were conducted to assess the
pharmacokinetics, pharmacodynamics, safety and tolerability of
Compound 2 in healthy, human volunteers in the presence and absence
of food. Compound 2 is also known as venglustat.
[0486] Study 1
[0487] Study 1 was a 2-part single-center trial in healthy adult
male volunteers. Part 1 was a double-blind, randomized,
placebo-controlled sequential ascending single-dose study of
Compound 2 for safety, tolerability, and PK. Part 2 was an
open-label, single-cohort, randomized, 2-sequence, 2-period,
2-treatment crossover study of Compound 2 for PK with and without a
high-fat meal.
[0488] Part 1 of the study enrolled and randomized 55 healthy men
(placebo, n=14; 2-, 5-, 15, 25-, 50, and 100-mg doses, n=6 each;
150-mg dose, n=5). Eight healthy men participated in Part 2.
[0489] In Part 1 the subjects were randomized to receive 2, 5, 15,
25, 50, 100, or 150 mg of Compound 2 (L-malic salt form) or
matching placebo on the morning of the first day after at least a
10-hour fast. In Part 2, the subjects were randomized to receive a
single oral dose of 5 mg Compound 2 either while fasting (at least
10 hours before and 4 hours after administration) or 30 minutes
after a standardized high-fat breakfast (.about.815 kcal). After a
7-day washout period, participants were crossed over to the other
condition.
[0490] In Study 1, Part 1, blood was sampled for plasma
concentrations of Compound 2 at the time of study drug
administration (0 hour) and 0.5, 1, 2, 3, 4, 5, 6, 8, 10, 12, 16,
24, 48, 72, and 96 hours post-dose. Urine samples were collected
for analysis of Compound 2 concentrations beginning 2 hours before
study drug administration through 48 hours afterward.
[0491] In Study 1, Part 2, blood was sampled for plasma
concentrations if Compound 2 at 0, 0.5, 1, 2, 3, 4, 5, 6, 8, 10,
12, 16, 24, and 48 hours post-dose.
[0492] From Part 1, it was found that following single oral doses
of 2 to 150 mg doses of Compound 2, maximal plasma concentration
(C.sub.max) occurred at a median time of 3-5.5 hours before plasma
concentrations began to decline exponentially, with a geometric
mean t.sub.1/2 of 28.9 hours. Exposure increased close to
dose-proportionally throughout the dose range: a 75-fold dose
increase resulted in 97.3-, 89.2-, and 85.9-fold increases in
geometric mean C.sub.max, AUC.sub.last, and AUC.sub.inf values,
respectively. PK results are shown in the following table (AUC=area
under the time concentration curve, either to last measurable
concentration or extrapolated to infinity; t.sub.1/2=terminal
half-life; CL/F=apparent total clearance from plasma;
CV=coefficient of variation; SD=standard deviation; t.sub.max=time
to C.sub.max; Vss/F=apparent volume of distribution at steady
state):
TABLE-US-00014 2 mg 5 mg 15 mg 25 mg 50 mg 100 mg 150 mg Parameter
(N = 6) (N = 6) (N = 6) (N = 6) (N = 6) (N = 6) (N = 5) C.sub.max,
ng/mL Mean (SD) 5.7 14.7 53.0 84.4 181 374 529 (1.2) (1.61) (16.7)
(31.8) (56) (38) (109) Geometric 5.6 14.6 50.7 79.9 173 372 520
mean (CV) (21.4) (10.9) (31.5) (37.7) (31) (10.3) (21) t.sub.max,
median 3.50 5.50 3.50 5.00 4.00 3.00 4.00 h (range) (3.00- (4.00-
(2.00- (4.00- (3.00- (2.00- (1.00- 8.00) 8.00) 5.00) 8.00) 6.00)
4.00) 8.00) AUC.sub.last, ng .cndot. h/mL Mean (SD) 214 560 1,830
3,380 6,310 13,000 18,600 (52) (71) (520) (1100) (1880) (2330)
(5480) Geometric 209 556 1,760 3,240 6,070 12,800 18,000 mean (CV)
(24.3) (12.7) (29) (33) (30) (18) (30) AUC.sub.inf, ng .cndot. h/mL
Mean (SD) 243 652 2,070 3,810 7,130 14,400 20,600 (61) (122) (600)
(1,080) (2,320) (3,010) (6,640) Geometric 237 643 1,990 3,690 6,800
14,100 19,900 mean (CV) (25) (19) (29) (28) (33) (21) (32)
t.sub.1/2, h Mean (SD) 29.2 33.3 29.7 30.2 28.9 27.8 26.9 (43)
(8.1) (7.1) (5.5) (5.3) (3.6) (5.7) Geometric 28.9 32.5 29.0 29.8
28.5 27.6 26.4 mean (CV) (14.8) (24.4) (24.0) (18.1) (18.4) (12.8)
(21.3) CL/F, L/h Mean (SD) 6.43 5.86 5.85 5.18 5.75 5.38 5.80
(1.41) (1.01) (1.89) (1.31) (2.01) (1.25) (1.55) Geometric 6.3 5.8
5.6 5.0 5.5 5.3 5.6 mean (CV) (22.0) (17.3) (32.2) (25.3) (34.9)
(23.4) (26.7) V.sub.ss/F, L Mean (SD) 275 274 245 240 239 213 228
(54) (30) (81) (78) (62) (22) (50) Geometric 270 273 233 228 232
212 223 mean (CV) (20) (11) (33) (33) (26) (10) (22)
[0493] From Part 2, it was found that administration of a 5 mg dose
with a high-fat meal had no effect on Compound 2 exposure compared
with fasting conditions. Median t.sub.max was 6.00 hours whether
fed or fasting. Fed/fasted geometric mean ratios were 0.92 and 0.91
for C.sub.max and AUC.sub.last, respectively. Within-subject
variability (i.e., fed vs fasted) accounted for less than half the
total subject variability.
[0494] Study 2
[0495] Study 2 was a single-center, double-blind, randomized,
placebo-controlled, sequential ascending repeated-dose study of the
safety, tolerability, PK, and pharmacodynamics of Compound 2 in
healthy adult male and female volunteers.
[0496] The study enrolled and randomized 36 healthy adults (19 men
and 17 women) (n=9 each to group). The subjects were randomized to
receive once-daily doses of Compound 2 at 5, 10, or 20 mg (provided
as 5-mg capsules of the L-malic salt form) or placebo for 14 days
after at least a 10-hour fast.
[0497] Blood was sampled for plasma concentrations of Compound 2 as
follows: Day 1 at 0, 0.5, 1, 2, 3, 4, 5, 6, 8, 10, 12, and 16 hours
post-dose; On Days 2-5, 8, 11, and 13, at 0 h; On Day 14, at 0.5,
1, 2, 3, 4, 5, 6, 8, 10, 12 hours post-dose; On Days 15-17, at 24,
48, and 72 hours, respectively, after the Day 14 dose. Urine
samples were collected for analysis of Compound 2 concentrations on
Day 1 (0 hours post-dose) and continuously on Day 14 from 0-24
hours post-dose. Pharmacodynamic endpoints (plasma GL-1, GL-3, and
GM3 concentrations) were assessed on Days 1-5, 8, 11, 13, and 14,
at 0 hours post-dose; and on Day 15, at 24 hours after the Day 14
dose.
[0498] It was found that in subjects receiving 5, 10, or 20 mg of
Compound 2 once daily for 14 days, plasma C.sub.max occurred at a
median time of 2-5 hours post-dose on Days 1 and 14. C.sub.trough
values reached a plateau after Day 5. Compound 2 exposure increased
close to dose-proportionally over the dose range of 5-20 mg: this
4-fold dose increase resulted in 3.76- and 3.69-fold increases in
geometric mean C.sub.max and AUC.sub.0-24 values on Day 14,
respectively. PK results from Study 2 are summarized in the
following table:
TABLE-US-00015 Parameter 5 mg (N = 9) 10 mg (N = 9) 20 mg (N = 9)
Day 1 C.sub.max, ng/mL Mean (SD) 18.5 (3.2) 38.5 (7.4) 68.0 (15.7)
Geometric mean 18.2 (17.3) 37.8 (19.3) 66.5 (23.1) (CV) t.sub.max,
median h 5.00 (2.00-8.17) 3.00 (2.00-5.00) 3.07 (2.00-6.00) (range)
AUC.sub.0-24, ng h/mL Mean (SD) 296 (54) 635 (132) 1,100 (211)
Geometric mean 292 (18) 623 (21) 1,080 (19) (CV) Day 14 C.sub.max,
ng/mL Mean (SD) 37.0 (6.4) 89.7 (29.1) 142 (40) Geometric mean 36.5
(17.2) 86.0 (32.5) 137 (28.3) (CV) t.sub.max, median h 3.00
(2.00-6.00) 2.00 (2.00-6.00) 3.00 (2.00-8.00) (range) AUC.sub.0-24,
ng h/mL Mean (SD) 642 (121) 1,550 (464) 2,420 (705) Geometric mean
632 (19) 1,490 (30) 2,340 (29) (CV) C.sub.trough, ng/mL Mean (SD)
19.4 (4.0) 49.9 (19.3) 73.3 (24.4) Geometric mean 19.0 (20.5) 47.5
(38.7) 69.9 (33.2) (CV) t.sub.1/2, h Mean (SD) 29.3 (4.6) 31.3
(3.3) 35.0 (6.3) Geometric mean 29.0 (15.8) 31.2 (10.5) 34.5 (18.0)
(CV) CL.sub.ss/F, L/h Mean (SD) 5.98 (1.17) 5.13 (1.25) 6.58 (1.70)
Geometric mean 5.9 (19.5) 5.0 (24.4) 6.4 (25.8) (CV)
CL.sub.R(0-24), L/h Mean (SD) 1.55 (0.68) 1.49 (0.41) 2.07 (0.58)
Geometric mean NA.sup.a (44.0) 1.4 (27.7) 2.0 (28.0) (CV)
[0499] After 14 once-daily doses of Compound 2, its 24-hour
unchanged urinary excretion fraction (mean fe.sub.0-24) ranged
between 26.3% and 33.1% without any obvious dose-relatedness. Mean
CL.sub.R(0-24) ranged between 1.49 L/h and 2.07 L/h, approximately
3.18-3.86-fold lower than observed plasma CL/F.
[0500] Plasma GL-1, GL-3, and GM3 in placebo recipients remained
similar to baseline throughout, whereas plasma GL-1 and GM3 levels
decreased from baseline time- and dose-dependently across the 3
Compound 2 dose groups, as shown in the following table (Point
estimates of treatment ratios for glucosylceramide (GL-1),
globotriaosylceramide (GL-3), and GM3 ganglioside (GM3) on Day 15
in the repeated ascending dose study):
TABLE-US-00016 90% Confidence Parameter Comparison Estimate
Interval GL-1 5 mg vs placebo 0.39 0.29-0.50 10 mg vs placebo 0.32
0.25-0.42 20 mg vs placebo 0.23 0.17-0.30 GL-3 5 mg vs placebo 0.61
0.47-0.79 10 mg vs placebo 0.69 0.53-0.89 20 mg vs placebo 0.67
0.51-0.89 GM3 5 mg vs placebo 0.56 0.45-0.70 10 mg vs placebo 0.49
0.39-0.60 20 mg vs placebo 0.40 0.32-0.50
[0501] Maximal sustained effects on GL-1 occurred on Day 11 in the
5- and 10-mg groups and by Day 8 in the 20-mg group. Mean
calculated GL-1 reductions from baseline at Day 15 were 41.9%,
69.6%, and 74.6% in the respective 5-, 10- and 20-mg groups. GL-1
values were below the lower limit quantification (LLOQ) at baseline
in 1 5-mg Compound 2 recipient and at Day 15 in 3, 5, and 9
subjects in the 5-, 10-, and 20-mg groups, respectively.
[0502] Maximal sustained GM3 decreases occurred across all Compound
2 dose groups starting on Day 13. Mean Day 15 plasma GM3 levels
were 42.7%, 49.4%, and 57.8% of baseline for the 5-, 10-, and 20-mg
dose groups, respectively. GM3 was below the LLOQ at Day 15 in 1
and 2 subjects in the 10- and 20-mg dose groups, respectively.
[0503] Plasma GL-3 also decreased with time in all Compound 2 dose
groups, but variable and low baseline GL-3 values relative to LLOQ
limited mean calculated GL-3 reductions. In the placebo, 5-, 10-,
and 20-mg dose groups, GL-3 values were below LLOQ in 1, 3, 1, and
6 subjects, respectively, at baseline and in 4, 9, 7, and 9
subjects, respectively, at Day 15.
[0504] Mean estimated plasma GL-1 reductions from baseline (90% CI)
attributable to Compound 2 C.sub.trough in the 5, 10, and 20 mg
dose groups (19.0, 47.5, and 69.9 ng/mL, respectively) were 67.0%
(54.4-79.7%), 74.4% (63.7-85.2%), and 76.3% (64.8-87.8%),
respectively.
[0505] Conclusions
[0506] In these studies, Compound 2 exposure in healthy subjects
(C.sub.max and AUC) was close-to-dose-proportional when
administered as single doses ranging from 2-150 mg or as repeated,
once-daily doses ranging from 5-20 mg for 14 days. Compared with
fasting, a high-fat meal had no effect on exposure in subjects who
received a single 5-mg dose. With repeated once-daily doses from
5-20 mg, steady state was achieved within 5 days; neither age nor
gender affected accumulation. Pharmacodynamically, repeated
once-daily doses of Compound 2 reduced plasma concentrations of
GL-1 and GM3 in a time- and dose-dependent manner, consistent with
Compound 2-mediated GCS inhibition, although baseline levels of
GL-3 were too low to be useful as a pharmacodynamic biomarker. The
dose-dependent GL-1 reduction corroborated the intended mechanism
of action of Compound 2: inhibition of GL-1 formation from ceramide
by GCS.
[0507] In all studies, safety profile was assessed by monitoring
treatment-emergent adverse events (TEAEs) through 10 days after
last dose of study medication, including serious adverse events
[SAEs]), ECG monitoring, laboratory values, and physical
examinations. There were no deaths, SAEs, severe TEAEs, or TEAEs
leading to study discontinuation in any of the studies.
[0508] No clinically relevant hematologic or biochemical
abnormalities were reported in any of the studies. Vital signs
showed no relevant changes from baseline in any of the studies. ECG
parameters showed no relevant changes in the single ascending dose
and food effect studies; in the multiple ascending dose study no
ECG parameters changed statistically significantly from average
baseline versus placebo in recipients of Compound 2 at any dose. It
is to be understood that while the invention has been described in
conjunction with the above embodiments, that the foregoing
description and examples are intended to illustrate and not limit
the scope of the invention. Other aspects, advantages and
modifications within the scope of the invention will be apparent to
those skilled in the art to which the invention pertains.
Example 7: Clinical Study of Compound 2 in Fabry Disease
Patients
[0509] Method
[0510] A three-year open-label investigation of Compound 2 in young
classic Fabry disease patients was conducted in order to evaluate
the long-term safety, pharmacodynamics and exploratory efficacy of
Compound 2 in adult male Fabry patients. Eleven subjects enrolled
in the study, and seven subjects completed all aspects of the
study. All subjects were males diagnosed with classic Fabry disease
confirmed by genotype and residual alpha-galactosidase activity
below detection levels (9 of the 11 had nonsense mutation in the
GLA gene). All subjects had lyso-GL3 levels in plasma of at least
65 ng/mL and had no prior Fabry disease-specific treatment. The
median age of subject was 24 years (range 19-37).
[0511] Patients were administered a daily oral dose of 15 mg of
Compound 2. The clearance of GL-3 deposits in the skin was
monitored by taking biopsies at weeks 12, 26, 52 and 156, which
were evaluated semi-quantitatively by light microscopy (focusing on
skin capillary endothelial cells). Each sample was scored
independently by 3 pathologists for the presence of GL-3 inclusions
on a four-point scale, and rated as 0 (no/trace), 1 (mild), 2
(moderate), or 3 (severe) according to Eng et al., N. Engl. J. Med.
345:9-16 (2001). A single score per patient per time point was
derived by taking the score rated by the majority of the three
pathologists. If a majority score could not be derived, then the
median score was used (resulting in some fractional scores). Plasma
samples were also analyzed for GL-3, lyso-GL-3, GL-1 and GM3 at
baseline and weeks 12, 26, 52 and 156. Pain scores and abdominal
symptoms were analyzed at baseline and weeks 12, 26, 52, 104 and
156 using the SF-36 scoring protocol.
[0512] Patients were assessed using the Short Form-36 (SF-36)
questionnaire on numerous visits from baseline to week 156. This is
a 36-item questionnaire used to measure 8 various aspects of health
(vitality, physical functioning, bodily pain, general health
perceptions, physical role functioning, emotional role functioning,
social role functioning and mental health). The score for each of
the eight aspects ranges from 0 (maximum disability) to 100 (no
disability), and thus, higher scores indicate good health
condition. In addition, gastrointestinal symptoms, including
abdominal pain, abdominal distention, and bowl movements, were
assessed using a modified version of the inflammatory bowel
severity scoring system. Particular questions asked as part of
these assessments included: (1) whether the patient suffered
abdominal pain within the last ten days, (2) what was the severity
of abdominal pain suffered in the last ten days using a 0 (no pain)
to 100 (very severe pain) scale, and (3) on how many days within
the last ten days did the patient have abdominal pain.
[0513] Results
[0514] By week 156, five patients showed a 1-point reduction in
skin GL-3 score, two patients had complete clearance of GL-3
inclusions, one patient had no change, and one patient lacked
samples. Over the course of the 156-week study, mean plasma GL-1
levels dropped by 69%, mean plasma GM3 levels dropped by 60%, mean
plasma GL-3 levels dropped by 77%, and mean plasma lyso-GL3 levels
dropped by 52%. Plasma GL-1 and GM3 showed a very rapid drop within
the first 2-4 weeks of treatment. All four measures showed a
sustained maintenance of reduced plasma load which was largely
stabilized by week 52.
[0515] Plasma and urine data are summarized in the table below:
TABLE-US-00017 Change from baseline measurement at: 26 weeks 52
weeks 104 weeks 156 weeks Parameter Measured: (n = 9) (n = 7) (n =
7) (n = 7) Plasma GL-3 (.mu.g/mL) -3.62 .mu.g/mL -5.06 .mu.g/mL
-6.32 .mu.g/mL -6.97 .mu.g/mL Plasma lyso-GL-3 (ng/mL) -30.99 ng/mL
-37.10 ng/mL -39.84 ng/mL -48.13 ng/mL Plasma GL-1 (.mu.g/mL) -3.26
.mu.g/mL -3.58 .mu.g/mL -3.70 .mu.g/mL -3.23 .mu.g/mL Plasma GM3
(.mu.g/mL) -10.77 .mu.g/mL -8.84 .mu.g/mL -9.92 .mu.g/mL -8.12
.mu.g/mL Urine GL-3 (mg per mmol -0.25 mg/mmol -0.20 mg/mmol -0.18
mg/mmol -0.18 mg/mmol urine creatinine) (n = 8) (n = 7) (n = 7) (n
= 7)
[0516] These results demonstrate that the Compound 2 administered
at 15mg/day consistently reduces the levels of GL-1, lyso-GL-1 and
GM3 in the body in a generally progressive manner.
[0517] In addition, data from a previously completed
placebo-controlled phase 3 trial of agalsidase beta (Fabrazyme) was
analyzed for comparison (see Eng et al., N. Eng. J. Med., 345:9
(2001). For comparison to agalsidase beta, the historical control
arm was patients treated with agalsidase beta in the phase 3 trial,
and change in plasma GL-3 was compared at multiple time points up
to three years. Entry criteria and baseline characteristics were
similar between the two studies. To strengthen the comparison,
patients receiving Compound 2 were matched with phase 3 study
patients based on propensity scores using baseline variables of
age, plasma GL-3, gender, UPCR (<500 mg/g versus 500-1000 mg/g
versus >1000 mg/g), and eGFR (<80 versus .gtoreq.80
mL/min/1.73m.sup.2). 11 patients receiving Compound 2 were matched
to 19 patients for the placebo comparison and to 28 patients for
the agalsidase beta comparison. All patients in all three groups
were male and demonstrated elevated plasma GL-3, UPCR of <500
mg/g, and eGFR .gtoreq.80 mL/min/1.73 m.sup.2. Mean ages were
similar across the three groups. The comparison shows that
treatment with Compound 2 for 26 weeks led to a significant
reduction in plasma GL-3 compared to placebo, -3.62 .mu.g/mL versus
-1.06 .mu.g/mL (P<0.0001). While treatment with Compound 2
yielded a similar reduction in plasma GL-3 at 52 weeks compared to
agalsidase beta, at 104 weeks and at 156 weeks, the plasma GL-3
reduction from Compound treatment was significantly greater
(p=0.0351 at 104 weeks; p=0.0081 at 156 weeks). Plasma GL-3 levels
after 156 weeks were 1.90 .mu.g/mL for patients treated with
Compound 2, compared to 4.44 .mu.g/mL for patients treated with
agalsidase beta.
[0518] Detailed results for the skin GL-3 inclusion scores are
shown in the tables below (score of 0 indicates no GL-3
inclusions):
TABLE-US-00018 Endothelial Cells, Superficial Vessels Number of
Patients in Each Change Category Skin GL-3 Score (changes compared
to baseline skin score) at: Change 12 weeks 26 weeks 52 weeks 156
weeks Score 1 dropped to 0 2/6 (33%) Score 1 unchanged 4/9 (44%)
4/9 (44%) 3/6 (50%) 1/6 (17%) Score 1 raised to 2 1/9 (11%) 1/9
(11%) Score 2 dropped to 1 3/9 (33%) 3/9 (33%) 2/6 (33%) 3/6 (50%)
Score 2 unchanged 1/9 (11%) 1/9 (11%) 1/6 (17%)
TABLE-US-00019 Endothelial Cells, Deep Vessels Number of Patients
in Each Change Category Skin GL-3 Score (changes compared to
baseline skin score) at: Change 12 weeks 26 weeks 52 weeks 156
weeks Score 1 unchanged 1/9 (11%) 1/9 (11%) 1/6 (17%) Score 2
dropped to 0.5 1/6 (17%) Score 2 dropped to 1 2/9 (22%) 2/9 (22%)
3/6 (50%) 3/6 (50%) Score 2 dropped to 1.5 1/6 (17%) Score 2
unchanged 6/9 (67%) 6/9 (67%) 2/6 (33%) 1/6 (17%)
TABLE-US-00020 Smooth Muscle Cells, Deep Vessels Number of Patients
in Each Change Category Skin GL-3 Score (changes compared to
baseline skin score) at: Change 12 weeks 26 weeks 52 weeks 156
weeks Score 1.5 unchanged 2/9 (22%) Score 1.5 raised to 2 2/9 (22%)
1/5 (20%) 1/6 (17%) Score 2 dropped to 0.5 Score 2 dropped to 1 1/6
(17%) Score 2 dropped to 1.5 1/6 (17%) Score 2 unchanged 7/9 (78%)
7/9 (78%) 4/5 (80%) 3/6 (50%)
TABLE-US-00021 Perineurium Cells Number of Patients in Each Change
Category Skin GL-3 Score (changes compared to baseline skin score)
at: Change 12 weeks 26 weeks 52 weeks 156 weeks Score 1 unchanged
1/6 (17%) Score 1 raised to 2 1/9 (11%) 1/9 (11%) 1/6 (17%) Score 2
dropped to 1 1/6 (17%) Score 2 dropped to 1.5 1/6 (17%) 1/6 (17%)
Score 2 unchanged 8/9 (89%) 8/9 (89%) 3/6 (50%) 4/6 (67%)
[0519] In addition to scoring GL-3 skin inclusions by light
microscopy, the fraction of the volume of endothelial cell
cytoplasm occupied by GL-3 inclusions was estimated using
point-counting of electron microscopic images by a masked reader.
Images from at least 50 superficial endothelial cell capillaries
were obtained using electron microscopy at 7500.times.
magnification. Two-sided t tests were used to evaluate differences
between baseline and post-treatment values at each time point. The
results are shown in the table below:
TABLE-US-00022 Baseline 3 months 6 months 3 years (N = 9) (N = 9)
(N = 8) (N = 6) Volume fraction 0.29 .+-. 0.03 0.29 .+-. 0.06 0.23
.+-. 0.04 0.18 .+-. 0.03 (volume of inclusions/total cytoplasmic
volume) Change from baseline -1.9% -21.1% -38.7% P value vs.
baseline 0.74 0.001 0.001
[0520] These results demonstrate that the Compound 2 administered
at 15 mg/day consistently reduces the levels of GL-3 inclusions in
the skin in a generally progressive manner. Results were generally
more pronounced for superficial vessel endothelial compared to deep
vessel endothelial cells and other skin tissues.
[0521] Seven of nine patients had improved overall bodily pain
scores at week 26 (SF-36), while three of six patients had improved
overall bodily pain scores at week 156 (SF-36). Among the patients
with gastrointestinal pain at baseline, the severity of the pain
(abdominal pain) decreased in four out of five at week 26, and in
four out of four at week 156. The number of days with
gastrointestinal pain was reduced in five of five patients at week
26 and in three of four patients at week 156.
[0522] Detailed results for abdominal pain measures are shown in
the tables below.
TABLE-US-00023 Abdominal Pain Number of Patients Reporting "Yes" to
Abdominal Pain in 10-Days Preceding Visit 4 12 26 52 104 156
Baseline weeks weeks weeks weeks weeks weeks 6/11 4/11 2/10 3/9 2/7
2/7 2/7 (55%) (36%) (20%) (33%) (28%) (28%) (28%) Abdominal Pain
Severity Score in 10-Days Preceding Visit (mean value, 0-100 scale)
4 12 26 52 104 156 Baseline weeks weeks weeks weeks weeks weeks (n
= 6) (n = 4) (n = 2) (n = 3) (n = 1) (n = 1) (n = 2) 52.50 29.75
40.00 31.33 21.00 21.00 15.00 Number of Days within Preceding 10
Days with Abdominal Pain (mean value) 4 12 26 52 104 156 Baseline
weeks weeks weeks weeks weeks weeks (n = 6) (n = 4) (n = 2) (n = 3)
(n = 3) (n = 2) (n = 2) 3.83 2.50 3.50 3.00 0.70 0.50 0.20
[0523] These results demonstrate that the Compound 2 administered
at 15mg/day consistently reduces abdominal pain and discomfort in
the body in a generally progressive manner.
[0524] In addition, where features or aspects of the invention are
described in terms of Markush groups, those skilled in the art will
recognize that the invention is also thereby described in terms of
any individual member or subgroup of members of the Markush
group.
[0525] All publications, patent applications, patents, and other
references mentioned herein are expressly incorporated by reference
in their entirety, to the same extent as if each were incorporated
by reference individually. In case of conflict, the present
specification, including definitions, will control.
* * * * *