Discovery of (R)-4-Cyclopropyl-7,8-difluoro-5-(4-(trifluoromethyl) phenylsulfonyl)-4,5-dihydro-1 H -pyrazolo[4,3- c ]quinoline (ELND006) and (R)-4-Cyclopropyl-8-fluoro-5-(6-(trifluoromethyl)pyridin-3-ylsulfonyl)-4, 5-dihydro-2 H -pyrazolo[4,3- c ]quinoline (ELND007): Metabolically stable γ-Secretase Inhibitors that selectively inhibit the production of amyloid-β over notch

Article abstract of DOI:10.1021/jm301741t

Herein, we describe our strategy to design metabolically stable γ-secretase inhibitors which are selective for inhibition of Aβ generation over Notch. We highlight our synthetic strategy to incorporate diversity and chirality. Compounds 30 (ELND006) and 34 (ELND007) both entered human clinical trials. The in vitro and in vivo characteristics for these two compounds are described. A comparison of inhibition of Aβ generation in vivo between 30, 34, Semagacestat 41, Begacestat 42, and Avagacestat 43 in mice is made. 30 lowered Aβ in the CSF of healthy human volunteers.

Full text of DOI:10.1021/jm301741t

Article
pubs.acs.org/jmc
Discovery of (R)‑4-Cyclopropyl-7,8-difluoro-5-
(4-(trifluoromethyl)phenylsulfonyl)-4,5-
dihydro‑1H‑pyrazolo[4,3‑c]quinoline (ELND006) and
(R)‑4-Cyclopropyl-8-fluoro-5-(6-(trifluoromethyl)pyridin-3-ylsulfonyl)-
4,5-dihydro‑2H‑pyrazolo[4,3‑c]quinoline (ELND007): Metabolically
Stable γ‑Secretase Inhibitors that Selectively Inhibit the Production
of Amyloid‑β over Notch
Gary Probst,*,† Danielle L. Aubele,† Simeon Bowers,† Darren Dressen,† Albert W. Garofalo,†
Roy K. Hom,† Andrei W. Konradi,† Jennifer L. Marugg,† Matthew N. Mattson,† Martin L. Neitzel,†
Chris M. Semko,† Hing L. Sham,†,‡ Jenifer Smith,† Minghua Sun,† Anh P. Truong,† Xiaocong M. Ye,†
Ying-zi Xu,† Michael S. Dappen,‡ Jacek J. Jagodzinski,‡ Pamela S. Keim,‡ Brian Peterson,‡ Lee H. Latimer,‡
David Quincy,‡ Jing Wu,‡ Erich Goldbach,§ Daniel K. Ness,§ Kevin P. Quinn,§ John-Michael Sauer,§
Karina Wong,§ Hongbin Zhang,§ Wes Zmolek,§ Elizabeth F. Brigham,∥ Dora Kholodenko,∥ Kang Hu,∥
Grace T. Kwong,∥ Michael Lee,∥ Anna Liao,∥ Ruth N. Motter,∥ Patricia Sacayon,∥ Pamela Santiago,∥
Christopher Willits,∥ Frederique Bard,⊥ Michael P. Bova,⊥ Susanna S. Hemphill,⊥ Lam Nguyen,⊥
́
́
Lany Ruslim,⊥ Kevin Tanaka,⊥ Pearl Tanaka,⊥ William Wallace,⊥ Ted A. Yednock,†,‡,§,∥,⊥
and Guriqbal S. Basi⊥
†Department of Medicinal Chemistry, Elan Pharmaceuticals, 180 Oyster Point Boulevard, South San Francisco, California 94080, United States
‡Department of Process and Analytical Chemistry, Elan Pharmaceuticals, 180 Oyster Point Boulevard, South San Francisco,
California 94080, United States
§Department of Lead Finding, Drug Disposition, and Safety Evaluation, Elan Pharmaceuticals, 180 Oyster Point Boulevard,
South San Francisco, California 94080, United States
∥Department of In Vivo Pharmacology, Elan Pharmaceuticals, 180 Oyster Point Boulevard, South San Francisco, California 94080,
United States
⊥Department of Biology, Elan Pharmaceuticals, 180 Oyster Point Boulevard, South San Francisco, California 94080, United States
S
* Supporting Information
ABSTRACT: Herein, we describe our strategy to design
metabolically stable γ-secretase inhibitors which are selective
for inhibition of Aβ generation over Notch. We highlight our
synthetic strategy to incorporate diversity and chirality. Com-
pounds 30 (ELND006) and 34 (ELND007) both entered
human clinical trials. The in vitro and in vivo characteristics for
these two compounds are described. A comparison of inhibi-
tion of Aβ generation in vivo between 30, 34, Semagacestat 41,
Begacestat 42, and Avagacestat 43 in mice is made. 30 lowered Aβ in the CSF of healthy human volunteers.
With escalating healthcare costs numbering in the hundreds of
billions of dollars to treat and care for AD patients, a disease-
modifying treatment is urgently needed.
The etiology of AD is not fully understood. However, hints
concerning the origins may be found in the pathology. The pathology
INTRODUCTION
■
Alzheimer’s disease (AD) is an insidious form of senile dementia
that causes one to lose their identity. Tens of millions of elderly
people worldwide are afflicted with AD.1 A massive progressive
loss of neurons in the hippocampus and cortex results in
impaired cognitive and functional ability. Current therapies,
N-methyl-D-aspartic acid antagonists and acetylcholine esterase
inhibitors, to treat those afflicted with AD are merely palliative.2
Received: November 29, 2012
Published: May 28, 2013
© 2013 American Chemical Society
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evinces itself with the presence of extraneuronal aggregation of
plaques composed of β-amyloid peptides (Aβ)3 and intracellular
neurofibrillary tangles (NFTs),4 aggregates of aberrantly hyper-
phosphorylated tau proteins, which are common to numerous
neurodegenerative diseases. The amyloid hypothesis5 states that
Aβ initiates a complex pathological cascade including the genera-
tion of NFTs, leading to neuronal death.
Aβ-peptides are derived by the sequential proteolytic cleavage
of the β-amyloid precursor protein (APP) by two aspartic acid
proteases, referred to as β- and γ-secretase, respectively.6 Inhibitors
of either protease offer attractive candidates as potentially disease-
modifying therapies for people afflicted with AD.7 γ-Secretase
processes a myriad of substrates8 in addition to C99, most notably
Notch, raising concerns about mechanism-based side-effects (e.g.,
goblet cell hyperplasia in the gastrointestinal tract)9 due to a lack
of selectivity.
Our objective was to avoid Notch-related toxicity by
developing γ-secretase inhibitors with a sufficient therapeutic
index. Thus, these inhibitors would be required to selectively
inhibit Aβ formation over Notch cleavage. The magnitude of
a selectivity ratio that would predict an acceptable in vivo
therapeutic index could not be determined a priori, and therefore
the strategy for the target product profile was to minimally
exceed the selectivity ratios for Semagacestat, 41, a relatively
nonselective γ-secretase inhibitor that had already entered
clinical development, by at least a half log interval. In this
paper, enzyme and cellular APP:Notch ratios are reported and
were used to guide SAR. The selectivity ratios for the bench-
mark 41 in these assays were 0.41-fold and 15-fold, respectively
(Table 4), and therefore the target product profile was to exceed
these values by approximately 3-fold. Acute biomarker data,
short-term-repeat dose studies in the mouse, and chronic repeat-
dose data collected in the Sprague−Dawley rat and cynomolgus
monkey retrospectively confirmed this product profile to be
adequate for clinical development, i.e., these data provided
evidence that brain Aβ could be continuously decreased in the
absence of Notch-related toxicity in vivo.
An active site inhibitor would not confer selectivity so we
sought an allosteric inhibitor. Existing literature suggested that an
allosteric inhibitor of γ-secretase was plausible.10 An additional
requirement was to incorporate favorable pharmacokinetic pro-
perties into these inhibitors to avoid peak-to-trough drug levels
that could rise above toxic exposures or fall below efficacious
concentrations, once-a-day dosing, and sustained levels of
exposure to reduce Aβ over a 24 h period.
RESULTS, DISCUSSION, AND CHEMISTRY
■
Three series of pyrazole/sulfonamide inhibitors 1−3 were under
development (Table 1). All three series were comparable in
terms of their in vitro selectively for inhibiting Aβ production
over Notch and low metabolic stability. When dosed in rat PK
screens,11 both compound 2 (67 mL/min/kg) and compound 3
(34 mL/min/kg) resulted in systemic clearances of greater than
60% of hepatic blood flow with oral bioavailabilities of 6% and
1%, respectively. Because of its low oxidative stability, compound
1 was not tested in vivo, however, these results demonstrated a
connectivity with low metabolic stability in vitro and high
clearance in vivo. Preventing the oxidation of aliphatic carbons
can be extremely challenging, however, mitigating the oxidation
of aryl rings can be more facile. Thus, the quinoline series 112
ostensibly offered the lowest hurdle to prepare metabolically
stable inhibitors. This series had a single aliphatic carbon (the site
of oxidation), whereas both the piperidine 213 and bicyclic 314
series bore multiple aliphatic carbons as potential sites for
oxidation (see refs 14b and c for early development of the series).
The initial synthesis of the quinolines was unreliable and offered
only limited access to functionality (Scheme 1). A reductive
amination between the β-keto ester 4 with 4-fluoroaniline
provided 5, which was sulfonylated to provide 6 and hydrolyzed
to afford 7. Formation of the acid chloride followed by
Friedel−Crafts cyclization yielded 8. This cyclization of 7 to 8
Table 1. SAR for the Three Series
a
b
See ref 14c for detailed experimental procedures. m = mouse, r = rat, d =dog, h = human.
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Scheme 1. Initial Synthetic Route
Scheme 2. Retrosynthetic Analysis
was capricious under multiple conditions. Alternatively, 5 could
be hydrolyzed and cyclized, but then the sulfonylation was
problematic especially in cases where the stereogenic substituent
was sterically encumbered by substituents larger than methyl. The
cyclization chemistry was not compatible with electron deficient
aryl rings (e.g., pyridines), further limiting this approach. Lastly,
incorporation of the pyrazole with reliable chemistry furnished 9.
This synthetic route was inefficient. The pyrazole was to be
maintained as a constant, and thus should be incorporated early
in the synthesis instead of the final step. Areas optimal for SAR were
installed in the initial steps of the synthesis rather than the latter
stages. Nevertheless, this route did furnish some initial compounds.
An investment into the development of a reliable synthetic
route that offered the opportunity to incorporate an eclectic mix
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Scheme 3. Synthesis of the Key Intermediate
Scheme 4. Completion of Final Target via the New Synthetic Route
of functionality in the latter stages of the synthesis was necessary
for the quinoline series to be successful. The pyrazole was to be
maintained as constant (any substitution at the C−H position of
the pyrazole was detrimental to the potency), while the other
regions of the template were to be varied. A retrosynthetic
analysis (Scheme 2) revealed that disconnecting the sulfonamide
nitrogen from the aryl ring in 10 would allow for the aliphatic
and sulfonamide region to be incorporated separately from the
aryl portion. The former areas could be installed via a Grignard
addition to a sulfonylimine 12, which in turn could be derived
from the condensation between a sulfonamide and an aldehyde
13. Biaryl rings can be built efficiently and reliably via transition-
metal mediated cross-coupling methodologies. Thus, a protected
pyrazole bearing two distinct handles 15 was required as the
key intermediate. Ethyl 3-amino-1H-pyrazole-4-carboxylate 16
served as the commercially available starting material.
This key intermediate 15 was prepared in multigram quantities
in five steps from commercially available starting material 16
(Scheme 3). Conversion of the amino group into an iodide
followed by protection of the pyrazole with (2-(chloromethoxy)-
ethyl)trimethylsilane afforded 17 and 18 as a mixture of
regioisomers15 that were separable by a laborious chromato-
graphic separation. This material could be advanced, however,
as a mixture through all of the subsequent chemistry without
incident. Saponification of the ester 18 followed by conversion to
the aldehyde yielded 15.16 An NOE enhancement between the
pyrazole C−H and hemiaminal protons was observed to confirm
the regiochemistry of 15 (see Supporting Information).
A Suzuki−Miyaura cross-coupling17 between 15 and commer-
cially available 2-bromo-4,5-dilfuorobenzeneboronic acid 19
installed the aryl portion (Scheme 4). Condensation of the
aldehyde 20 with a sulfonamide followed by Grignard addition to
the sulfonylimine afforded the cyclization precursor 21. A copper-
mediated reaction18 cleanly yielded the cyclized material 22 that
was subsequently deprotected to complete the synthesis of 23.
This copper-mediated reaction was extremely robust in that clean
starting material was not required, it worked with heterocyclic aryl
rings, and was complete within minutes. Hydrodebromination
gave rise to the major undesired byproduct, but by placing the
reaction mixture into a preheated oil bath this byproduct could
largely be suppressed.
Identification of the metabolites was crucial in order to make
rational modifications to improve the metabolic stability.19
Metabolite identification studies with uridine 5′-diphosphate-
glucuronic acid (UDP-GA) supplemented rat and human liver
microsomses, as well as from bile cannulated rat studies for both
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Table 2. Quinoline Series SAR
a
b
See ref 14c for detailed experimental procedures. m = mouse, r = rat, d = dog, h = human.
compounds 1 and 3, demonstrated that the pyrazole underwent
direct N-glucuronidation. Glucuronidation is a low affinity, high
capacity pathway leading to high clearance in vivo which needed
to be addressed. Additionally, cleavage of the glucuronide in
the gastrointestinal tract could lead to enterohepatic circulation
of the inhibitor, resulting in prolonged plasma concentration−
time profile and increased variability between patients. Initial
studies indicated that incorporation of a fluoride at C8, as in 24,
attenuated the N-glucuronidation. Metabolite identification
studies of 24 demonstrated that the methylene of the quinoline
ring was susceptible to oxidation (Table 2). A methyl group at
C4, as in 26, was installed to block the metabolism. Virtually no
improvement was observed in the in vitro microsomal stability,
but fortuitous introduction of the methyl in 26 led to an increase
in potency (0.62 vs 22 nM) and an improvement in the
biochemical Notch/Aβ selectivity (23 vs 7.3) relative to 24.
Metabolite identification studies of 26 surprisingly revealed
that the major site of oxidation was now the aryl ring of the
sulfonamide. To prevent this oxidation and retain the potency,
electron withdrawing lipophilic functional groups20 were placed
at the para position of the sulfonamide. Introduction of a
fluoride, as in 27, in place of the chloride resulted in only a slight
improvement in the oxidative stability but resulted in reduced
formation of reactive metabolite in trapping studies with
glutathione. Virtually no improvement in the oxidative stability
was observed with the stronger electron withdrawing trifluor-
omethyl group as in 28. The major site of oxidation of 27
appeared relocated onto the quinoline ring system with a minor
metabolite was located on the aryl ring of the sulfonamide.
We hypothesized that the methyl was the most likely site of
oxidation. Cyclopropane has a ring strain of 27 kcal/mol, and
incorporation of a single sp2 center increases the ring strain to
41 kcal/mol.21b,c Also the C−H bond strength of cyclopropane
(106 kcal/mol) is greater than ethane (101 kcal/mol).21d,e Thus,
replacing the methyl group with a cyclopropyl ring could pre-
vent hydrogen atom extraction, thereby mitigating the oxidation
of the aliphatic region. At this juncture, we began to employ
the second synthetic route (Scheme 4) and used commercially
available 2-bromo-4,5-difluoroboronic acid 19 in order to
expedite the production of targets rather than 2-bromo-4-
fluororoboronic acid 36 (Scheme 5) which had to be synthesized.
The SAR indicated that the additional C7 fluoride 25 would not
be detrimental. The cyclopropyl moiety in 29 enhanced the
potency and improved the metabolic stability in rodents but not
in humans. At this point, we postulated that the site of metabolic
oxidation must have relocated once again, this time back to the
aryl ring of the sulfonamide. A trifluoromethyl group 30 was
used to prevent the oxidation of aryl ring of the sulfonamide and
maintain potency. Furthermore, glutathione adducts were not
observed with this substitution. To provide additional support
for the hypothesis that the cyclopropyl group prevented the
oxidation, the isopropyl derivative 31 was prepared and
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Scheme 5. Asymmetric Synthesis of 34
demonstrated very little stability toward oxidative metabolism.
Recall that early in the development of the template, the fluoride
at C8 was necessary to mitigate glucuronidation. Removal of the
fluorides on the quinoline ring system 32 demonstrated that the
C8 fluoride was not necessary to improve the stability against
glucuronidation. The cyclopropyl group (or any alkyl group) became
the key to abrogate glucuronidation. A single fluoride as in 33 was
necessary for stability against oxidation in phase 1 metabolism.
On the basis of these data and the pharmacological data that
follows, compound 30 was advanced as far as possible into
preclinical development to uncover any hurdles for clinical
development for this series because it was the first inhibitor in
this genre. Both the in vitro and in vivo profiles for 30, (R)-4-
cyclopropyl-7,8-difluoro-5-(4-(trifluoromethyl)phenylsulfonyl)-
4,5-dihydro-1H-pyrazolo[4,3-c]quinoline (ELND006),22 sup-
ported IND, enabling advancement into human clinical trials
(ELND numbered compounds are nominated for development).
Inhibitor 34, (R)-4-cyclopropyl-8-fluoro-5-(6-(trifluoromethyl)-
pyridin-3-ylsulfonyl)-4,5-dihydro-2H-pyrazolo[4,3-c]quinoline
(ELND007),22 was optimized for both potency and water
solubility and eventually advanced into human clinical trials as
well. Details regarding the pharmacokinetic/pharmacodynamic
(PK/PD) relationship, in vivo Notch selectivity, and other
pharmacokinetic properties will be published separately.
the (S)-(−)-tert-butanesulfinamide23 with the aldehyde 37
afforded the sulfinimine to which cyclopropylmagnesium bromide
was added in 9:1 dr as measured by HPLC. Removal of the sulfinyl
group afforded the amine, which was sulfonylated with 38 to yield
the cyclization precursor 39. Cyclization followed by deprotection
afforded a 9:1 mixture of enantiomers of 34 that were separable via
chiral HPLC.
The pharmacological and drug properties for 30 and 34 were
suitable for advancing into human clinical trials. The overall
chemotype demonstrates favorable physiochemical properties
for an orally available central nervous system (CNS) compound
with range of lipophilicity (2 < log P < 4), topographical polar
surface area (<70 Å2), hydrogen bond donors (0−1), and
moderate molecular weight (≤450 amu).24 Compound 34
demonstrated a lower octanol−water partition coefficient of
3.65 compared to 3.91 for 30 as measured by the shaker-flask
method.11 This reduced lipophilicity translated to better aqueous
solubility for 34 (∼100 μg/mL) compared to 30 (∼3 μg/mL).11
In addition, compound 34 demonstrated a high rate of passive
permeability (204 nm/s) in wild-type Madin−Darby canine kidney
(MDCK) cells compared to the moderate rate of permeability for
compound 30 (88 nm/s), relative to the high permeability control,
propranolol (178 nm/s). Furthermore, both compounds were
negligible substrates for permeability-glycoprotein (P-gp) efflux in
MDR1 transfected MDCK cells. These properties25 afforded good
to excellent oral bioavailabilities in rats for 34 (92%) and 30 (30%)
following administration of an aqueous suspension formulated in
2% methylcellulose and 0.5% Tween 80. Furthermore, their
metabolic stabilities conferred a moderately low systemic clearance
of 22 mL/min/kg (40% of hepatic blood flow (QH)) and
23 mL/min/kg (42% QH) for 30 and 34, respectively, in the rat.
The preclinical studies required copious quantities of 30 and
34. An asymmetric synthesis, which was incorporated into the
initial synthetic route, could potentially double the overall
yield of the route (Scheme 5) and was implemented immediately
upon the discovery of this genre of inhibitors. The commercially
available starting material 35 was used to generate the boronic
acid 36 that was cross-coupled with 15 to afford 37. Condensing
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Table 3. Compound Levels 3 h After Administration
Table 5. Efficacy Comparison 3 h After Administration
a
dose in mice for efficacy studies in Table5
dose
a,b
brain and plasma concentration
(% reduction of cortical Aβx-40 in mice)
1 mg/kg 3 mg/kg 10 mg/kg 30 mg/kg
inhibitor
1 mg/kg
3 mg/kg
10 mg/kg
30 mg/kg
30
30
34
41
42
43
32
34
5
48
56
19
18
10
61
61
31
20
19
64
62
47
33
[brain] 3 h (μM)
0.12
1.0
0.52
4.5
2.1
7.2
62
[brain unbound] 3 h (nM)
[brain] AUC(0−3 h) (μM)
[plasma] 3 h (μM)
18
b
b
c
0.64
0.031
0.28
0.19
34
2.1
7.0
ND
3.0
27
ND
0.19
1.7
0.90
8.1
7
35
a
b
c
[plasma unbound] 3 h (nM)
[plasma] AUC(0−3 h) (μM)
See ref 14c for detailed experimental procedures. *p < 0.05. Not
determined.
0.68
0.68
ND
[brain] 3 h (μM)
0.045
0.63
0.24
0.014
0.87
0.14
0.22
3.1
1.0
14
2.6
36
the other agents at 30 mg/kg) owing to their greater metabolic
stability in rodents as well as potency. The effective unbound
brain concentration (ECu25) estimated to achieve a 25% reduction
of brain Aβx-40 in mice was 18-fold lower for 34 (0.034 nM) than
30 (0.62 nM). Sustained reduction of cortical Aβx-40 above the
25% threshold over a 24 h period was achieved in mice with a single
oral 30 mg/kg dose of 30 with a maximal effect of 82% reduction at
6 h post administration (the companion PK/PD study in mice for
34 was not conducted).19 The ability to reduce central Aβ robustly
over a 24 h period with a moderate single daily dose of inhibitor
ratified our strategy of focusing on improving the metabolic stability
to enhance the exposure. Additionally, the respective in vitro
biochemical and cellular selectivity ratios for inhibition of
Notch cleavage over Aβ production for 30 (19 and 78) and
34 (17 and 71) were nearly identical and comparable to both
43 (12 and 59) and 42 (6.9 and 45) but superior to 41
(0.41 and 15).
[brain unbound] 3 h (nM)
[brain] AUC(0−3 h) (μM)
[plasma] 3 h (μM)
0.92
0.079
4.9
4.1
1.2
74
11
3.5
220
12
[plasma unbound] 3 h (nM)
[plasma] AUC(0−3 h) (μM)
0.57
4.7
a
b
See ref 14c for detailed experimental procedures. ND = not
determined.
Both 30 and 34 also readily partitioned into the CNS (Table 3),
which was consistent with their favorable ratio of unbound plasma
to unbound brain ratio of 1 and 4, respectively, as determined by
equilibrium dialysis for 30 (fu,plasma = 0.0090, fu,brain = 0.0086) and
34 (fu,plasma = 0.062, fu,brain = 0.014).26 The CNS partitioning was
further corroborated by the significant pharmacodynamic effect in
the reduction of brain Aβx-40, which plateaued at approximately
60% for all of the compounds in this genre by 3 h following a single
oral dose. Both 30 and 34 were superior to 41,27 Begacestat 42,28
and Avagacestat 4329 (Table 4) when comparing the cortical
Aβx-40 reduction in mice 3 h after a single oral dose (Table 5).
Both 30 and 34 achieved similar Aβ reduction, but at a dose 30-fold
lower than other agents (compare 30 and 34 at 1 mg/kg to
The magnitude of a selectivity ratio that would predict an
acceptable in vivo therapeutic index could not be determined a
priori, and therefore the strategy for the target product profile
was to minimally exceed the selectivity ratios for 41, a relatively
nonselective γ-secretase inhibitor that had already entered clinical
Table 4. Competitors Clinical γ-Secretase Inhibitors
a
b
See ref 14c for detailed experimental procedures. m = mouse, r = rat, d = dog, h = human.
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10 μm. Separation of the enantiomers is indicated for each compound
(vide infra).
development, by at least a half log interval. It was impractical
to assess in vivo selectivity across a large set of compounds to
fully model in vitro to in vivo relationships, but for a subset of
analogues of interest for potential advancement into preclinical
development, acute rodent models were used to explore this
translation, i.e., could reductions in CNS Aβ be achieved in the
absence of peripheral Notch inhibition in vivo? First, an acute
biomarker study was conducted in mice, comparing plasma
exposures that decrease brain Aβ with those that decrease the
Notch transcription factor Hairy and Enhancer of Split
homologue-1 (HES-1) in thymus (Table 6). Plasma compound
1H NMR data were recorded on a 400 MHz Bruker, 300 MHz Bruker,
or 300 MHz Varian Gemini. Chemical shifts are reported in parts per
million (ppm) downfield relative to tetramethylsilane (TMS) or to
proton resonances resulting from incomplete deuteration of the NMR
solvent (d scale). The 2D ROSEY NMR was taken on a 700 MHz
Bruker.
HMRS data were collected using a LC/MS analysis on a Q-TOFMS.
Stock solutions (20 mM) for all standard compounds prepared in
DMSO were diluted in water to 5 μM for positive mode mass spectro-
metric analysis. This typically produced maximum ion abundances on
the order of one to five million ions/scan, although for data processing,
spectra with ion abundances of 500000 ions/scan were averaged.
A tripleTOF 5600 quadrupole time-of-flight hybrid mass spectrometer
(AB Sciex, Foster City, CA, USA) equipped with an LC system
consisting of a LEAP PAL HTC auto sampler and an Agilent 1290
Infinity LC pump (Agilent, Santa Clara, CA) was used for LC/MS
analysis. The mass spectrometer was operated in the positive ionization
mode with a spray voltage of 5.5 kV. Ion source gas 1, ion gas source 2,
and curtain gas were set at 40, 45, and 25 psi, respectively, and the source
temperature was maintained at 500 °C. Samples (5 μL) were introduced
to the mass spectrometer via the HPLC system using an Agilent EC-C18
column (2.1 mm × 75 mm, 2.7 μm) under isocratic conditions (50:50
acetonitrile/water with 0.1% formic acid) at a flow rate of 0.8 mL/min.
Full scan mass spectra were acquired over the range of 120−900 m/z
with an accumulation time of 0.25 s. Total acquisition time was 2 min
per sample, and all four time-to-digital converter channels were used for
detection. The tripleTOF instrument was calibrated by an integrated
calibration delivery system (AB Sciex) using the manufacturer’s positive
calibration solution, with an injection flow rate of 300 μL/min. These
calibrant ions were introduced via an orthogonal atmospheric pressure
chemical ionization source in the presence of mobile phase flow. Mass
spectral data acquisition and processing were performed using Analyst
TF 1.5 and Peak View 1.1.1.2 software.
Mass spectra were recorded on an Agilent series 1100 mass spectro-
meter connected to an Agilent series 1100 HPLC. Retention times
(tR) are given in minutes. Three methods used for purity analysis: (1)
Method 1 = 20% [B], 80% [A] to 70% [B], 30% [A] gradient in
10.0 min, then hold, at 1.5 mL/min, where [A] = 0.1% trifluoroacetic
acid in water, [B] = 0.1% trifluoroacetic acid in acetonitrile on a
Phenomenex Luna C18 (2) 4.6 mm × 3 cm column, 3 μm packing,
210 nm detection, at 35 °C. (2) Method 2 = 50% [B], 50% [A] to 95%
[B], 5% [A] gradient in 10.0 min, then hold, at 1.5 mL/min, where
[A] = 0.1% trifluoroacetic acid in water, [B] = 0.1% trifluoroacetic acid in
acetonitrile a Phenomenex Luna C18 (2) 4.6 mm × 3 cm column, 3 μm
packing, 210 nm detection, at 35 °C. (3) Method 3 = 10% [B], 90%
[A] to 40% [B], 60% [A] gradient in 10.0 min, then hold, at 1.5 mL/min,
where [A] = 0.1% trifluoroacetic acid in water, [B] = 0.1% trifluoroacetic
acid in acetonitrile on a Phenomenex Luna C18 (2) 4.6 mm × 3 cm
column, 3 μm packing, 210 nm detection, at 35 °C.
Table 6. Changes in Thymic HES-1 mRNA 3 h Post
Administration
dose
a
(% change of thymic HES-1 mRNA in mice)
inhibitor
1 mg/kg
3 mg/kg
10 mg/kg
30 mg/kg
b
c
d
e
30
34
ND
−17
−14
−25
d
f
f
f
+11
−21
−44
−46
a
b
See reference 14c for detailed experimental procedures. Not
determined. *p < 0.05. Not statistically significant. *p < 0.001.
c
d
e
f
*p < 0.01.
concentrations were assessed over a range of doses and plasma
concentrations associated with a 25% reduction (EC25) in brain
Aβ were compared to EC25 for thymus HES-1 reduction. To
achieve these reductions, 30 afforded a 57-fold difference in
plasma levels.30 Second, 7-day studies in mice dosed twice daily
were conducted, examining brain Aβ concentrations biochemi-
cally and Notch-responsive tissues histologically (e.g., goblet
cell hyperplasia in intestine, atrophy of thymus). Both studies
demonstrated a broader therapeutic index relative to the
benchmark comparator. Furthermore, longer-term repeat dose
studies in the cynomolgus monkey and Sprague−Dawley rat
confirmed that acceptable margins of safety were achievable, even
after chronic administration. In these repeat dose studies, the
cell-based IC50 for Notch inhibition was the most informative in
predicting plasma concentrations associated with adverse effects.
Retrospectively, in vivo studies supported the target product
profile for Notch selectivity as being acceptable for clinical
development.
In summary, we were able to improve the metabolic stability
through rational design based on our understanding of physical
properties of functional groups. Fortunately, these perturbations
improved the potency and did not adversely affect the Notch
selectivity. A single 30 mg oral dose of 30 crystalline material in
a gelatin capsule lowered Aβ in the CSF of healthy human
volunteers by 31−46% 5 h after administration, however, it was
discontinued due to an adverse liver event unrelated to its
mechanism of action. The observed liver effects in the clinic were
not predicted by repeat-dose toxicity studies up to six months of
dosing in the rat and one year of dosing in the cynomolgus
monkey. 34 was advanced into human clinical trials but was
discontinued due to a concern about the potential for an adverse
liver event.
The specific optical rotation values for samples were obtained using
a Rudolf Instruments, Model Autopol V polarimeter. The polarimeter
was zeroed with 95% ethanol, and a sample was tested at 589 nm. The
analysis was performed per USP <781S>.
5-(4-Chlorophenylsulfonyl)-4,5-dihydro-1H-pyrazolo[4,3-c]-
1
quinoline (1). Synthesized in a Similar Manner to Scheme 1. H
NMR (300 MHz, DMSO-d6) δ 7.68−7.62 (m, 2H), 7.46−7.41 (m, 3H),
7.32 (d, J = 8.2 Hz, 2H), 7.16 (d, J = 8.8 Hz, 2H), 4.93 (s, 2H). HMRS:
calcd for C16H12ClN3O2S (M + H)+, 346.04115; found, 346.04057.
HPLC purity (method 1): tR = 4.08, 100%. LCMS m/z 346.0 and 348.0
(M + H)+. HPLC purity (method 2): tR = 0.87, 100%. LCMS m/z 346.0
and 348.0 (M + H)+. HPLC purity (method 3): tR = 9.79, 100%. LCMS
m/z 346.0 and 348.0 (M + H)+.
(4R,6S)-5-((4-Chlorophenyl)sulfonyl)-4,6-diethyl-4,5,6,7-tet-
rahydro-2H-pyrazolo[4,3-c]pyridine (2).31 1H NMR (400 MHz,
CDCl3) δ 7.70 (d, J = 8.8 Hz, 2H), 7.41 (s, 1H), 7.35 (d, J = 8.8 Hz, 2H),
4.96 (t, J = 7.2 Hz, 1H), 4.17 (q, J = 7.2 Hz, 1H), 2.52 (d, J = 16.0 Hz,
1H), 2.24 (dd, J = 16.0, 6.4 Hz, 1H), 1.96 (m, 1H), 1.79 (m, 1H), 1.65
(m, 1H), 1.45 (m, 1H), 1.19 (t, J = 7.2 Hz, 3H), 0.98 (t, J = 7.2 Hz, 3H).
EXPERIMENTAL METHODS
■
All reagents and solvents were used as purchased from commercial
sources. All reactions were run under nitrogen. All of the final
compounds were purified on a Varian Prepstar HPLC with a Varian
Prostar detector using on a Phenomenex Luna C18 column of either of
the following sizes 250 mm × 21.20 mm, 5 μm, or 250 mm × 50.00 mm,
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HMRS: calcd for C16H20ClN3O2S (M + H)+, 354.10375; found,
354.10350. HPLC purity (method 1): tR = 5.18, 100%. LCMS m/z 354.1
and 356.1 (M + H)+. HPLC purity (method 2): tR = 0.85, 100%. LCMS
m/z 354.1 and 356.1 (M + H)+. HPLC purity (method 3): tR = 10.21,
100%. LCMS m/z 354.1 and 356.1 (M + H)+.
(4R,8S)-10-((4-Chlorophenyl)sulfonyl)-4,5,6,7,8,9-hexahy-
dro-2H-4,8-epiminocycloocta[c]pyrazole (3).32 1H NMR (400
MHz, CDCl3) δ 7.65 (d, J = 8.8 Hz, 2H), 7.48 (s, 1H), 7.36 (d, J =
8.8 Hz, 2H), 5.28 (broad s, 1H), 4.48 (t, J = 5.6 Hz, 1H), 2.96 (dd, J =
17.6, 7.6 Hz, 1H), 2.61 (d, J = 17.6 Hz, 1H), 1.93 (broad m, 2H), 1.61
(broad m, 3H), 1.30 (qt, J = 14.0, 4.0 Hz, 1H). HMRS: calcd for
C15H16ClN3O2S (M + H)+, 338.07245; found, 338.087212. HPLC
purity (method 1): tR = 2.74, 99%. LCMS m/z 338.0 and 340.1 (M +
H)+. HPLC purity (method 2): tR = 0.49, 100%. LCMS m/z 338.0 and
340.1 (M + H)+. HPLC purity (method 3): tR = 7.38, 100%. LCMS m/z
338.0 and 340.1 (M + H)+.
tert-Butyl 3-((4-Fluorophenyl)amino)butanoate (5). To a
1,2-dichloroethane solution (146 mL) of 4-fluoroaniline (4.06 g,
36.5 mmol) was added tert-butyl acetoacetate 4 (6.05 mL, 36.5 mmol)
and glacial acetic acid (3.0 mL, 47.5 mmol). Sodium triacetoxyborohy-
dride (11.6 g, 54.8 mmol) was added in portions, and the reaction was
stirred overnight at room temperature. Saturated aqueous sodium
bicarbonate (250 mL) was slowly added, and the two-phase solution was
well stirred. The separated organic portion was washed with saturated
aqueous sodium bicarbonate (1 × 100 mL), water (1 × 100 mL), and
brine (1 × 20 mL), dried over anhydrous magnesium sulfate, filtered,
and concentrated to yield 5 (9.03 g, 98%) as an oil. 1H NMR (400 MHz,
CDCl3) δ 6.88 (t, J = 8.8 Hz, 2H), 6.55 (dd, J = 8.8, 4.4 Hz, 2H), 3.80
(sextet, J = 6.4 Hz, 1H), 2.49 (dd, J = 14.8, 5.6 Hz, 1H), 2.33 (dd, J =
14.8, 6.8 Hz, 1H), 1.44 (s, 9H), 1.25 (d, J =6.4Hz, 3H). HPLC(method1):
tR = 3.28. LCMS m/z 254.1 (M + H)+. HPLC (method 3): tR = 6.93.
LCMS m/z 254.1 (M + H)+.
tert-Butyl 3-(4-Chloro-N-(4-fluorophenyl)phenylsulfonamido)-
butanoate (6). To a solution of tert-butyl 3-((4-fluorophenyl)amino)-
butanoate 5 (3.08 g, 12.2 mmol) in pyridine (35 mL) at 0 °C was added
4-chlorobenzenesulfonyl chloride (8.98 g, 42.6 mmol) portionwise. The
reaction was warmed to room temperature and then heated at 70 °C for
5.5 h then cooled to room temperature. Dimethylaminopropylamine
(40 mL) was added to the reaction mixture, which was then stirred
overnight. The reaction mixture was then concentrated and the residue
taken up in ethyl acetate (120 mL). The organic phase was washed with
cold 1N aqueous hydrogen chloride (6 × 100 mL), water (1 × 100 mL),
saturated aqueous sodium bicarbonate (3× 100 mL), brine (1 × 100 mL),
dried over anhydrous magnesium sulfate, filtered, and concentrated to
yield 6 (5.2 g, 99%) as an oil. 1H NMR (400 MHz, CDCl3) δ 7.65 (dd,
J = 6.8, 1.6 Hz, 2H), 7.43 (dd, J = 6.8, 1.6 Hz, 2H), 7.02 (broad m, 4H),
4.90 (sextet, J = 6.8 Hz, 1H), 2.33 (dd, J = 15.2, 7.6 Hz, 1H), 2.12 (dd, J =
15.2, 7.2 Hz, 1H), 1.46 (s, 9H), 1.13 (d, J = 6.8 Hz, 3H). HPLC (method 1):
tR = 10.14. LCMS m/z 372.0 and 374.0 (M − C4H7)+, and 450.1 and 452.1
(M + Na)+. HPLC (method 2): tR = 4.43. LCMS m/z 372.0 and 374.0
(M − C4H7)+, and 450.1 and 452.1 (M + Na)+.
residue taken up in diethyl ether, filtered through a plug of glass wool, and
concentrated to give the acid chloride (4.09 g, 99%). A dichloromethane
(20 mL) solution of the acid chloride was added dropwise over a 20 min
period to a chilled (−5 °C) suspension of aluminum chloride (1.81,
13.6 mmol) in dichloromethane (20 mL). After stirring for 0 °C for 1 h, the
reaction was warmed to room temperature and stirred overnight. The
reaction mixture was then chilled in an ice bath to which was added 10%
aqueous hydrogen chloride (15 mL) dropwise over 15 min. The organic
portion was washed with 10% aqueous hydrogen chloride (2 × 50 mL),
water (1 × 20 mL), saturated aqueous sodium bicarbonate (3 × 50 mL),
brine (1 × 20 mL), dried over anhydrous magnesium sulfate, filtered, and
concentrated. The crude product was then dissolved in tetrahydrofuran
(40 mL) to which was added saturated aqueous sodium bicarbonate
(40 mL). This mixture was stirred for 7 h and then concentrated. The
residue was taken up in water and ethyl acetate. The aqueous layer was
extracted with ethyl acetate (4 × 30 mL), and the combined ethyl acetate
extracts washed with brine (1 × 20 mL), dried over anhydrous magnesium
sulfate, filtered, and concentrated to the give the crude product, which was
purified by flash chromatography eluting with 8:1 hexanes:ethyl acetate to
give 8 (980 mg, 26%). 1H NMR (400 MHz, CDCl3) δ 7.91 (dd, J = 9.2,
4.8 Hz, 1H), 7.66 (dd, J = 8.4, 3.2 Hz, 1H), 7.57 (dd, J = 6.8, 1.6 Hz, 2H),
7.45 (dd, J = 6.8, 1.6 Hz, 2H), 7.36 (ddd, J = 9.2, 7.6, 3.2 Hz, 1H), 4.91
(quintuplet d, J = 7.2, 2.0 Hz, 1H), 2.40 (dd, J = 18.0, 5.6 Hz, 1H), 2.30 (dd,
J = 18.0, 2.0 Hz, 1H), 1.31 (d, J = 7.2 Hz, 3H). HPLC (method 1): tR =
7.80, LCMS m/z 354.0 and 356.0 (M + H)+. HPLC (method 2): tR = 2.38,
LCMS m/z 354.0 and 356.0 (M + H)+.
5-((4-Chlorophenyl)sulfonyl)-8-fluoro-4-methyl-4,5-dihydro-
2H-pyrazolo[4,3-c]quinoline (9). Dimethylformamide−dimethyl
acetal (5.2 mL) solution of 1-((4-chlorophenyl)sulfonyl)-6-fluoro-2-
methyl-2,3-dihydroquinolin-4(1H)-one 8 (922 mg, 2.61 mmol) was
heated at 100 °C for 3 h. HPLC/MS analysis of the reaction mixture
determined no starting material remained, so the reaction was cooled to
room temperature and poured into ethyl acetate and water (50 mL
each). The organic portion was washed with water (3 × 50 mL) and
brine (1 × 20 mL), dried over anhydrous magnesium sulfate, filtered,
and concentrated to give a reddish foam which was used directly in
the next reaction. To an ethanol:glacial acetic acid solution (25:1 v/v,
12 mL) of the intermediate (982 mg, 2.40 mmol) was added hydrazine
hydrate (582 mL, 12.0 mmol). The reaction mixture was stirred
overnight at room temperature, concentrated, and the residue taken up
in ethyl acetate (20 mL). The organic phase was washed with saturated
aqueous sodium bicarbonate (3 × 25 mL) and brine (1 × 20 mL), dried over
anhydrous magnesium sulfate, filtered, and concentrated. The crude product
was purified by flash chromatography eluting with 2:1 hexanes:ethyl acetate
to give 9 as a yellow solid. Chiral separation of the enantiomers of 9 provided
26. See 26 for details concerning chiral separation and characterization.
3-Iodo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-4-
carbaldehyde (15). Ethyl 3-iodo-1-((2-(trimethylsilyl)ethoxy)-
methyl)-1H-pyrazole-4-carboxylate 18 (22.50 g, 56.8 mmol) and 3N
aqueous sodium hydroxide (50.0 mL, 150 mmol) in dioxane (200 mL)
was stirred for four days. The solution was brought to pH = 3 with 10%
aqueous hydrogen chloride and extracted with methylene chloride. The
combined organic extracts were dried over anhydrous magnesium
sulfate, filtered, and concentrated under reduced pressure to yield
20.91 g (100%) of 3-iodo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-
pyrazole-4-carboxylic acid as a light-yellow solid. The material was
used in the subsequent reaction without further purification. HPLC
(method 1): tR = 6.22. LCMS m/z 369.0 (M + H)+ and 391.0
(M + Na)+. Neat borane dimethylsulfide ca. 10 M (25 mL, 250 mmol)
was slowly added to 3-iodo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-
pyrazole-4-carboxylic acid (20.91 g, 56.8 mmol) in tetrahydrofuran
(250 mL) at 0 °C. After stirring for 30 min, the solution was placed into
a preheated oil bath at 50 °C. After stirring for 18 h, the solution was
cooled to 0 °C and ice was added. After stirring for 30 min, saturated
aqueous ammonium chloride was added and stirred for an additional
30 min. The solution was extracted with methylene chloride, and the
combined organic extracts were dried over anhydrous magnesium
sulfate, filtered, and concentrated under reduced pressure to yield
19.50 g (97%) of 3-iodo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-
pyrazol-4-yl)methanol as a light-yellow liquid. The material was used
3-(4-Chloro-N-(4-fluorophenyl)phenylsulfonamido)butanoic
acid (7). To a solution of tert-butyl 3-(4-chloro-N-(4-fluorophenyl)-
phenylsulfonamido)butanoate 6 (5.19 g, 12.13 mmol) in dichloro-
methane (10 mL) at 0 °C was added trifluoroacetic acid (35 mL). The
reaction mixture was allowed to warm to room temperature, stirred
overnight, and then concentrated to give 7 (4.51 g, 100%) as a solid.
1H NMR (400 MHz, CDCl3) δ 7.65 (dd, J = 6.8, 1.6 Hz, 2H), 7.43 (dd,
J = 6.8, 1.6 Hz, 2H), 7.02 (d, J = 6.4 Hz, 4H), 4.92 (sextet, J = 6.8 Hz,
1H), 2.50 (dd, J = 15.6, 8.0 Hz, 1H), 2.31 (dd, J = 15.6, 6.4 Hz, 1H), 1.20
(d, J = 6.8 Hz, 3H). HPLC (method 1): tR = 6.16. LCMS m/z 372.0 and
374.0 (M + H)+. HPLC (method 2): tR = 1.25. LCMS m/z 372.0 and
374.0 (M + H)+.
1-((4-Chlorophenyl)sulfonyl)-6-fluoro-2-methyl-2,3-dihydro-
quinolin-4(1H)-one (8). To an ice chilled solution of 3-(4-chloro-N-
(4-fluorophenyl)phenylsulfonamido)butanoic acid 7 (3.91 g, 10.52 mmol)
in dichloromethane (45 mL) and dimethylformamide (4 drops) was added
oxalyl chloride (3.67 mL, 42.10 mmol) dropwise. The resulting solution
was stirred at −5 °C for 15 min and then warmed to room temperature
and stirred for 3 h. The reaction mixture was then concentrated and the
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in the subsequent reaction without further purification. HPLC (method 1):
tR = 5.72. LCMS m/z 355.0 (M + H)+. Dess−Martin periodinane (13.80 g,
32.5 mmol) was added to a heterogeneous mixture of (3-iodo-1-
((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)methanol (7.19 g,
20.3 mmol) and sodium bicarbonate (17.44 g, 208 mmol) in methylene
chloride (200 mL). After stirring for 24 h, the heterogeneous mixture was
diluted with saturated aqueous sodium sulfite and water and then extracted
with diethyl ether. The combined organic extracts were dried over
anhydrous magnesium sulfate, filtered, and concentrated. The residue was
flash chromatographed on silica using 99:1 49:1, 24:1, 23:2, 22:3, 21:4, and
4:1 hexanes:ethyl acetate as the eluant to yield 7.15 g (100%) of 15 as a
white solid. 1H NMR (300 MHz, CDCl3) δ 9.71 (s, 1H), 8.00 (s, 1H), 5.44
(s, 2H), 3.58 (m, 2H), 0.90 (m, 2H), −0.03 (s, 9H). HPLC (method 1):
tR = 7.51. LCMS m/z 294.9 (M + H)+. HPLC (method 2): tR = 2.23.
LCMS m/z 294.9 (M + H)+.
N-((3-(2-Bromo-4,5-difluorophenyl)-1-((2-(trimethylsilyl)ethoxy)-
methyl)-1H-pyrazol-4-yl)(cyclopropyl)methyl)-4-(trifluoromethyl)-
benzenesulfonamide (21). Titanium(IV) isopropoxide (3.0 mL,
10.2 mmol) was added to a solution of 3-(2-bromo-4,5-difluorophenyl)-
1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-4-carbaldehyde
(slightly impure 1.00 g, < 2.40 mmol) and 4-(trifluoromethyl)-
benzenesulfonamide (970 mg, 4.31 mmol) in tetrahydrofuran (10 mL).
After stirring for 18 h, the solution was diluted with methylene chloride
and then water, and the heterogeneous solution was filtered through a
pad of Celite. The filtrate was extracted with methylene chloride, the
combined organic extracts were dried over anhydrous magnesium sulfate,
filtered, and concentrated. The residue was flash chromatographed on
silica using 9:1, 4:1, 7:3, and 3:2 hexanes:ethyl acetate as the eluant to
afford a mixture of products containing N-((3-(2-bromo-4,5-difluor-
ophenyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)-
methylene)-4-(trifluoromethyl)benzenesulfonamide which was used
without further purification. A 1.0 M solution of cyclopropyl magnesium
bromide in tetrahydrofuran (20.0 mL, 20.0 mmol) was slowly added along
the walls of the flask to a solution of N-((3-(2-bromo-4,5-difluorophenyl)-
1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)methylene)-4-
(trifluoromethyl)benzenesulfonamide in tetrahydrofuran (20 mL) at
−78 °C. After stirring for 30 min, the dry ice/acetone bath was removed
and saturated aqueous ammonium chloride was added followed by water
until the heterogeneous mixture became homogeneous. The biphasic
solution was extracted with methylene chloride. The combined organic
extracts were dried over anhydrous magnesium sulfate, filtered, and
concentrated. The residue was flash chromatographed on silica using 9:1,
4:1, 7:3, and 3:2 hexanes:ethyl acetate as the eluant to yield 1.13 g (>60%
yield over two steps) of 21 as a yellow oil. 1H NMR (400 MHz, CDCl3) δ
7.45 (dd, J = 9.2, 7.2 Hz, 1H), 7.11 (dd, J = 10.4, 8.0 Hz, 1H), 5.37 (d, J =
10.4 Hz, 1H), 5.33 (d, J = 10.4 Hz, 1H), 4.88 (d, J = 6.0 Hz, 1H), 3.76 (dd,
J = 8.8, 6.0 Hz, 1H), 3.59 (dd, J = 8.8, 7.6 Hz, 2H), 1.01 (m, 1H), 0.95 (t,
J = 10.4 Hz, 2H), 0.37 (broad m, 2H), 0.06(m, 2H), −0.02(s, 9H). HPLC
(method 1): tR = 12.03. LCMS m/z 666.1 and 668.1 (M + H)+. HPLC
(method 2): tR = 6.04. LCMS m/z 666.1 and 668.1 (M + H)+.
Ethyl 5-Iodo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyra-
zole-4-carboxylate (17) and Ethyl 3-Iodo-1-((2-(trimethylsilyl)-
ethoxy)methyl)-1H-pyrazole-4-carboxylate (18). Isoamylnitrite
(400 mL, 2.98 mol) was added to a heterogeneous mixture of ethyl
3-amino-1H-pyrazole-4-carboxylate 16 (47.69 g, 307 mmol) in
diiodomethane (800 mL) at −10 °C over a period of 30 min. The
heterogeneous mixture was placed into a preheated oil bath at 100 °C
for 2 h. The solution was cooled to ambient temperature, diluted with
saturated aqueous sodium sulfite, and extracted with ethyl acetate. The
combined organic extracts were dried over anhydrous magnesium
sulfate, filtered, and concentrated under reduced pressure. The residue
was flash chromatographed on silica using 9:1, 4:1, 7:3, 3:2, and 1:1
hexanes:ethyl acetate as the eluant to yield 53.85 g (66%) of ethyl
3-iodo-1H-pyrazole-4-carboxylate as a light-yellow solid. Sodium
hydride (6.00 g, 150 mmol) as a 60% dispersion in mineral oil was
added to a solution of ethyl 3-iodo-1H-pyrazole-4-carboxylate (28.21 g,
106 mmol) in tetrahydrofuran (400 mL). After stirring for 1 h,
(2-(chloromethoxy)ethyl)trimethylsilane (25.0 mL, 142 mmol) was
added. After stirring for 18 h, the heterogeneous mixture was diluted with
saturated aqueous sodium bicarbonate and extracted with diethyl ether. The
combined organic extracts were dried over anhydrous magnesium sulfate,
filtered, and concentrated under reduced pressure. The residue was flash
chromatographed with 49:1, 24:1, 23:2, 22:3, 21:4, and 4:1 hexanes:ethyl
acetate as the eluant to first 18.00 g (43%) of ethyl 5-iodo-1-((2-
(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-4-carboxylate 17 as a light-
yellow liquid then 22.50 g (54%) of ethyl 3-iodo-1-((2-(trimethylsilyl)-
ethoxy)methyl)-1H-pyrazole-4-carboxylate 18 as a light-yellow liquid.
Ethyl 5-Iodo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-4-
4-Cyclopropyl-7,8-difluoro-5-(4-(trifluoromethyl)-
phenylsulfonyl)-2-((2-(trimethylsilyl)ethoxy)methyl)-4,5-dihy-
dro-2H-pyrazolo[4,3-c]quinoline (22). Cesium acetate (2.71g, 14.1
mmol), cuprous iodide (797 mg, 4.19 mmol), and N-((3-(2-bromo-4,5-
difluorophenyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-
yl)(cyclopropyl)methyl)-4-(trifluoromethyl)benzenesulfonamide 21
(1.13 g, 1.69 mmol) were placed into a flask that was then evacuated
and refilled with nitrogen. Degassed dimethylsulfoxide (1.7 mL) was
added, and the heterogeneous mixture was placed into a preheated oil
bath at 120 °C. After stirring for 2 h, the solution was cooled to ambient
temperature. The solid that formed was washed with methylene chloride
and filtered. The filtrate was concentrated and the residue was flash
chromatographed on silica using 19:1, 9:1, 17:3, and 4:1 hexanes:ethyl
1
carboxylate (17). H NMR (300 MHz, CDCl3) δ 8.02 (s, 1H), 5.58
(s, 2H), 4.33 (q, J = 7.2 Hz, 2H), 3.60 (t, J = 8.1 Hz, 2H), 1.37 (t, J =
7.2 Hz, 3H), 0.91 (t, J = 8.1 Hz, 2H), −0.03 (s, 9H).
Ethyl 3-Iodo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-4-
1
carboxylate (18). H NMR (300 MHz, CDCl3) δ 7.97 (s, 1H), 5.40
1
(s, 2H), 4.31 (q, J = 7.2 Hz, 2H), 3.58 (t, J = 8.4 Hz, 2H), 1.35 (t, J =
7.2 Hz, 3H), 0.90 (t, J = 8.4 Hz, 2H), −0.03 (s, 9H).
acetate as the eluant to yield 645 mg (65%) of 22 as a yellow oil. H
NMR (400 MHz, CDCl3) δ 7.74 (dd, J = 11.2, 7.2 Hz, 1H), 7.52 (dd, J =
10.0, 8.4 Hz, 1H), 7.39 (d, J = 8.4 Hz, 2H), 7.34 (d, J = 8.4 Hz, 2H), 7.15
(s, 1H), 5.23 (s, 2H), 4.92 (d, J = 7.6 Hz, 1H), 3.50 (t, J = 8.0 Hz, 2H),
1.04 (m, 1H), 0.90 (m, 2H), 0.54 (m, 1H), 0.37 (m, 2H), 0.07 (m, 1H),
−0.01 (s, 9H). HPLC (method 2): tR = 7.22. LCMS m/z 586.2 (M + H)+.
4-Cyclopropyl-7,8-difluoro-5-(4-(trifluoromethyl)-
phenylsulfonyl)-4,5-dihydro-2H-pyrazolo[4,3-c]quinoline (23).
4-Cyclopropyl-7,8-difluoro-5-(4-(trifluoromethyl)phenylsulfonyl)-2-
((2-(trimethylsilyl)ethoxy)methyl)-4,5-dihydro-2H-pyrazolo[4,3-c]-
quinoline 22 (645 mg, 1.10 mmol) was stirred in 4N hydrogen chloride
in dioxane (10 mL) and 10% aqueous hydrogen chloride (2 mL) for
18 h. The solution was concentrated to yield 500 mg (100%) of 23 as a
light-yellow solid. Chiral separation of the enantiomers of 23 provided
29. See 29 for details concerning chiral separation and characterization.
5-(4-Chlorophenylsulfonyl)-8-fluoro-4,5-dihydro-1H-
pyrazolo[4,3-c]quinoline (24). Synthesized in a similar manner to
3-(2-Bromo-4,5-difluorophenyl)-1-((2-(trimethylsilyl)-
ethoxy)methyl)-1H-pyrazole-4-carbaldehyde (20). Tetrakis-
(triphenylphosphine)palladium(0) (1.29 g, 1.12 mmol), 3-iodo-1-
((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-4-carbaldehyde 15
(3.53 g, 10.0 mmol), 2-bromo-4,5-difluorophenylboronic acid 19
(10.17 g, 42.9 mmol), and potassium carbonate (15.91 g, 115 mmol)
was placed into a flask that was then evacuated and refilled with nitrogen
three times. Water (40 mL) and 1,2-dimethoxyethane (40 mL) were
added, and the solution placed into a preheated oil bath at 80 °C.
After stirring for 18 h, the solution was extracted with diethyl ether.
The combined organic extracts were dried over anhydrous magnesium
sulfate, filtered, and concentrated. The residue was flash chromato-
graphed on silica using 49:1, 24:1, 23:2, 22:3, 21:4, and 4:1 hexanes:ethyl
acetate as the eluant to yield 2.10 g pure (50%) and 1.00 g slightly
impure (<24%) of 20 as an orange oil. 1H NMR (300 MHz, CDCl3) δ
9.76 (s, 1H), 8.19 (s, 1H), 7.53 (dd, J = 9.3, 7.2 Hz, 1H), 7.30 (dd,
J = 10.2, 8.4 Hz, 1H), 5.50 (s, 2H), 3.65 (m, 2H), 0.94 (m, 2H), −0.03
(s, 9H). HPLC (method 1): tR = 9.67. LCMS m/z 417.0 and 419.0
(M + H)+. HPLC (method 2): tR = 3.96. LCMS m/z 417.0 and 419.0
(M + H)+.
1
Scheme 1. H NMR (300 MHz, CD3OD) δ 7.77−7.74 (dd, J = 8.9,
5.0 Hz, 1H), 7.40 (s, 1H), 7.36−7.33 (dd, J = 8.8, 3.0 Hz, 1H), 7.20−7.14
(m, 5H), 4.95 (s, 2H). HMRS: calcd for C16H11ClFN3O2S (M + H)+,
364.03173; found, 364.03148. HPLC purity (method 1): tR = 5.61, 96%.
LCMS m/z 364.0 and 366.0 (M + H)+. HPLCpurity(method 2):tR = 1.09,
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95%. LCMS m/z 364.0 and 366.0 (M + H)+. HPLC purity (method 3):
tR = 10.79, 97%. LCMS m/z 364.0 and 366.0 (M + H)+.
HMRS: calcd for C20H14F5N3O2S (M + H)+, 456.08051; found, 456.07959.
HPLC purity (method 1): tR = 8.24, 100%. LCMS m/z 456.1 (M + H)+.
HPLC purity (method 2): tR = 2.62, 100%. LCMS m/z 456.1 (M + H)+.
[α]D25.0 = −133.40° (c = 5 mg/mL in 95% ethanol).
5-(4-Chlorophenylsulfonyl)-7-fluoro-4,5-dihydro-1H-
pyrazolo[4,3-c]quinoline (25). Synthesized in a similar manner to
1
Scheme 1. H NMR (300 MHz, CD3OD) δ 7.68−7.63 (dd, J = 8.6,
(R)-7,8-Difluoro-4-isopropyl-5-(4-(trifluoromethyl)-
phenylsulfonyl)-4,5-dihydro-2H-pyrazolo[4,3-c]quinoline (31).
6.2 Hz, 1H), 7.53−7.49 (dd, J = 10.1, 2.6 Hz, 1H), 7.38 (s, 1H), 7.22−
7.14 (m, 5H), 4.96 (s, 2H). HMRS: calcd for C16H11ClFN3O2S (M +
H)+, 364.03173; found, 364.03157. HPLC purity (method 1): tR = 5.64,
100%. LCMS m/z 364.0 and 366.0 (M + H)+. HPLC purity (method 2):
tR = 1.07, 100%. LCMS m/z 364.0 and 366.0 (M + H)+. HPLC purity
(method 3): tR = 10.86, 100%. LCMS m/z 364.0 and 366.0 (M + H)+.
(R)-5-((4-Chlorophenyl)sulfonyl)-8-fluoro-4-methyl-4,5-dihydro-
2H-pyrazolo[4,3-c]quinoline (26). Separation of the enantiomers of 9
was achieved using 10% [A]:90% [B] isocrat, 1.0 mL/min, where [A] =
2-propanol, [B] = hexanes, on a Chiral Technologies OD column
(4.6 mm × 250 mm), 10 μm packing, 220 nm detection at ambient
temperature. 1H NMR (300 MHz, CDCl3) δ 7.78 (dd, J = 6.0, 5.1 Hz,
1H), 7.38 (dd, J = 6.0, 3.3 Hz, 1H), 7.21 (s, 1H), 7.10 (broad m, 5H),
5.60 (q, J = 6.6 Hz, 1H), 1.29 (d, J = 6.6 Hz, 3H). HMRS: calcd for
C17H13ClFN3O2S (M + H)+, 378.04738; found, 378.04726. HPLC
purity (method 1): tR = 6.21, 97%. LCMS m/z 378.0 and 380.0
(M + H)+. HPLC purity (method 2): tR = 1.33, 95%. LCMS m/z 378.0
and 380.0 (M + H)+.
1
Synthesized in a similar manner to Scheme 4. H NMR (400 MHz,
CDCl3) δ 7.70 (dd, J = 11.2, 7.2 Hz, 1H), 7.48 (d, J = 10.0, 8.4 Hz, 1H),
7.39 (d, J = 8.8 Hz, 2H), 7.34 (d, J = 8.8 H, 2H), 7.18 (s, 1H), 4.89 (d, J =
9.6 Hz, 1H), 2.69 (s, 1H), 1.51 (m, 1H), 1.09 (d, J = 6.8 Hz, 3H), 0.83
(d, J = 6.4 Hz, 3H). HMRS: calcd for C20H16F5N3O2S (M + H)+,
458.09562; found, 458.09518. HPLC purity (method 1): tR = 8.69,
100%. LCMS m/z 458.1 (M + H)+. HPLC purity (method 2): tR = 3.02,
100%. LCMS m/z 458.1 (M + H)+.
(R)-4-Cyclopropyl-5-(4-(trifluoromethyl)phenylsulfonyl)-4,5-
dihydro-1H-pyrazolo[4,3-c]quinoline (32). Synthesized in a similar
manner to Scheme 5. Separation of the major and minor enantiomers
was achieved using 10% [A]:90% [B] isocrat, 1.0 mL/min, where [A] =
ethanol, [B] = hexanes, on a Chiral Technologies OD column (4.6 mm ×
250 mm), 10 μm packing, 220 nm detection at ambient temperature.
1H NMR (300 MHz, CDCl3) δ 7.90 (dd, J = 9.2, 2.0 Hz, 1H), 7.64 (dd,
J = 7.0, 1.5 Hz, 1H), 7.48−7.32 (m, 4H), 7.28−7.25 (m, 2H), 7.21 (s,
1H), 4.95 (d, J = 7.8 Hz, 1H), 1.06−1.03 (m, 1H), 0.52−0.34 (m, 3H),
0.15−0.05 (m, 1H). HMRS: calcd for C20H16F3N3O2S (M + H)+,
420.09881; found, 420.09842. HPLC purity (method 1): tR = 7.12, 100%.
LCMS m/z 420.1 (M + H)+. HPLC purity (method 2): tR = 1.80, 100%.
LCMS m/z 420.1 (M + H)+.
(R)-4-Cyclopropyl-8-fluoro-5-(4-(trifluoromethyl)-
phenylsulfonyl)-4,5-dihydro-1H-pyrazolo[4,3-c]quinoline (33).
Synthesized in a similar manner to Scheme 5. Separation of the major
and minor enantiomers was achieved using 10% [A]:90% [B] isocrat,
1.0 mL/min, where [A] = ethanol, [B] = hexanes, on a Chiral Technologies
OD column (4.6 mm × 250 mm), 10 μm packing, 220 nm detection at
ambient temperature. 1H NMR (300 MHz, CD3OD) δ 7.85−7.80 (dd, J =
8.9, 5.1 Hz, 1H), 7.50−7.46 (m, 2H) 7.40−7.34 (m, 4H), 7.22−7.16 (td,
J = 8.7, 2.9 Hz, 1H), 5.07−5.04 (d, J = 7.5 Hz, 1H), 1.02−0.94 (m, 1H),
0.54−0.47 (m, 1H), 0.41−0.31 (m, 2H), 0.17−0.12 (m, 1H). HMRS: calcd
for C20H15F4N3O2S (M + H)+, 438.08939; found, 438.08958. HPLCpurity
(method 1): tR = 7.65, 100%. LCMS m/z 438.1 (M + H)+. HPLC purity
(method 2): tR = 2.16, 100%. LCMS m/z 438.1 (M + H)+.
(R)-8-Fluoro-5-(4-fluorophenylsulfonyl)-4-methyl-4,5-dihydro-
1H-pyrazolo[4,3-c]quinoline (27). Synthesized in a similar manner to
Scheme 1. Separation of the enantiomers was achieved using 15%
[A]:85% [B] isocrat, 1.0 mL/min, where [A] = ethanol, [B] = hexanes,
on a Chiral Technologies OD column (4.6 mm × 250 mm), 10 μm
1
packing, 220 nm detection at ambient temperature. H NMR (300
MHz, CDCl3) δ 7.84−7.79 (dd, J = 5.1, 3.8 Hz, 1H), 7.38−7.34 (dd, J =
5.5, 2.9 Hz, 1H), 7.23 (s, 1H), 7.22−7.16 (m, 2H), 7.15−7.08 (dt, J =
5.7, 2.9 Hz, 1H), 6.80−6.75 (m, 2H), 5.67−5.60 (q, J = 6.9 Hz, 1H),
1.33−1.30 (d, J = 6.8 Hz, 3H). HMRS: calcd for C17H13F2N3O2S
(M + H)+, 362.07693; found, 362.07682. HPLC purity (method 1): tR =
5.53, 100%. LCMS m/z 362.0 (M + H)+. HPLC purity (method 2): tR =
1.04, 100%. LCMS m/z 362.1 (M + H)+.
(R)-7,8-Difluoro-4-methyl-5-(4-(trifluoromethyl)-
phenylsulfonyl)-4,5-dihydro-1H-pyrazolo[4,3-c]quinoline (28).
Synthesized in a similar manner to Scheme 4. Separation of the
enantiomers was achieved using 10% [A]:90% [B] isocrat, 1.0 mL/min,
where [A] = ethanol, [B] = hexanes, on a Chiral Technologies OD
column (4.6 mm × 250 mm), 10 μm packing, 220 nm detection at
(R)-4-Cyclopropyl-8-fluoro-5-(6-(trifluoromethyl)pyridin-3-
ylsulfonyl)-4,5-dihydro-2H-pyrazolo[4,3-c]quinoline (34). Hy-
drogen chloride (4N) in dioxane (10 mL) was added to a solution of
(R)-4-cyclopropyl-8-fluoro-5-(6-(trifluoromethyl)pyridin-3-ylsulfonyl)-
2-((2-(trimethylsilyl)ethoxy)methyl)-4,5-dihydro-2H-pyrazolo[4,3-c]-
quinoline 40 (1.30 g, 2.29 mmol) in methanol (20 mL), which was placed
into a preheated oil bath at 50 °C. After stirring for 1.5 h, the solution was
concentrated. The residue was diluted with saturated aqueous sodium
bicarbonate and extracted with methylene chloride. The combined
organic extracts were dried over anhydrous magnesium sulfate, filtered,
and concentrated. The residue was flash chromatographed on silica using
9:1, 4:1, 7:3, and 3:2 hexanes:ethyl acetate as eluant to yield 1.00 g
(100%) of 34 as a light-yellow oil. Separation of the major and minor
enantiomers was achieved using HPLC 5% [A]:95% [B] isocrat,
55 mL/min, where [A] = 2-propanol, [B] = hexanes, on a Chiral
Technologies AD column (5 cm × 50 cm), 20 μm packing, 220 nM
detection at ambient temperature. 1H NMR (400 MHz, CDCl3) δ 9.95
(braod s, 1H), 8.53 (s, 1H), 7.85 (dd, J = 8.8 and 4.8 Hz, 1H), 7.40 (m,
3H), 7.14 (dt, J = 2.8, 8.4 Hz, 1H), 5.00 (d, J = 7.6 Hz, 1H), 1.01 (m, 1H),
0.54 (m, 1H), 0.39 (m, 2H), 0.09 (m, 1H). HMRS: calcd for
C19H14F4N4O2S (M + H)+, 439.08464; found, 439.08427. HPLC purity
1
ambient temperature. H NMR (300 MHz, CDCl3) δ 8.84 (broad s,
1H), 7.74 (dd, J = 11.1, 7.5 Hz, 1H), 7.41 (m, 3H), 7.32 (d, J = 9.0 Hz, 2H),
7.29 (s, 1H), 5.63 (q, J = 6.9 Hz, 1H), 1.32 (q, J = 6.9Hz, 3H). HMRS: calcd
for C18H12F5N3O2S (M + H)+, 430.06432; found, 430.06420. HPLC purity
(method 1): tR = 7.52, 100%. LCMS m/z 430.0 (M + H)+. HPLC purity
(method 2): tR = 2.08, 100%. LCMS m/z 430.0 (M + H)+.
(R)-4-Cyclopropyl-7,8-difluoro-5-(4-fluorophenylsulfonyl)-
4,5-dihydro-2H-pyrazolo[4,3-c]quinoline (29). Synthesized in a
similar manner to Scheme 4. Separation of the enantiomers was
achieved using 10% [A]:90% [B] isocrat, 1.0 mL/min, where [A] =
2-propanol, [B] = hexanes, on a Chiral Technologies IA column
(4.6 mm × 250 mm), 5 μm packing, 220 nm detection at ambient
temperature. 1H NMR (300 MHz, CDCl3) δ 7.75 (dd, J = 11.4, 7.5 Hz,
1H), 7.48 (dd, J = 9.9, 8. Hz, 1H), 7.20 (m, 4H), 6.79 (m, 2H), 4.96 (d,
J = 7.8 Hz, 1H), 1.02 (m, 1H), 0.51 (m, 1H), 0.36 (m, 2H), 0.06 (m,
1H). HMRS: calcd for C19H14F3N3O2S (M + H)+, 406.08316; found,
406.08314. HPLC purity (method 1): tR = 7.04, 100%. LCMS m/z 406.1
(M + H)+. HPLC purity (method 2): tR = 1.75, 100%. LCMS m/z 406.1
(M + H)+.
(R)-4-Cyclopropyl-7,8-difluoro-5-(4-(trifluoromethyl)-
phenylsulfonyl)-4,5-dihydro-1H-pyrazolo[4,3-c]quinoline (30).
Separation of the enantiomers of 23 was achieved using 10% [A]:90%
[B] isocrat, 1.0 mL/min, where [A] = 2-propanol, [B] = hexanes, on a
Chiral Technologies OD column (4.6 mm × 250 mm), 10 μm packing,
220 nm detection at ambient temperature. 1H NMR (400 MHz, CDCl3)
δ 9.90 (broad s, 1H), 7.76 (dd, J = 11.2, 7.2 Hz, 1H), 7.50 (t, J = 9.2 Hz,
1H), 7.38 (d, J = 8.4 Hz, 2H), 7.33 (d, J = 8.4 Hz, 2H), 7.18 (s, 1H), 4.96
(d, J = 7.6 Hz, 1H), 1.03 (m, 1H), 0.54 (m, 1H), 0.40 (m, 2H), 0.09 (m, 1H).
(method 1): tR = 6.98, 100%. LCMS m/z 439.1 (M + H)+. HPLC purity
25.0
(method 2): tR = 1.74, 100%. LCMS m/z 439.1 (M + H)+. [α]D
=
−91.36° (c = 5 mg/mL in 95% ethanol).
2-Bromo-5-fluorophenylboronic acid (36).33 Isopropylmagne-
sium chloride (36.5 mmol, 2.0 M in tetrahydrofuran) was slowly added
to a −42 °C tetrahydrofuran (2.0 M) solution of 1-bromo-4-fluoro-2-
iodobenzene 35 (10 g, 33.2 mmol) under nitrogen and stirred at −42 °C
for 2 h. Triisopropylborate (11.4 mL, 49.8 mmol) was added, and
the reaction was warmed to room temperature and stirred overnight.
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Then 1N aqueous sodium hydroxide (20 mL) was added and the mixture
stirred for 1h. The pH was adjusted to 3 with 3N aqueous hydrogen
chloride, and the sample was extracted (2 × 20 mL) with ethyl acetate.
The combined organics were dried over anhydrous magnesium sulfate
and filtered through Celite, and the solvent was removed to obtain
(5.75 g, 79% yield) of 36 as a white solid. 1H NMR (300 MHz, CDCl3) δ
7.68−7.64 (dd, J = 9.1, 3.2 Hz, 1H), 7.54−7.49 (dd, J = 8.8, 4.8 Hz, 1H),
7.08−7.01 (1H, m), 5.76 (2H, s).
1H-pyrazol-4-yl)(cyclopropyl)methyl)-2-methylpropane-2-sulfinamide as
a brown oil which was used without further purification. HPLC
(method 1): tR = 9.63, LCMS m/z 544.0 and 546.0 (M + H)+ for the
major diastereomer and tR = 10.03, LCMS m/z 544.0 and 546.0 (M +
H)+ for the minor diastereomer. Aqueous hydrogen chloride (4N) in
dioxane (1 mL) was added to a solution of (S)-N-((R)-(3-(2-bromo-5-
fluorophenyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)-
(cyclopropyl)methyl)-2-methylpropane-2-sulfinamide in methanol
(20 mL). After stirring for 1 h, the solution was concentrated. The
residue was diluted with 1N aqueous sodium hydroxide and extracted
with methylene chloride, and the combined organic extracts were dried
over anhydrous magnesium sulfate, filtered, and concentrated to yield
(R)-(3-(2-bromo-5-fluorophenyl)-1-((2-(trimethylsilyl)ethoxy)-
methyl)-1H-pyrazol-4-yl)(cyclopropyl)methanamine as a brown oil
which was used without further purification. HPLC (method 1): tR =
5.57, LCMS m/z 423.0 and 425.0 (M − NH2)+, 440.0 and 442.0 (M +
H)+. 6-(Trifluoromethyl)pyridine-3-sulfonyl chloride 38 (835 mg,
3.40 mmol) was added to a solution of (R)-(3-(2-bromo-5-fluorophenyl)-
1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)(cyclopropyl)-
methanamine and 4-dimethylaminopyridine (120 mg, 982 mmol) in
methylene chloride (5 mL) and triethylamine (1 mL). After stirring for
18 h, the solution was concentrated and the residue was flash
chromatographed on silica using 9:1, 4:1, 7:3, and 3:2 hexanes:ethyl
acetate as eluant to yield 525 mg (55% over four steps) of 39 as an oil.
1H NMR (400 MHz, CDCl3) δ 9.03 (s, 1H), 8.16 (d, J = 8.0 Hz, 1H), 7.74
(d, J = 8.0 Hz, 1H), 7.53 (s, 1H), 7.37 (dd, J = 8.0, 2.4 Hz, 1H), 7.24 (m,
1H), 7.07 (dt, J = 2.4, 8.0 Hz, 1H), 5.39 (d, J = 10.8 Hz, 1H), 5.35 (d, J =
10.8 Hz, 1H), 5.00 (d, J = 6.4 Hz, 1H), 3.82 (dd, J = 8.8, 6.4 Hz, 1H), 3.61
(t, J = 8.4 Hz, 2H), 1.04 (m, 1H), 0.92 (t, 8.4 Hz, 2H), 0.45 (m, 1H), 0.37
(m, 1H), 0.18 (m, 1H), 0.09 (m, 1H), −0.02 (s, 9H). HPLC (method 1):
tR = 11.01 LCMS m/z 649.1 and 651.1 (M + H)+. HPLC (method 2): tR =
5.30. LCMS m/z 649.1 and 651.1 (M + H)+.
(R)-4-Cyclopropyl-8-fluoro-5-(6-(trifluoromethyl)pyridin-3-
ylsulfonyl)-2-((2-(trimethylsilyl)ethoxy)methyl)-4,5-dihydro-
2H-pyrazolo[4,3-c]quinoline (40). Cesium acetate (1.46 g, 7.61 mmol),
cuprous iodide (446 mg, 2.34 mmol), and (R)-N-((3-(2-bromo-5-
fluorophenyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)-
(cyclopropyl)methyl)-6-(trifluoromethyl)pyridine-3-sulfonamide 39
(613 mg, 944 mmol) were placed into a flask that was evacuated and
refilled with nitrogen. Degassed dimethylsulfoxide (1.0 mL) was added,
and the heterogeneous mixture was placed into a preheated oil bath at
160 °C. After stirring for 30 min, the solution was cooled to ambient
temperature, the solid that formed was washed with methylene chloride
and filtered through Celite. The filtrate was concentrated, and the residue
was flash chromatographed on silica using 19:1, 9:1, 17:3, 4:1, and 3:1
hexanes:ethyl acetate as eluant to yield 405 mg (75%) of 40 as a yellow
oil. 1H NMR (400 MHz, CDCl3) δ 8.57 (d, J = 1.6 Hz, 1H), 7.70 (dd, J = 8.8,
6.0 Hz, 1H), 7.62 (dd, J =9.6, 2.4Hz, 1H), 7.44(dd, J = 8.0, 1.6 Hz, 1H), 7.36
(d, J = 8.4 Hz, 1H), 7.23 (s, 1H), 7.11 (dt, J = 2.4, 8.4 Hz, 1H), 5.29 (d, J =
10.8 Hz, 1H), 5.22 (d, J = 10.8 Hz, 1H), 4.99 (d, J = 7.6 Hz, 1H), 3.53 (t, J =
8.4 Hz, 2H), 1.04 (m, 1H), 0.89 (m, 2H), 0.55 (m, 1H), 0.37 (m, 2H), 0.08
(m, 1H), −0.02 (s, 9H). HPLC (method 1): tR = 12.08, LCMS m/z 569.1
(M + H)+. HPLC (method 2): tR = 6.18, LCMS m/z 569.1 (M + H)+.
3-(2-Bromo-5-fluorophenyl)-1-((2-(trimethylsilyl)ethoxy)-
methyl)-1H-pyrazole-4-carbaldehyde (37). Tetrakis-
(triphenylphosphine)palladium(0) (224 mg, 194 mmol), 3-iodo-1-
((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-4-carbaldehyde 15
(1.71 g, 10.0 mmol), 2-bromo-5-fluorophenylboronic acid 36 (1.36 g,
6.23 mmol), and potassium carbonate (2.86 g, 20.7 mmol) were placed
into a flask that was evacuated and refilled with nitrogen three times.
Water (5 mL) and 1,2-dimethoxyethane (5 mL) were added and the
solution placed into a preheated oil bath at 80 °C. After stirring for 18 h,
the solution was extracted with diethyl ether. The combined organic
extracts were dried over anhydrous magnesium sulfate, filtered, and
concentrated. The residue was flash chromatographed on silica using
49:1, 24:1, 23:2, 22:3, 21:4, and 4:1 hexanes:ethyl acetate as eluant to
yield 590 mg (73%) of 37 as a brown oil. 1H NMR (400 MHz, CDCl3) δ
9.73 (s, 1H), 8.19 (s, 1H), 7.43 (m, 2H), 7.14 (dt, J = 1.8, 8.0 Hz, 1H), 5.52
(s, 2H), 3.67 (t, J = 8.4 Hz, 2H), 0.95 (t, J = 8.4 Hz, 2H), 0.01 (s, 9H).
HPLC (method 1): tR = 9.26. LCMS m/z 399.0 and 401.0 (M + H)+.
HPLC (method 2): tR = 3.58. LCMS m/z 399.0 and 401.0 (M + H)+.
6-(Trifluoromethyl)pyridine-3-sulfonyl chloride (38).34 Thi-
onyl chloride (31 mL, 0.43 mol) was slowly added to 175 mL of water at
0 °C. During the addition, the temperature was maintained between
0 and 5 °C. After addition, the solution was warmed to 15 °C and 0.47 g
(4.8 mmol) of cuprous chloride was added. The solution was diluted
with 100 mL of water and cooled back to 0 °C. A solution of 7.21 g
(0.10 mol) of sodium nitrite in 100 mL of water was slowly added to a
solution of 15.39 g (0.10 mol) of 5-amino-2-(trifluoromethyl)pyridine
in concentrated aqueous hydrogen chloride (125 mL) at 0 °C. During
addition, the temperature was maintained between 0 and 5 °C. This
mixture was then slowly added to the above prepared solution so as to
maintain a temperature between 0 and 5 °C. A voluminous precipitate
formed. The mixture was stirred for an additional 30 min after addition,
and the solid was then collected by filtration. The solid was washed with
water and dissolved in chloroform. The solution was dried over
anhydrous magnesium sulfate and filtered, and the solvent was removed
to afford 18.03 g (77%) of 38 as a tan solid. 1H NMR (400 MHz, CDCl3)
δ 9.35 (s, 1H), 8.53 (d, J = 8.4 Hz, 1H), 7.98 (d, J = 8.4 Hz, 1H).
(R)-N-((3-(2-Bromo-5-fluorophenyl)-1-((2-(trimethylsilyl)-
ethoxy)methyl)-1H-pyrazol-4-yl)(cyclopropyl)methyl)-6-
(trifluoromethyl)pyridine-3-sulfonamide (39). Titanium(IV) iso-
propoxide (1.1 mL, 3.75 mmol) was added to a solution of 3-(2-bromo-
5-fluorophenyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazole-4-
carbaldehyde 37 (590 mg, 1.48 mmol) and (S)-(−)-tert-butanesulfina-
mide (254 mg, 2.10 mmol) in tetrahydrofuran (10 mL). After stirring for
18 h, the solution was placed into a preheated oil bath at 60 °C. After
stirring for 2 h, the solution was cooled to ambient temperature, diluted
with methylene chloride and then water, and the heterogeneous solution
was filtered through Celite. The filtrate was extracted with methylene
chloride, and the combined organic extracts were dried over anhydrous
magnesium sulfate, filtered, and concentrated to yield (S)-(E)-N-((3-(2-
bromo-5-fluorophenyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-
4-yl)methylene)-2-methylpropane-2-sulfinamide as an oil which was used
without further purification. A 1.0 M solution of cyclopropyl magnesium
bromide in tetrahydrofuran (15.0 mL, 15.0 mmol) was slowly added
dropwise to a solution of (S)-(E)-N-((3-(2-bromo-5-fluorophenyl)-1-((2-
(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)methylene)-2-methylpro-
pane-2-sulfinamide in methylene chloride (15mL) at−78 °C. After stirring
for 30 min, the dry ice/acetone bath was removed and saturated aqueous
sodium bicarbonate was added followed by water until the heterogeneous
mixture became homogeneous. The biphasic solution was extracted
with methylene chloride, the combined organic extracts were dried over
anhydrous magnesium sulfate, filtered, and concentrated to yield (S)-N-
((R)-(3-(2-bromo-5-fluorophenyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-
ASSOCIATED CONTENT
■
S
* Supporting Information
NMR and HPLC/MS data for all numbered compounds as well
as the DMPK experimental protocols are provided. This material
is available free of charge via the Internet at http://pubs.acs.org.
AUTHOR INFORMATION
■
Corresponding Author
*Phone: (415)317-2669. E-mail: E-mail:garybruin@hotmail.
com.
Notes
The authors declare no competing financial interest.
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Journal of Medicinal Chemistry
Article
3−28. (c) Lleo, A. Activity of γ-Secretase on Substrates Other than APP.
Curr. Top. Med. Chem. 2008, 8, 9−16.
ACKNOWLEDGMENTS
■
We thank Jiangli Yan for providing the 2-D NMR and John Erve
for providing the HMRS data.
(9) (a) Wong, G. T.; Manfras, D.; Poulet, F. M.; Zhang, Q.; Josien, H.;
Bara, T.; Engstrom, L.; Pinzon-Ortiz, M.; Fine, J. S.; Lee, H. J. J.; Zhang,
L.; Higgins, G. A.; Parker, E. M. Chronic Treatment with the γ-Secretase
Inhibitor LY-411,575 Inhibits β-Amyloid Peptide Production and Alters
Lymphopoiesis and Intestinal Cell Differentiation. J. Biol. Chem. 2004,
279, 12876−12882. (b) Milano, J.; McKay, J.; Dagenais, C.; Foster-
Brown, L.; Pognan, F.; Gadient, R.; Jacobs, R. T.; Zacco, A.; Greenberg,
B.; Ciaccio, P. J. Modulation of Notch Processing by γ-Secretase
Inhibitors Causes Intestinal Goblet Cell Metaplasia and Induction of
Genes Known to Specify Gut Secretory Lineage Differentiation. Toxicol.
Sci. 2004, 82, 341−358.
ABBREVIATIONS USED
■
AcOH, acetic acid; AD, Alzheimer’s disease; Aβ, β-amyloid
peptides; amu, atomic mass units; APP, amyloid precursor
protein; AUC, area under the curve; CNS, central nervous
system; c-PrMgBr, cyclopropylmagnesium bromide; DCE, 1,2-
dichloroethane; DMAP, 4-dimethylaminopyridine; DME, 1,2-
dimethoxyethane; DMF, dimethylformamide; DMF-DMA,
dimethylformamide−dimethylacetal; DMP, Dess−Martin peri-
odinane; DMPK, drug metabolism and pharmacokinetics; DMS,
dimethylsulfide; DMSO, dimethylsulfoxide; dr, diastereomeric
ratio; EtOH, ethanol; Gluc Met, glucuronidation metabolism;
HES-1, hairy and enhancer of split homologue-1; HPLC, High
pressure liquid chromatography; IND, investigational new drug;
i-PrMgCl, isopropylmagnesium chloride; log P, octanol−water
partition coefficient; MDCK cells, Madin−Darby canine kidney
cells; MDR1-MDCK cells, multidrug resistance gene 1 trans-
fected Madin−Darby canine kidney cells; MeOH, methanol;
ND, not determined; NFT, neurofibrillary tangle; Ox Met,
oxidative metabolism; Pd(PPh3)4, tetrakis(triphenylphosphine)-
palladium(0); P-gp, permeability-glycoprotein; PK/PD, phar-
macokinetic/pharmacodynamic; po, oral administration; QH,
hepatic blood flow; SAR, structure−activity relationship; SEM-
Cl, (2-(chloromethoxy)ethyl)trimethylsilane; TEA, triethyl-
amine; TFA, trifluoroacetic acid; THF, tetrahydrofuran; Ti(O-
i-Pr), titanium(IV) isopropoxide; UDP-GA, uridine 5′-diphos-
phate−glucuronic acid
(10) (a) Clarke, E. E.; Churcher, I.; Ellis, S.; Wrigley, J. D. J.; Lewis, H.
D.; Harrison, T.; Shearman, M. S.; Beher, D. Intra- or Intercomplex
Binding to the γ-Secretase Enzyme. A Model to Differentiate Inhibitor
Classes. J. Biol. Chem. 2006, 281, 31279−31289. (b) Tian, G.; Sobotka-
Briner, C. D.; Zysk, J.; Liu, X.; Birr, C.; Sylvester, M. A.; Edwards, P. D.;
Scott, C. D.; Greenberg, B. D. Linear Non-competitive Inhibition of
Solubilized Human γ-Secretase by Pepstatin A Methylester, L685458,
Sulfonamides, and Benzodiazepines. J. Biol. Chem. 2002, 277, 31499−
31505.
(11) See Supporting Information for experimental details.
(12) Truong, A. P.; Aubele, D. L.; Probst, G. D.; Neitzel, M. L.; Semko,
C. M.; Bowers, S.; Dressen, D.; Hom, R. K.; Konradi, A. W.; Sham, H. L.;
Garofalo, A. W.; Keim, P. S.; Wu, J.; Dappen, M. S.; Wong, K.; Goldbach,
E.; Quinn, K. P.; Sauer, J.-M.; Brigham, E. F.; Wallace, W.; Nguyen, L.;
Hemphill, S. S.; Bova, M. P.; Bard, F.; Yednock, T. A.; Basi, G. Design,
synthesis, and structure−activity relationship of novel orally efficacious
pyrazole/sulfonamide based dihydroquinoline γ-secretase inhibitors.
Bioorg. Med. Chem. Lett. 2009, 19, 4920−4923.
(13) (a) Ye, X. M.; Konradi, A. W.; Smith, J.; Xu, Y-z.; Dressen, D.;
Garofalo, A. W.; Marugg, J.; Sham, H. L.; Truong, A. P.; Jagodzinski, J.;
Pleiss, M.; Zhang, H.; Goldbach, E.; Sauer, J.-M.; Brigham, E.; Bova, M.;
Basi, G. S. Discovery of a novel sulfonamide−pyrazolopiperidine series
as potent and efficacious γ-secretase inhibitors. Bioorg. Med. Chem. Lett.
2010, 20, 2195−2199. (b) Ye, X. M.; Konradi, A. W.; Smith, J.; Aubele,
D. L.; Garofalo, A. W.; Marugg, J.; Neitzel, M. L.; Semko, C. M.; Sham,
H. L.; Sun, M.; Truong, A. P.; Wu, J.; Zhang, H.; Goldbach, E.; Sauer, J.-
M.; Brigham, E. F.; Bova, M.; Basi, G. S. Discovery of a novel
sulfonamide-pyrazolopiperidine series as potent and efficacious γ-
secretase inhibitors (Part II). Bioorg. Med. Chem. Lett. 2010, 20, 3502−
3506.
(14) (a) Aubele, D. L.; Truong, A. P.; Dressen, D. B.; Probst, G. D.;
Bowers, S.; Mattson, M. N.; Semko, C. M.; Sun, M.; Garofalo, A. W.;
Konradi, A. W.; Sham, H. L.; Zmolek, W.; Wong, K.; Goldbach, E.;
Quinn, K. P.; Sauer, J.-M.; Brigham, E. F.; Wallace, W.; Nguyen, L.;
Bova, M. P.; Hemphill, S. S.; Basi, G. Design, synthesis and structure−
activity relationship of novel [3.3.1] bicyclic sulfonamide-pyrazoles as
potent γ-secretase inhibitors. Bioorg. Med. Chem. Lett. 2011, 21, 5791−
5794. (b) Mattson, M. N.; Neitzel, M. L.; Quincy, D. A.; Semko, C. M.;
Garofalo, A. W.; Keim, P. S.; Konradi, A. W.; Pleiss, M. A.; Sham, H. L.;
Brigham, E. F.; Goldbach, E. G.; Zhang, H.; Sauer, J.-M.; Basi, G. S.
Discovery of sulfonamide-pyrazole γ-secretase inhibitors. Bioorg. Med.
Chem. Lett. 2010, 20, 2148−2150. (c) Basi, G. S.; Hemphill, S.; Brigham,
E. F.; Liao, A.; Aubele, D. L.; Baker, J.; Barbour, R.; Bova, M.; Chen, X.-
H.; Dappen, M. S.; Eichenbaum, T.; Goldbach, E.; Hawkinson, J.;
Lawler-Herbold, R.; Hu, K.; Hui, T.; Jagodzinski, J. J.; Keim, P. S.;
Kholodenko, D.; Latimer, L. H.; Lee, M.; Marugg, J.; Mattson, M. N.;
McCauley, S.; Miller, J. L.; Motter, R.; Mutter, L.; Neitzel, M. L.; Ni, H.;
Nguyen, L.; Quinn, K.; Ruslim, L.; Semko, C. M.; Shapiro, P.; Smith, J.;
Soriano, F.; Szoke, B.; Tanaka, K.; Tang, P.; Tucker, J. A.; Ye, X. M.; Yu,
M.; Wu, J.; Xu, Y-z.; Garofalo, A. W.; Sauer, J. M.; Konradi, A. W.; Ness,
D.; Shopp, G.; Pleiss, M. A.; Freedman, S. B.; Schenk, D. Amyloid
precursor protein selective gamma-secretase inhibitors for treatment of
Alzheimer’s disease. Alzheimer’s Res. Ther. 2010, 2, 36−56. (d) Bowers,
S.; Probst, G. D.; Truong, A. P.; Hom, R. K.; Konradi, A. W.; Sham, H.
L.; Garofalo, A. W.; Wong, K.; Goldbach, E.; Quinn, K. P.; Sauer, J.-M.;
Wallace, W.; Nguyen, L.; Hemphill, S. S.; Bova, M. P.; Basi, G. S. N-
REFERENCES
■
(1) Alzheimer’s Association. 2012 Alzheimer’s disease facts and figures.
Alzheimer’s Dementia2012, 8, 131−168.
(2) (a) Querfurth, H. W.; LaFerla, F. M. Alzheimer’s Disease. N. Engl. J.
Med. 2010, 362, 329−344. (b) Ballard, C.; Gauthier, S.; Corbett, A.;
Brayne, C.; Aarsland, D.; Jones, E. Alzheimer’s disease. Lancet 2011, 377,
1019−1031.
(3) Hardy, J.; Allsop, D. Amyloid deposition as the central event in the
aetiology of Alzheimer’s disease. Trends Pharmacol. Sci. 1991, 12, 383−
388.
(4) Lee, V. M. Y.; Goedert, M.; Trojanowski, J. Q. Neurodegenerative
tauopathies. Annu. Rev. Neurosci. 2001, 24, 1121−1159.
(5) (a) Hardy, J. A.; Higgins, G. A. Alzheimer’s Disease: The Amyloid
Cascade Hypothesis. Science 1992, 256, 184−185. (b) Selkoe, D. J. The
molecular pathology of Alzheimer’s disease. Neuron 1991, 6, 487−498.
(6) Sinha, S.; Lieberburg, I. Cellular mechanisms of β-amyloid
production and secretion. Proc. Natl. Acad. Sci. U. S. A. 1999, 96, 11049−
11053.
(7) (a) D’Onofrio, G.; Panza, F.; Frisardi, V.; Solfrizzi, V.; Imbimbo, B.
P.; Paroni, G.; Cascavilla, L.; Seripa, D.; Pilotto, A. Advances in the
identification of γ-secretase inhibitors for the treatment of Alzheimer’s
disease. Expert Opin. Drug Discovery 2012, 7, 19−37. (b) De Strooper,
B.; Vassar, R.; Golde, T. The secretases: enzymes with therapeutic
potential in Alzheimer disease. Nature Rev. Neurol. 2010, 6, 99−107.
(c) Citron, M. Alzheimer’s disease: strategies for disease modification.
Nature Rev. Drug Discovery 2010, 9, 387−398. (d) Kreft, A. F.; Martone,
R.; Porte, A. Recent Advances in the Identification of γ-Secretase
Inhibitors To Clinically Test the Aβ Oligomer Hypothesis of
Alzheimer’s Disease. J. Med. Chem. 2009, 52, 6169−6188.
(8) (a) Lleo, A.; Saura, C. A. γ-Secretase Substrates and their
Implications for Drug Development in Alzheimer’s Disease. Curr. Top.
Med. Chem. 2011, 11, 1513−1527. (b) Haapasalo, A.; Kovacs, D. M. The
Many Substrates of Presenilin/γ-Secretase. J. Alzheimer’s Dis. 2011, 25,
5273
dx.doi.org/10.1021/jm301741t | J. Med. Chem. 2013, 56, 5261−5274

Journal of Medicinal Chemistry
Article
Bridged bicyclic sulfonamides as inhibitors of γ-secretase. Bioorg. Med.
Chem. Lett. 2009, 19, 6952−6956.
(15) Gavina, B. D.; Forte, B.; Mantegani, S.; Varasi, M.; Vianello, P.
Pyrazolyl-indole derivatives active as kinase inhibitors, process for their
preparation and pharmaceutical compositions comprising them.
WO2005005414, 2005.
(16) Dess, D. B.; Martin, J. C. Readily accessible 12-I-5 oxidant for the
conversion of primary and secondary alcohols to aldehydes and ketones.
J. Org. Chem. 1983, 48, 4155−4156.
(17) Miyaura, N.; Suzuki, A. Palladium-Catalyzed Cross-Coupling
Reactions of Organoboron Compounds. Chem. Rev. 1995, 95, 2457−
2483.
(18) Yamada, K.; Kubo, T.; Tokuyama, H.; Fukuyama, T. A Mild
Copper-Mediated Intramolecular Amination of Aryl Halides. Synlett
2002, 231−234.
(19) Experimental findings regarding the metabolite identification,
PK/PD, and other pharmacokinetic characteristics will be published
separately.
(20) (a) Smart, B. E. Fluorine substituent effects (on bioactivity). J.
Fluorine Chem. 2001, 109, 3−11. (b) Hansch, C.; Leo, A.; Taft, R. W. A
Survey of Hammett Substituents Constants and Resonance and
Parameters. Chem. Rev. 1991, 91, 165−195. (c) McDaniel, D. H.;
Brown, H. C. An Extended Table of Hammett Substituent Constants
based on the Ionization of Substituted Benzoic Acids. J. Org. Chem.
1958, 23, 420−427.
(21) (a) Staas, D. D.; Savage, K. L.; Sherman, V. L.; Shimp, H. L.; Lyle,
T. A.; Tran, L. O.; Wiscount, C. M.; McMasters, D. L.; Sanderson, P. E.
J.; Williams, P. D.; Lucas, B. J.; Krueger, J. A.; Lewis, S. D.; White, R. B.;
Yu, S.; Wong, B. K.; Kochansky, C. J.; Anari, M. R.; Yan, Y.; Vacca, J. P.
Discovery of potent, selective 4-fluoroproline-based thrombin inhibitors
with improved metabolic stability. Bioorg. Med. Chem. 2006, 14, 6900−
6916. (b) Wiberg, K. B. The Concept of Strain in Organic Chemistry.
Angew. Chem., Int. Ed. Engl. 1986, 25, 312. (c) Wiberg, K. B.; Fenoglio, R.
A. Heats of Formation of C4H6 Hydrocarbons. J. Am. Chem. Soc. 1968,
90, 3395−3397. (d) Baghal-Vayjooee, M. H.; Benson, S. W. Kinetics and
Thermochemistry of the Reaction Cl + Cyclpropane ↔ HCl +
Cyclopropyl. Heat of Formation of the Cyclopropyl Radical. J. Am.
Chem. Soc. 1979, 101, 2838−2840. (e) Blanksby, S. J.; Ellison, G. B.
Bond Dissociation Energies of Organic Molecules. Acc. Chem. Res. 2003,
36, 255−263.
(LY450139), a functional γ-secretase inhibitor, for the treatment of
Alzheimer’s disease. Expert Opin. Pharmacother. 2009, 10, 1657−1664.
(28) (a) Martone, R. L.; Zhou, H.; Atchison, K.; Comery, T.; Xu, J. Z.;
Huang, X.; Gong, X.; Jin, M.; Kreft, A.; Harrison, B.; Mayer, S. C.;
Aschmies, S.; Gonzales, C.; Zaleska, M. M.; Riddell, D. R.; Wagner, E.;
Lu, P.; Sun, S.-C.; Sonnenberg-Reines, J.; Oganesian, A.; Adkins, K.;
Leach, M. W.; Clarke, D. W.; Huryn, D.; Abou-Gharbia, M.; Magolda,
R.; Bard, J.; Frick, G.; Raje, S.; Forlow, S. B.; Balliet, C.; Burczynski, M.
E.; Reinhart, P. H.; Wan, H. I.; Pangalos, M. N.; Jacobsen, J. S.
Begacestat (GSI-953): a novel, selective thiophene sulfonamide
inhibitor of amyloid precursor protein γ-secretase for the treatment of
Alzheimer’s disease. J. Pharmacol. Exp. Ther. 2009, 331, 598−608.
(b) Mayer, S. C.; Kreft, A. F.; Harrison, B.; Abou-Gharbia, M.; Antane,
M.; Aschmies, S.; Atchison, K.; Chlenov, M.; Cole, D. C.; Comery, T.;
Diamantidis, G.; Ellingboe, J.; Fan, K.; Galante, R.; Gonzales, C.; Ho, D.
M.; Hoke, M. E.; Hu, Y.; Huryn, D.; Jain, U.; Jin, M.; Kremer, K.;
Kubrak, D.; Lin, M.; Lu, P.; Magolda, R.; Martone, R.; Moore, W.;
Oganesian, A.; Pangalos, M. N.; Porte, A.; Reinhart, P.; Resnick, L.;
Riddell, D. R.; Sonnenberg-Reines, J.; Stock, J. R.; Sun, S.-C.; Wagner,
E.; Wang, T.; Woller, K.; Xu, Z.; Zaleska, M. M.; Zeldis, J.; Zhang, M.;
Zhou, H.; Jacobsen, J. S. Discovery of Begacestat, a Notch-1-Sparing γ-
Secretase Inhibitor for the Treatment of Alzheimer’s Disease. J. Med.
Chem. 2008, 51, 7348−7351.
(29) (a) Tong, G.; Wang, J.-S.; Sverdlov, O.; Huang, S.-P.; Slemmon,
R.; Croop, R.; Castaneda, L.; Gu, H.; Wong, O.; Li, H.; Berman, R. M.;
Smith, C.; Albright, C. F.; Dockens, R. C. Multicenter, Randomized,
Double-Blind, Placebo-Controlled, Single-Ascending Dose Study of the
Oral γ-Secretase Inhibitor BMS-708163 (Avagacestat): Tolerability
Profile, Pharmacokinetic Parameters, and Pharmacodynamic Markers.
Clin. Ther. 2012, 34, 654−667. (b) Gillman, K. W.; Starrett, J. E.; Parker,
M. F.; Xie, K.; Bronson, J. J.; Marcin, L. R.; McElhone, K. E.; Bergstrom,
C. P.; Mate, R. A.; Williams, R.; Meredith, J. E.; Burton, C. R.; Barten, D.
M.; Toyn, J. H.; Roberts, S. B.; Lentz, K. A.; Houston, J. G.; Zaczek, R.;
Albright, C. F.; Decicco, C. P.; Macor, J. E.; Olson, R. E. Discovery and
Evaluation of BMS-708163, a Potent, Selective and Orally Bioavailable
γ-Secretase Inhibitor. ACS Med. Chem. Lett. 2010, 1, 120−124.
(30) Ye, X. M.; Konradi, A. W.; Sun, M.; Yuan, S.; Aubele, D. L.;
Dappen, M.; Dressen, D.; Garofalo, A. W.; Jagodzinski, J. J.; Latimer, L.;
Probst, G. D.; Sham, H. L.; Wone, D.; Xu, Y-z.; Ness, D.; Brigham, E.;
Kwong, G. T.; Willits, C.; Tonn, G.; Goldbach, E.; Quinn, K. P.; Zhang,
H. H.; Sauer, J.-M.; Bova, M.; Basi, G. S. Discovery of a novel [3.2.1]
benzo fused bicyclic sulfonamide-pyrazoles as potent, selective and
efficacious γ-secretase inhibitors. Bioorg. Med. Chem. Lett. 2013, 23,
996−1000.
(22) Neitzel, M. L.; Aubele, D. L.; Hom, R.; Konradi, A. W.; Probst, G.;
Semko, C. M.; Truong, A. P.; Garofalo, A. W. Fused, tricyclic
sulfonamide inhibitors of gamma secretase. WO2007143523, 2007.
(23) Robak, M. T.; Herbage, M. A.; Ellman, J. A. Synthesis and
Applications of tert-Butanesulfinamide. Chem. Rev. 2010, 110, 3600−
3740.
(31) Garofalo, A. W.; Jagodzinski, J. J.; Konradi, A. W.; Semko, C. M.;
Smith, J. L.; Xu, Y-z.; Ye, X. M. 5-(Arylsulfonyl)-pyrazolopiperidines.
WO2007064914, 2007.
(32) (a) Konradi, A. W.; Ye, X. M.; Bowers, S.; Garofalo, A. W.; Aubele,
D. L.; Dressen, D.; Ng, R.; Probst, G.; Semko, C. M.; Sun, M.; Truong,
A. P.; Dappen, M. S. N-Sulfonamido polycyclic pyrazolyl compounds.
WO2010028213, 2010; (b) Bowers, S.; Garofalo, A. W.; Hom, R. K.;
Konradi, A. W.; Mattson, M. N.; Neitzel, M. L.; Semko, C. M.; Truong,
A. P.; Wu, J.; Xu, Y.-z. Bridged N-cyclic sulfonamido inhibitors of gamma
secretase. WO2007022502, 2007.
(33) 2-Bromo-5-fluorophenylboronic acid is now commercially
available from Combi-Blocks.
(34) 6-(Trifluoromethyl)pyridine-3-sulfonyl chloride is now commer-
cially available from Matrix Scientific.
(24) Hitchcock, S. A.; Pennington, L. D. Structure−Brain Exposure
Relationships. J. Med. Chem. 2006, 49, 7559−7583.
(25) (a) Varma, M. V. S.; Obach, R. S.; Rotter, C.; Miller, H. R.; Chang,
G.; Steyn, S. J.; El-Kattan; Troutman, M. D. Physicochemical Space for
Optimum Oral Bioavailability Contribution of Human Intestinal
Absorption and First-Pass Elimination. J. Med. Chem. 2010, 53, 1098−
1108. (b) Veber, D. F.; Johnson, S. R.; Cheng, H. Y.; Smith, B. R.; Ward,
K. W.; Kopple, K. D. Molecular Properties that Influence the Oral
Bioavailability of Drug Candidates. J. Med. Chem. 2002, 45, 2615−2623.
(c) Wils, P.; Warnery, A.; Phung-Ba, V.; Legrain, S.; Scherman, D. High
Lipophilicity Decreases Drug Transport Across Intestinal Epithelial
Cells. J. Pharmacol. Exp. Ther. 1994, 269, 654−658.
(26) (a) Di, L.; Umland, J. P.; Chang, G.; Huang, Y.; Lin, Z.; Scott, D.
O.; Troutman, M. D.; Liston, T. E. Species Independence in Brain
Tissue Binding Using Brain Homogenate. Drug Metab. Dispos. 2011, 39,
1270−1277. (b) Read, K. D.; Braggio, S. Assessing brain free fraction in
early drug discovery. Expert Opin. Drug Metab. Toxicol. 2010, 6, 337−
334.
(27) (a) Yi, P.; Hadden, C.; Kulanthaivel, P.; Calvert, N.; Annes, W.;
Brown, T.; Barbuch, R. J.; Chaudhary, A.; Ayan-Oshodi, M. A.; Ring, B.
J. Disposition and metabolism of semagacestat, a γ-secretase inhibitor, in
humans. Drug Metab. Dispos. 2010, 38, 554−565. (b) Henley, D. B.;
May, P. C.; Dean, R. A.; Siemers, E. R. Development of semagacestat
5274
dx.doi.org/10.1021/jm301741t | J. Med. Chem. 2013, 56, 5261−5274