Translocator protein ligands based on N-methyl-(quinolin-4-yl)oxypropanamides with properties suitable for PET radioligand development

Article abstract of DOI:10.1016/j.ejmech.2016.08.046

Modifications to an N-methyl-(quinolin-4-yl)oxypropanamide scaffold were explored to discover leads for developing new radioligands for PET imaging of brain TSPO (translocator protein), a biomarker of neuroinflammation. Whereas contraction of the quinolin

Full text of DOI:10.1016/j.ejmech.2016.08.046

European Journal of Medicinal Chemistry 124 (2016) 677e688
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Research paper
Translocator protein ligands based on N-methyl-(quinolin-4-yl)
oxypropanamides with properties suitable for PET radioligand
development
Chad Brouwer, Kimberly J. Jenko, Sami S. Zoghbi, Cheryl L. Morse, Robert B. Innis,
Victor W. Pike*
Molecular Imaging Branch, National Institute of Mental Health, National Institutes of Health, Building 10, Room B3 C346A, 10 Center Drive, Bethesda, MD
20892, United States
a r t i c l e i n f o
a b s t r a c t
Article history:
Modifications to an N-methyl-(quinolin-4-yl)oxypropanamide scaffold were explored to discover leads
for developing new radioligands for PET imaging of brain TSPO (translocator protein), a biomarker of
neuroinflammation. Whereas contraction of the quinolinyl portion of the scaffold or cyclization of the
tertiary amido group abolished high TSPO affinity, insertion of an extra nitrogen atom into the 2-
arylquinolinyl portion was effective in retaining sub-nanomolar affinity for rat TSPO, while also
decreasing lipophilicity to within the moderate range deemed preferable for a PET radioligand.
Replacement of a phenyl group on the amido nitrogen with an isopropyl group was similarly effective.
Among others, compound 20 (N-methyl-N-phenyl-2-[2-(pyridin-2-yl)-1,8-naphthyridin-4-yloxy]prop-
anamide) appears especially appealing for PET radioligand development, based on high selectivity and
high affinity (Ki ¼ 0.5 nM) for rat TSPO, moderate lipophilicity (logD ¼ 2.48), and demonstrated
amenability to labeling with carbon-11.
Received 8 June 2016
Received in revised form
28 July 2016
Accepted 21 August 2016
Available online 25 August 2016
Keywords:
Translocator protein
PET
Carbon-11
Ligand
Binding affinity
Lipophilic efficiency
Published by Elsevier Masson SAS.
1. Introduction
important biomarker for neuroinflammation. Moreover, ligands for
TSPO have also been explored as possible drugs, particularly for
Translocator protein 18 kDa (TSPO), formerly known as the pe-
ripheral benzodiazepine receptor [1], is located predominantly at
the outer mitochondrial membrane in association with a voltage-
dependent anion channel and an adenine nucleotide transporter
[2]. TSPO is present in several major organs, and is particularly
dense in adrenal gland, heart, kidney, and testis [2]. Low amounts
are present in normal human brain, primarily in microglia [3].
Activated microglia upregulate TSPO in instances of neuronal
damage [4] as seen in many neurological disorders [5e7] including
Alzheimer's disease, movement disorders, stroke, multiple scle-
rosis, and major depression [8]. Therefore, TSPO can serve as an
anxiety [9].
For more than three decades, PET imaging of human TSPO has
been carried out with [11C]PK11195 ([11C]1) [10] or its (R)-enan-
tiomer ([11C](R)-1) [11] (Chart 1) for biomedical investigations of
neuroinflammation. [11C](R)-1 has been by far the most employed
radioligand for this purpose despite limited brain uptake [12], low
specific binding [12], and an undesirable metabolic profile [13].
Efforts to tackle these shortcomings of [11C](R)-1 have resulted in
several new structural classes of TSPO radioligand with superior
imaging characteristics (Chart 1). Examples include [11C]PBR28
([11C]2) [14,15], [11C]DAA1106 ([11C]3) [16], [11C]DPA-713 ([11C]4)
[17], [18F]DPA-714 ([18F]5) [18], [18F]FBR ([18F]6) [19], [18F]PBR111
([18F]7) [20], [18F]FEPPA ([18F]8) [21], [18F]FEMPA ([18F]9) [22], and
[
11C]ER176 ([11C]10) [23]. Nonetheless, many of these new radio-
Abbreviations: DIPEA, diisopropylethylamine; DMF, N,N-dimethylformamide;
DMSO, dimethylsulfoxide; HFIP, hexafluoroisopropyl alcohol; HRMS-ESI, high res-
olution mass spectrometry electrospray ionization; LDA, lithium diisopropylamide;
MTBE, methyl tert-butyl ether; PFP, pentafluorophenyl; PyBroP, bromo-
tripyrrolidinophosphonium hexafluorophosphate; TEA, triethylamine; TSPO,
translocator protein (18 kDa); LipE, lipophilic efficiency.
ligands also suffer particular deficiencies, most prevalent of which
is sensitivity to the rs6971 polymorphism in human subjects
[20,21,24,25].
Successful PET radioligands for imaging specific proteins in
brain are required to display a wide array of properties [26]. Among
* Corresponding author.
E-mail address: pikev@mail.nih.gov (V.W. Pike).
http://dx.doi.org/10.1016/j.ejmech.2016.08.046
0223-5234/Published by Elsevier Masson SAS.

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C. Brouwer et al. / European Journal of Medicinal Chemistry 124 (2016) 677e688
Chart 1. Structures of some notable TSPO PET radioligands.
these properties are: i) high affinity and selectivity for the target
protein; ii) low molecular weight; iii) intermediate polar surface
area for blood-brain barrier penetration; iv) moderate lipophilicity
for adequate brain entry in the absence of excessive non-specific
binding; and v) amenability to labeling with a positron-emitter,
either carbon-11 (t1/2 ¼ 20.4 min) or fluorine-18 (t1/2 ¼ 110 min).
This study aimed to develop TSPO ligands as leads with a desirable
combination of properties for PET radioligand development. We
have previously explored a series of N-methyl-(quinolin-4-yl)oxy-
propanamides as prospective TSPO ligands [27], and encouragingly
many of these ligands have shown low TSPO genotype sensitivity
in vitro. Here, we further explore structure-affinity relationships in
this structural class. High-affinity TSPO ligands emerged from this
effort and a few of these are promising new leads to PET
radioligands.
replacement of the pendant aryl ring with methoxy, 2-pyrimidinyl
or N-pyrrolidinyl, iv) the effect of cyclization to eliminate amide
bond rotation, and v) contraction of the bicyclic quinolinyl nucleus.
Generally, PET radioligands are required to have high affinity with
KD in the low nM range, and moderate lipophilicity with measured
(or computed) logD in the 2e4 range [26]. Most of the changes that
we made to the lead ligand scaffold were intended to retain the
very high TSPO affinity (Ki ¼ 0.07 nM for rat TSPO) seen in the
previously reported example 11 (Chart 2; scaffold with Y ¼ 2-
pyridinyl, and R ¼ Ph), as well as to decrease ligand computed
lipophilicity (clogD) from 4.73 towards the desirable range. Usually,
the overall shape of the scaffold was modified little to retain high
affinity, although the effects of scaffold pruning were also investi-
gated. The main strategy for lowering lipophilicity was to introduce
nitrogen into one or more of the aryl rings. The lipophilicity cost for
high ligand affinity may be indexed as a lipophilicity efficiency
parameter (LipE), defined as ligand pIC50 (or pKi) minus clogD [28].
Therefore, our overall aim encompasses the discovery of ligands
with high LipE scores (>6). The amide N-methyl substituent was
retained in all new ligands as a site that should be amenable to
labeling with carbon-11 through 11C-methylation of N-desmethyl
precursors.
2. Results and discussion
In this study, we identified new leads to PET radioligands for
imaging TSPO based on modifications to the previously reported
[27] N-methyl-(quinolin-4-yl)oxypropanamide TSPO ligand scaf-
fold (Chart 2). These modifications were aimed at exploring, i)
variation of substituents on the amide nitrogen, ii) introduction of
nitrogen into the quinolin-4-yl group or pendant aryl ring, iii)
2.1. Chemistry
As
prospective
TSPO
ligands,
the
2-heteroaryl-4-
alkoxyquinolines 18e23 (Scheme 1) were synthesized in three
steps, proceeding with acylations of the requisite 2-amino-aryle-
thanones [29]. The resultant amides 12 and 13 were subjected to
Camps cyclization [30] to yield the 2-heteroarylquinolin-4-ones 14
and 15, respectively, which were then chemoselectively O-alky-
lated to the desired ligands 18e23 (Scheme 1). The required
a-
bromoamides 16 and 17 were made under SchottenꢀBaumann
conditions.
Chart 2. The N-methyl-(quinolin-4-yl)oxypropanamide scaffold used as a basis for
new TSPO ligand development.
The 2,4-dialkoxy-1,8-naphthyridine ligand 27 was made by first

C. Brouwer et al. / European Journal of Medicinal Chemistry 124 (2016) 677e688
679
Scheme 1. Syntheses of 2-heteroaryl-4-alkoxyquinolines 18e23. Reagents and conditions: (i) for 12 (X ¼ Cl): TEA, CHCl3, 4 ꢁC; for 13 (X ¼ ONa): DMF, DIPEA, PyBroP, CH2Cl2, rt; (ii)
NaOH, dioxane, 110 ꢁC; (iii) KOH, EtOAc, H2O, 4 ꢁC; (iv) for 18: NaH, DMSO, 2-bromo-N-phenylpropanamide, 80 ꢁC; for 19: K2CO3, MeCN, 2-bromo-N-phenylpropanamide, 55 ꢁC; for
20e21; K2CO3, MeCN, 2-bromo-N-methyl-N-phenylpropanamide, 55e60 ꢁC; for 22e23: K2CO3, MeCN, 16e17, 55e60 ꢁC.
Scheme 2. Synthesis of the 2,4-dialkoxy-1,8-naphthyridine analog 27. Reagents and conditions: (i) MeC(OMe)3, Ac2O, 110 ꢁC; (ii) LDA, THF, hexane, ꢀ50 ꢁC; (iii) POCl3, 110 ꢁC; (iv)
NaOMe, toluene, rt; (v) NaH, DMF, DMSO, rt.

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accomplishing a regioselective methoxylation [31,32] at C2 of 2,4-
dichloro-1,8-naphthyridine to yield 2-methoxy-4-chloro-1,8-
naphthyridine 26 (Method A, Scheme 2). The position of methox-
ylation was confirmed by an alternative but much lower yielding
regiospecific synthesis wherein methyl 2-aminopyridine-3-
carboxylate was first converted into the methyl acetimidate 24.
Dieckmann condensation [33] of 24 then gave 2-methoxy-1,8-
naphthyridin-4-one 25. Compound 25 was then converted into
26 by treatment with phosphoryl chloride (Method B, Scheme 2),
but in only 3% yield. Finally, alkoxylation at C4 of 26 gave 27
(Scheme 2).
alkylation [39] of this oxide in situ with
(Scheme 5).
a-bromoamide 16
2.2. Ligand pharmacology: TSPO binding affinity, cLogD, and LipE
The binding affinities (1/Ki) of all TSPO ligands, including the
previously reported ligand 11, were determined on rat brain ho-
mogenates (Table 1). We tested all ligands as racemates only, with
the assumption that the high-affinity enantiomer has twofold
higher affinity than that recorded for the racemate [27].
The N-(p-fluorophenyl) analog 23 of the previously described
high-affinity TSPO ligand 11 maintained very high binding affinity
and a comparably high LipE value (Table 1). Ligand 23 opens up the
possibility to prepare an 18F-labeled ligand, according to modern
18F-labeling techniques [40,41].
The 2-pyrrolequinoline ligand 29 was made by chemoselective
O-alkylation [34] of 2-amino-4-hydroxyquinoline to yield 28 fol-
lowed by treatment of its free base with dimethoxytetrahydrofuran
in acetic acid (Scheme 3) [35].
Two analogs of ligand 22, namely 33 and 34, in which rotation of
the amide bond was eliminated through cyclization, were synthe-
sized in two steps. First, an appropriate lactam was subjected to a-
chlorination with hexachloroethane to give compounds 31 and 32
in moderate yields [36,37]. Alkylation of 2-(pyridin-2-yl)-quinolin-
4(1H)-one with each of these a-chlorolactams required prolonged
heating but eventually gave the desired ligands 33 and 34,
respectively, in moderate yields (Scheme 4).
Finally, a ring-contracted analog of 11, namely 36, was made
through nucleophilic amination [38] of 1-fluoro-2-nitrobenzene to
yield 2-nitro-N-(pyridin-2-ylmethyl)aniline 35, followed by treat-
ment of 35 with two equivalents of sodium hydride to give the
cyclized intermediate benzimidazole-N-oxide, and finally
Previously, we achieved an increase in LipE by introducing a
nitrogen atom at either the 20 position of the pendant phenyl group
or the 8 position of the quinolinyl scaffold (Chart 2) [27]. We pre-
sumed that by having nitrogen atoms at both positions in the same
ligand that we might significantly improve LipE, and, indeed, this
turned out to be the case in ligand 20 (Table 1). We obtained a
similar result when the pendant phenyl group alone was modified
to have nitrogen atoms at positions 20 and 60, as in ligand 21
(Table 1). Previously, we had established the tolerance of the TSPO
binding pocket for ligands containing nitrogen atoms at positions
20, 30, and 40 of the pendant aryl group of the lead scaffold [27].
Nitrogen at the 10 position in a pyrrolyl ring was no exception,
yielding ligand 29 with high affinity (Ki ¼ 0.22 nM) and high LipE
Scheme 3. Synthesis of 2-pyrrolequinoline analog 29. Reagents and conditions: (i) NaOEt, EtOH, 70 ꢁC, then -bromoamide, DMF, 90 ꢁC; (ii) AcOH, 120 ꢁC.
a
Scheme 4. Synthesis of
a
-quinolinyloxylactam analogs 33 and 34 with eliminated amide bond rotation. Reagents and conditions: (i) t-BuLi, pentane, THF, ꢀ60 to ꢀ70 ꢁC,
then ꢀ20 ꢁC; (ii) THF, ꢀ70 ꢁC, then 0 ꢁC; iii) C2Cl6 THF, ꢀ78 to ꢀ70 ꢁC, and then rt; (iv) K2CO3, MeCN, 80 ꢁC.
Scheme 5. Synthesis of ring-contracted analog 36. Reagents and conditions: (i) K2CO3, DMF, 90 ꢁC; (ii) 16, NaH, DMF, 55 ꢁC.

C. Brouwer et al. / European Journal of Medicinal Chemistry 124 (2016) 677e688
681
Table 1
Ki, clogD and LipE values for new oxypropanamide TSPO ligands; comparison with those of the classical ligand 1 and the progenitor ligand 11.
Ligand
A
Y
R1
R2
R3
Rat Ki (nM)a
cLogD
LipE
1 (PK11195)b
0.5 0.3
0.10 0.05
0.5 0.1
0.53 0.08
0.76 0.09
0.09 0.01
7.8 0.6
0.22 0.03
>1000
3.97
4.3 0.2
5.9 0.2
6.5 0.1
6.6 0.1
6.58 0.05
5.80 0.06
6.09 0.04
6.13 0.07
<3.93
11b
20
21
22
23
27
29
33
34
36
CH
N
CH
CH
CH
N
CH
CH
CH
2-Pyridinyl
2-Pyridinyl
2-Pyrimidinyl
2-Pyridinyl
2-Pyridinyl
OMe
1-Pyrrolyl
2-Pyridinyl
2-Pyridinyl
2-Pyridinyl
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Ph
Ph
4.18 (3.11 0.02)c
2.80 (2.48 0.06)c
Ph
2.71
2.54
4.25
2.02
3.72
2.07
2.64
3.53
iPr
p-F-C6H4
Ph
Ph
CH2-R2
CH2-R2
Me
CMe2-R1
CH2-R1
Me
Me
iPr
>1000
>1000
<3.36
<2.47
iPr
a
Mean SD for n ¼ 6, except for 1 (n ¼ 60).
Binding data from reference 27.
Measured value, n ¼ 6.
b
c
(6.3) (Table 1). In addition, we had found that removal of the
pendant phenyl group in the scaffold does not seriously affect
binding affinity and that LipE was maintained as a result of a
decrease in lipophilicity (clogD) by about 2 units [27]. Seeking to
exploit the apparent benefit of reducing the size of this substituent
and concomitantly molecular weight, we swapped the large and
lipophilic pendant aryl group in 20 for the small and polar methoxy
group (Table 1, 27). This replacement conserved high LipE despite
some decrease in TSPO affinity and established that TSPO is quite
tolerant for such substitutions.
Tertiary amides are present in nearly all high-affinity TSPO li-
gands (e.g., see Chart 1). We sought further improvements to LipE
by manipulation of substituents on the tertiary amido group. Pre-
viously, we also reported some TSPO ligands based on the scaffold
in Chart 2 that incorporated a methylene tether to the amide in
place of the oxygen tether [27]. As a group, these methylene-
tethered ligands had lower LipE scores than the analogous
oxygen-tethered ligands. Because LipE improved by about 1 unit
when a phenylamide was exchanged for an isopropylamide in the
methylene-tethered series [27], we expected a similar improve-
ment in the oxygen-tethered series, and indeed this was found
when comparing the new ligand 22 with 11 (Table 1).
(Table 1, 36) has a fused bicyclic scaffold whose structure is rather
similar to that found in many literature TSPO ligands, as exempli-
fied by DPA-713 (4), PBR111 (7) and IGA-1 (37) [45] (Chart 3). Ligand
36 appears to occupy essentially the same chemical space as these
compounds, so we were surprised that 36 showed no binding to
TSPO (Ki > 1000 nM). However, simple comparison of the formal 2-
dimensional structure of 36 with those of other known TSPO li-
gands with similar core scaffolds shows that the amido carbonyl
group, a key pharmacophoric element, is not aligned with the
carbonyl groups of the other ligands. Hence, the inability of 36 to
form a required directional hydrogen bond with TSPO may explain
its lack of affinity.
Previously, we had found that the elimination of amide bond
rotation in a methylene-tethered TSPO ligand, 1-methyl-3-[(2-
phenylquinolin-4-yl)methyl]pyrrolidin-2-one (30), increased LipE
by about 1 unit [27]. Therefore, we were interested to exploit this
effect again by making a locked amide rotamer of ligand 22. There
are two possible ligands that may result from tethering the
a-
methyl group in 22 to one of its amide substituents, namely the
ligands 33 and 34 (Table 1). Unlike the locked rotamer from our
earlier study, both 33 and 34 showed no binding affinity for TSPO
(Table 1). These results add to previous observations of locked
amide rotamers showing much reduced affinity for TSPO [42].
Moreover, evidence indicates that TSPO has a preference to bind the
E-rotamer of PK11195 (1) [43,44]. Thus, it seems that TSPO is highly
sensitive to the spatial arrangement of the substituents on the
requisite tertiary amide in TSPO ligands.
Chart 3. Comparison of formal structures of three high-affinity TSPO ligands with 36.
Bold bonds indicate scaffold matching that in ligand 4. For ligands 4, 7, and 37, a
carbonyl group, shown in red, is part of the matching scaffold, but not for compound
36. (For interpretation of the references to color in this figure legend, the reader is
referred to the web version of this article.)
Finally, we tested a truncated scaffold analog of 11. This ligand

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2.3. Pharmacological screen
4. Experimental section
Ligand 11, at 10
mM concentration, showed <50% inhibition of
4.1. Materials and methods
specific binding to all tested receptors and transporters, except the
5-HT1A receptor (Ki ¼ 366 nM).
Literature methods were used to prepare ligand 11 [27], 2-
(pyridin-2-yl)-quinolin-4(1H)-one [30], 2-bromo-N-methyl-N-
phenylpropanamide
[47],
2-hydroxy-N-methyl-N-phenyl-
2.4. Radioligand syntheses
propanamide [48], 1,5,5-trimethylpyrrolidin-2-one [49], and 1-
isopropylpyrrolidin-2-one [50]. All other reagents and solvents
were purchased. Air-sensitive reagents were stored under N2 in a
PureLab HE glovebox (Innovative Technology; Amesbury, MA).
Melting points were determined on an SMP30 apparatus (Stuart;
Staffordshire, UK). Boiling point vacuum pressures were deter-
mined on a DDR-1200 apparatus (J-Kem Scientific Inc.; St. Louis,
MO). Reactions in dry solvent were performed with dried reagents
under an inert atmosphere. Solutions were taken to dryness by
treatment with MgSO4 (unless stated otherwise), followed by
filtration and evaporation. 1H (400 MHz), 13C NMR (100 MHz), and
19F NMR (376 MHz) spectra were recorded on an Avance 400 in-
strument (Bruker; Billerica, MA). Chemical shifts for 19F are re-
The radioligands [11C]11 and [11C]20 were produced in practi-
cally useful radiochemical yields in only 5 min at room temperature
by treating their respective N-desmethyl precursors, 19 and 18,
with [11C]methyl triflate [46] (Scheme 6). Each radioligand was
separated with reversed phase HPLC and readily formulated in
sterile ethanol with 10% saline, for possible use in PET scanning.
Each radioligand was obtained with high radiochemical purity
(>99%) and with high specific activities (244 and 126 GBq/mmol,
respectively).
ported relative to neat TFA in a coaxial insert (
d
¼ ꢀ76.6). HRMS
data were obtained at the University of Illinois Urbana-Champaign
(Mass Spectrometry Laboratory, School of Chemical Sciences) with
a Micromass Q-Tof Ultima instrument for ESI (Waters; Milford, MA).
Preparative HPLC was performed with elution at 30 mL/min on
either a Luna PFP(2) column (5
nomenex; Torrance, CA), or a Gemini C18 column (10
30 ꢂ 250 mm; Phenomenex). Compound purities were established
on either a Luna PFP(2) column (5
m; 100 Å; 4.6 ꢂ 250 mm;
Phenomenex) or a Gemini C18 column (5
m
m; 100 Å; 30 ꢂ 250 mm; Phe-
m
m; 110 Å;
m
mm; 110 Å; 4.6 ꢂ 250 mm;
Scheme 6. Radiosyntheses of [11C]11 and [11C]20. Reagents and conditions: (i) for [11C]
11; NaOH, MeCN, rt, 5 min; for [11C]20; NaH, MeCN, rt, 5 min.
Phenomenex). All compounds were >95% pure and typically >99%
pure, as monitored by absorbance at 220 nm. Radioligands were
isolated with HPLC on an XBridge C18 column (5
mm; 130 Å;
10 ꢂ 250 mm; Waters) or a Luna C18(2) column (10
mm; 100 Å;
2.5. Lipophilicity
10 ꢂ 250 mm; Phenomenex). Radiochemical purities were deter-
mined with HPLC on an XTerra RP18 column (10
mm, 125 Å, 7.8 ꢂ
The accessibility of [11C]11 and [11C]20 afforded the opportunity
to measure their lipophilicities accurately for comparison to their
computed values. Prolonged incubation of these radioligands in
phosphate buffer (pH 7.4) at room temperature left them un-
changed chemically. Although the measured logD for [11C]11 (3.11)
was appreciably lower than the computed value (4.18), the
measured logD of [11C]20 (2.48) was quite similar to the computed
value (2.80) (Table 1). The computed values are therefore likely to
be reasonable estimates (within one log unit of true values), and are
also expected to be in approximately correct rank order for the
closely related compounds. The measured values for both radio-
ligands placed them near or within the desirable logD range of 2e4.
300 mm; Waters) using a Gold HPLC apparatus (Beckman Coulter,
Inc.; Fullerton, CA) equipped with an in-line Flow-Count NaI scin-
tillation detector (Bioscan, Inc.; Washington, DC) and a UV absor-
bance detector (Beckman Coulter, Inc.) operating at 254 nm.
Radioactive decay events were counted in an automatic gamma
counter (1480 Wizard 3; PerkinElmer Life Sciences: Wallac Oy;
Turku, Finland) with an electronic window set between 360 and
1800 keV. cLogD values were computed with Pallas for Windows
software version 3.8 in default option (CompuDrug; Bal Harbor, FL).
4.2. Chemistry
4.2.1. N-(3-Acetylpyridin-2-yl)picolinamide (12)
Pyridine-2-carbonyl chloride hydrochloride (4.89 g, 27.5 mmol)
was added portion-wise to a solution of 1-(2-amino-3-pyridinyl)-
1-ethanone (5.15 g, 37.8 mmol) and TEA (9.2 mL, 65.0 mmol) in dry
CHCl3 (100 mL) at 4 ꢁC. The mixture turned from tan to deep blue-
green and the temperature rose to 15 ꢁC. After 29 h, the mixture
was diluted with CHCl3 (100 mL) and washed with hydrochloric
acid (1 M; 100 mL ꢂ 2), water (100 mL), and brine (100 mL ꢂ 2), and
then dried. The product was recrystallized (MTBEeCHCl3) to give a
3. Conclusions
Modifications to N-methyl-(quinolin-4-yl)oxypropanamides,
especially introduction of nitrogen into the 2-arylquinolin-4-yl
scaffold, and use of an isopropyl substituent instead of a phenyl
substituent on the tertiary amido nitrogen, were effective in
retaining LipE to within a desirable range for PET radioligands.
Cyclization of the amido group or contraction of the quinolinyl ring
abolished high affinity. Three compounds (20e22) showed favor-
able properties for PET radioligand development, including high
TSPO affinity and moderate computed lipophilicity. Ligands 11 and
20 were shown further to be amenable to labeling with carbon-11,
to have acceptable moderate measured lipophilicity, and to be se-
lective for binding to TSPO.
tan powder (714 mg, 11%).
A
second recrystallization
(MTBEeMeOH) gave 12 as a cream-white powder. mp 151e154 ꢁC.
HRMS-ESI (m/z): [MþH]þ calcd for C13H12N3O2, 242.0930; found,
242.0929. 1H NMR (CDCl3):
d
13.22 (bs, 1H), 8.79 (dq, J ¼ 4.8, 0.8 Hz,
1H), 8.74 (dd, J ¼ 4.8, 2.0 Hz, 1H), 8.35 (td, J ¼ 8.0, 1.2 Hz, 1H), 8.24
(dd, J ¼ 8.0, 2.0 Hz, 1H), 7.91 (dt, J ¼ 8.0, 2.0 Hz, 1H), 7.51 (ddd,
J ¼ 7.6, 4.8, 1.2 Hz, 1H), 7.18 (dd, J ¼ 7.6, 4.8 Hz, 1H), 2.71 (s, 3H). 13
C
NMR (CDCl3):
d 200.2, 162.6, 153.2, 151.1, 150.1, 148.5, 139.8, 137.5,

C. Brouwer et al. / European Journal of Medicinal Chemistry 124 (2016) 677e688
683
126.7, 123.2, 119.3, 118.5, 28.0.
biphasic solution of KOH (1.67 g, 29.7 mmol) in water (10 mL) and
EtOAc (10 mL). This mixture was stirred rapidly and cooled to 4 ꢁC
whereupon 2-bromopropanoyl chloride (1.5 mL, 15 mmol) was
added dropwise with the temperature kept below 11 ꢁC. The ice
bath was removed and stirring continued for 1.5 h. The organic
phase was separated off and the aqueous phase extracted with
EtOAc (5 mL ꢂ 2). The combined extracts were washed with brine
(10 mL) and dried. The residue was distilled (bp 48e50 ꢁC at
0.2 mmHg), but was still contaminated with 2-bromopropanoic
acid. The oil was taken up in CHCl3 (10 mL), washed with NH4OH
(5 mL ꢂ 3), water (5 mL), and brine (5 mL ꢂ 2), and then dried to
give 16 as a colorless oil (588 mg, 29%). d 1.38 g/mL. HRMS-ESI (m/
z): [MþH]þ calcd for C7H15BrNO, 208.0337; found, 208.0341. 1H
4.2.2. N-(2-Acetylphenyl)pyrimidine-2-carboxamide (13)
DMF (30 mL) was added dropwise to a slurry of sodium py-
rimidine-2-carboxylate (5.03 g, 34.4 mmol), 20-amino-
acetophenone (4.6 mL, 37 mmol), DIPEA (6.5 mL, 37 mmol) and
PyBroP (17.4 g, 37.4 mmol) in CH2Cl2 (40 mL) held in a water bath,
wherein the temperature rose from 21 to 29 ꢁC. As the colorless
slurry dissolved, the solution turned from orange to a deep forest
green over the course of 18 h. The solvent was removed. The residue
was taken up in EtOAc (370 mL), then washed successively with aq
KHSO4 (5% w/v, 370 mL ꢂ 3), brine (370 mL), aq NaHCO3 (5% w/v,
370 mL ꢂ 3), brine (370 mL), and dried (Na2SO4). The residue was
washed with ether and dioxane and recrystallized (MTBEeCHCl3;
charcoal to decolorize) to give 13 as golden-yellow plates (435 mg,
5%). mp 201e203 ꢁC. HRMS-ESI (m/z): [MþH]þ calcd for
NMR (CDCl3, cisetrans; 1.4:1.0):
d
4.84 (sept, J ¼ 6.8 Hz,1H, cis), 4.60
(q, J ¼ 6.4 Hz, 1H, trans), 4.53 (q, J ¼ 6.4 Hz, 1H, cis), 4.20 (sept,
J ¼ 6.8 Hz, 1H, trans), 2.91 (s, 3H, cis), 2.81 (s, 3H, trans), 1.84 (d,
J ¼ 5.6 Hz, 3H, trans),1.83 (d, J ¼ 6.4 Hz, 3H, trans), 1.27 (d, J ¼ 6.8 Hz,
3H, trans), 1.21 (d, J ¼ 6.8 Hz, 3H, cis), 1.12 (d, J ¼ 7.2 Hz, 3H, cis), 1.10
C
13H12N3O2, 242.0930; found, 242.0928. 1H NMR (CDCl3):
d 13.65
(bs, 1H), 9.09 (dd, J ¼ 8.4, 0.8 Hz, 1H), 9.03 (d, J ¼ 5.2 Hz, 2H), 7.98
(dd, J ¼ 8.0, 1.6 Hz, 1H), 7.66 (ddd, J ¼ 8.4, 7.6, 1.2 Hz, 1H), 7.50 (t,
(d, J ¼ 6.8 Hz, 3H, cis). 13C NMR (CDCl3):
d 168.6 (cis), 168.4 (trans),
J ¼ 4.8 Hz, 1H), 7.22 (ddd, J ¼ 8.4, 7.6, 1.2 Hz, 1H), 2.74 (s, 3H). 13
C
48.3 (trans), 44.8 (trans), 39.5 (cis), 38.5 (trans), 28.3 (cis), 26.6
(trans), 22.1 (trans), 21.7 (cis), 20.8 (trans), 19.9 (trans),19.4 (cis),18.8
(cis).
NMR (CDCl3):
d 202.4, 161.3, 158.2, 157.8, 139.9, 135.1, 131.7, 123.2,
122.9, 122.6, 121.2, 28.6.
4.2.3. 2-(Pyridin-2-yl)-1,8-naphthyridin-4(1H)-one (14)
4.2.6. 2-Bromo-N-(4-fluorophenyl)-N-methylpropanamide (17)
The method for 16 was applied to 4-fluoro-N-methylaniline
(3.5 mL, 29 mmol) and 2-bromopropanoyl chloride (4.4 mL,
44 mmol). The crude product oil was fractionally distilled. The
forerun (bp 37e39 ꢁC at 7.2 mmHg; 0.2 mL) was discarded and 17
(6.7 g, 88%) was collected as a yellow oil (bp 84e88 ꢁC at
5.7 mmHg; d 1.5 g/mL). HRMS-ESI (m/z): [MþH]þ calcd for
Compound 12 (716 mg, 2.97 mmol) and NaOH (355 mg,
8.88 mmol) in dry dioxane (40 mL) were heated to 95 ꢁC for 3 h,
whereupon the tan solution precipitated a brown solid. The
mixture was cooled to rt, and the solid filtered off, washed with
toluene (10 mL ꢂ 3), and taken up in H2O (20 mL). The blood-red
solution was neutralized with hydrochloric acid (1 M, ~7 mL) to
precipitate a tan solid, which was filtered off, washed with water
(10 mL ꢂ 2) followed by ether (10 mL ꢂ 2), and then recrystallized
(toluene) to give 14 as a pink-tan solid with a cotton candy texture
(165 mg). More 14 (60 mg) was obtained from the mother liquoþr
(225 mg total, 34%). mp 205e206 ꢁC. HRMS-ESI (m/z): [MþH]
calcd for C13H10N3O, 224.0824; found, 224.0829. 1H NMR (CDCl3):
C
10H12BrFNO, 260.0086; found, 260.0090. 1H NMR (CDCl3):
d 7.29
(m, 2H), 7.14 (m, 2H), 4.22 (q, J ¼ 6.8 Hz, 1H), 3.28 (s, 3H), 1.74 (d,
J ¼ 6.4 Hz, 3H). 13C NMR (CDCl3):
169.6, 162.1 (d, J ¼ 247 Hz), 138.8
d
(d, J ¼ 3 Hz), 129.1 (d, J ¼ 9 Hz), 116.9 (d, J ¼ 22 Hz), 38.8, 38.2, 21.7.
19F NMR (CDCl3):
d
ꢀ113.0.
d
10.78 (bs, 1H), 8.76 (td, J ¼ 2.8, 1.2 Hz, 1H), 8.74 (dd, J ¼ 4.4, 1.6 Hz,
4.2.7. N-Phenyl-2-[2-(pyridin-2-yl)-1,8-naphthyridin-4-yloxy]
propanamide (18)
1H), 8.68 (dd, J ¼ 7.6, 1.6 Hz, 1H), 8.01 (td, J ¼ 7.2, 0.8 Hz, 1H), 7.92
(dt, J ¼ 7.6, 1.6 Hz, 1H), 7.47 (ddd, J ¼ 7.2, 4.8, 0.8 Hz, 1H), 7.35 (dd,
NaH (20 mg, 0.50 mmol; 60% in mineral oil) was added to a
solution of 14 (60 mg, 0.27 mmol) in dry DMSO (2 mL). The orange
effervescent solution was stirred for 1 h, whereupon 2-bromo-N-
phenylpropanamide (192 mg, 0.84 mmol) was added. The mixture
was heated to 80 ꢁC for 8 h, cooled to rt and quenched with water
(15 mL). The precipitate was filtered off, washed with water and
purified with HPLC [PFP column; MeOHeNH4CH3CO2 buffer
(25 mM, pH 5); 70:30] to give a peach solid (14 mg, 14%), which was
recrystallized (tolueneeMTBE) to give 18. mp 222e223 ꢁC. HRMS-
ESI (m/z): [MþH]þ calcd for C22H19N4O2, 371.1508; found, 371.1503.
J ¼ 8.0, 3.6 Hz, 1H), 7.00 (br s, 1H). 13C NMR (CDCl3):
d 179.8, 153.6,
150.1, 149.3, 148.2, 145.6, 137.7, 135.5, 125.5, 120.9, 120.5, 120.0,
107.2.
4.2.4. 2-(Pyrimidin-2-yl)quinolin-4(1H)-one (15)
Compound 13 (407 mg, 1.69 mmol) and NaOH (202 mg,
5.05 mmol) in dry dioxane (15 mL) were heated in a pressure vessel
to 110 ꢁC for 3 h, during which time the pale yellow solution turned
yellow-brown and a red-orange solid precipitated. After cooling the
mixture to rt, the solid was filtered off, washed with dioxane, taken
up in water (20 mL), and brought to pH 5 with acetic acid,
whereupon the orange solution gave a salmon-pink precipitate.
The solid was filtered off, washed with water, and recrystallized
1H NMR (CDCl3):
d
9.13 (dd, J ¼ 4.0, 2.0 Hz, 1H), 8.75 (d, J ¼ 8.0 Hz,
1H), 8.74 (ddd, J ¼ 4.8, 2.8, 0.8 Hz, 1H), 8.57 (dd, J ¼ 8.4, 2.0 Hz, 1H),
8.44 (bs, 1H), 8.20 (s, 1H), 7.82 (dt, J ¼ 7.6, 1.6 Hz, 1H), 7.61 (dd,
J ¼ 8.8, 1.2 Hz, 2H), 7.46 (dd, J ¼ 8.4, 4.4 Hz, 1H), 7.38e7.31 (3H), 7.14
(tt, J ¼ 7.6, 0.8 Hz,1H), 5.37 (q, J ¼ 6.8 Hz,1H),1.85 (d, J ¼ 6.8 Hz, 3H).
(toluene) to give 15 as a light pink solid (187 mg, 50%). mp 258 ꢁ
C
(dec). HRMS-ESI (m/z): [MþH]þ calcd for C13H10N3O, 224.0824;
13C NMR (CDCl3):
d 168.4, 160.5, 160.5, 157.0, 154.8, 154.0, 149.0,
found, 224.0827. 1H NMR (DMSO-d6, ketoeenol; 1.4:1.0):
d
12.14 (bs,
137.0, 136.9, 131.2, 129.1,125.0,124.9, 122.4, 121.5, 120.4,116.1, 100.2,
75.7, 18.2.
1H, keto), 9.10 (d, J ¼ 4.8 Hz, 2H, keto þ enol), 8.12 (d, J ¼ 8.8 Hz, 2H,
keto þ enol), 7.74e7.68 (m, 2H, keto þ enol), 7.37 (dt, J ¼ 6.8, 0.8 Hz,
1H, keto þ enol), 7.15 (s, 1H, keto þ enol), 3.34 (s, 1H, enol). 13C NMR
4.2.8. N-Phenyl-2-[2-(pyridin-2-yl)quinolin-4-yloxy]propanamide
(19)
(DMSO-d6):
d 177.7 (keto), 158.3 (enol), 158.0 (keto), 144.7 (enol),
144.6, 140.3 (enol), 140.2 (keto þ enol), 132.2 (keto þ enol), 125.8
(keto þ enol), 124.7 (keto þ enol), 123.6 (keto þ enol), 122.3
(keto þ enol), 119.6 (keto þ enol), 108.0 (keto þ enol).
2-(Pyridin-2-yl)quinolin-4(1H)-one (222 mg, 1.00 mmol), 2-
bromo-N-phenylpropanamide (255 mg, 1.12 mmol), and K2CO3
(834 mg, 6.03 mmol) in MeCN (35 mL) were heated to 55 ꢁC for 8 h.
The mixture was cooled to rt and poured into water (175 mL). The
precipitate was filtered off and recrystallized (aq dioxane) to give 19
as a colorless powder (283 mg, 77%). mp 205e206 ꢁC. 1H NMR
4.2.5. 2-Bromo-N-isopropyl-N-methylpropanamide (16)
N-Isopropylmethylamine (1.0 mL, 9.8 mmol) was added to a

684
C. Brouwer et al. / European Journal of Medicinal Chemistry 124 (2016) 677e688
(CDCl3):
d
8.70 (qd, J ¼ 4.8, 0.8 Hz, 1H), 8.63 (md, J ¼ 9.2, 0.8 Hz, 1H),
J ¼ 6.4 Hz, 1H, trans), 3.02 (s, 3H, cis), 2.84 (s, 3H, trans), 1.78 (d,
J ¼ 6.4 Hz, 3H, trans), 1.77 (d, J ¼ 6.8 Hz, 3H, cis), 1.38 (d, J ¼ 6.4 Hz,
3H, trans), 1.17 (d, J ¼ 6.4 Hz, 3H, trans), 1.16 (d, J ¼ 6.8 Hz, 3H, cis),
8.29 (dd, J ¼ 8.4, 0.8 Hz, 1H), 8.22 (bs, 1H), 8.17 (d, J ¼ 8.4 Hz, 1H),
8.13 (s,1H), 7.85 (dt, J ¼ 8.0, 2.0 Hz,1H), 7.80 (ddd, J ¼ 7.2, 5.6, 0.6 Hz,
1H), 7.61 (ddd, J ¼ 8.4, 6.8, 0.6 Hz, 1H), 7.55e7.52 (2H), 7.35 (ddd,
J ¼ 7.6, 4.8, 1.2 Hz, 1H), 7.34e7.29 (2H), 7.12 (m, 1H), 5.40 (q,
1.10 (d, J ¼ 6.8 Hz, 3H, cis). 13C NMR (CDCl3):
d 169.4 (cis), 169.2
(trans), 160.8 (cis), 160.7 (trans), 157.3 (trans), 157.2 (cis), 156.2
(trans), 156.1 (cis), 149.2 (cis þ trans), 148.9 (cis þ trans), 136.8
(cis þ trans), 129.9 (cis), 129.3 (trans), 125.9 (cis þ trans), 124.1
(cis þ trans), 122.1 (cis þ trans), 121.7 (trans), 121.6 (cis), 121.3
(cis þ trans), 98.7 (cis), 98.6 (trans), 72.3 (cis), 72.1 (trans), 47.7
(trans), 44.8 (cis), 39.5 (cis), 38.5 (trans), 28.3 (cis), 26.6 (trans), 22.1
(trans), 21.7 (cis), 20.8 (trans), 19.9 (trans), 19.4 (cis), 18.8 (cis).
J ¼ 6.8 Hz, 1H), 1.87 (d, J ¼ 6.8 Hz, 3H). 13C NMR (CDCl3):
d 168.9,
159.9, 157.6, 155.7, 149.3, 149.1, 136.9, 136.9, 130.3, 129.8, 129.1,
126.5, 124.9, 124.3, 121.7, 121.2, 121.2, 120.2, 99.6, 75.2, 18.4.
4.2.9. N-Methyl-N-phenyl-2-[2-(pyridin-2-yl)-1,8-naphthyridin-4-
yloxy]propanamide (20)
2-Bromo-N-methyl-N-phenylpropanamide
(122
mg,
0.50 mmol), 14 (100 mg, 0.45 mmol), and K2CO3 (371 mg,
2.69 mmol) in dry MeCN (10 mL) were heated to 55 ꢁC for 18 h. The
mixture was cooled to rt and filtered through diatomaceous earth.
The solvent was removed, the residue taken up in CH2Cl2 (10 mL),
extracted into hydrochloric acid (2 M; 30 mL ꢂ 2), and neutralized
with satd. NaHCO3. The solution was extracted with CH2Cl2
(50 mL ꢂ 3) and the combined organic layers washed with water
(50 mL) and brine (50 mL ꢂ 2), and then dried. The product was
isolated by HPLC [PFP column; MeOHephosphate buffer (25 mM,
pH 6); 80:20] followed by recrystallization (cyclohexaneedioxane)
to give 20 as a cream solid (17 mg, 10%). mp 193e194 ꢁC. HRMS-ESI
(m/z): [MþH]þ calcd for C23H21N4O2, 385.1665; found, 385.1664. 1H
4.2.12. N-(4-Fluorophenyl)-N-methyl-2-[2-(pyridin-2-yl)quinolin-
4-yloxy]propanamide (23)
The method for 21 was applied to 2-(pyridin-2-yl)-quinolin-
4(1H)-one (222 mg, 1.00 mmol) and 17 (195 mL, 1.12 mmol). The
crude product was recrystallized (aq dioxane) to give white crys-
tals. These were dried in an Abderhalden pistol under high vacuum
(T ¼ 110 ꢁC) in the presence of P2O5 for 1 d to give 23 (293 mg, 73%).
mp 179e180 ꢁC. HRMS-ESI (m/z): [MþH]þ calcd for C24H21FN3O2,
402.1618; found, 402.1613. 1H NMR (CDCl3):
d
8.75 (d, J ¼ 4.0 Hz,
1H), 8.70 (d, J ¼ 8.0 Hz, 1H), 8.27 (dd, J ¼ 8.4, 0.8 Hz, 1H), 8.08 (d,
J ¼ 8.0 Hz, 1H), 7.88 (dt, J ¼ 7.6, 1.6 Hz, 1H), 7.82 (s, 1H), 7.70 (dt,
J ¼ 8.0, 1.2 Hz, 1H), 7.68 (br s, 2H), 7.50 (dt, J ¼ 8.0, 0.8 Hz, 1H), 7.38
(ddd, J ¼ 7.6, 4.8, 1.2 Hz, 1H), 7.15 (dt, J ¼ 8.0, 1.2 Hz, 2H), 5.04 (q,
J ¼ 6.4 Hz, 1H), 3.31 (s, 3H), 1.87 (d, J ¼ 6.4 Hz, 3H). 13C NMR (CDCl3):
NMR (CDCl3):
d
9.08 (dd, J ¼ 4.4, 2.4 Hz, 1H), 8.90 (d, J ¼ 8.0 Hz, 1H),
8.78 (d, J ¼ 4.0 Hz,1H), 8.66 (dd, J ¼ 8.0, 2.0 Hz,1H), 8.03 (s,1H), 7.89
(dt, J ¼ 8.0, 2.0 Hz, 1H), 7.68 (bs, 2H), 7.50 (t, J ¼ 7.6 Hz, 2H),
7.45e7.38 (3H), 5.08 (q, J ¼ 6.4 Hz, 1H), 3.33 (s, 3H), 1.69 (d,
d
170.3, 162.3 (d, J ¼ 248 Hz), 160.7, 156.9, 156.2, 149.2, 148.6, 138.6
(d, J ¼ 3 Hz), 137.0, 130.0, 129.8 (d, J ¼ 8 Hz), 129.1, 125.9, 124.2,
J ¼ 6.4 Hz, 3H). 13C NMR (CDCl3):
d 169.9, 161.4, 159.9, 157.0, 155.5,
153.7, 148.6, 142.5, 137.0, 132.3, 130.3, 128.8, 127.8, 124.7, 122.5121.0,
116.3, 99.7, 71.7, 50.7, 38.1, 18.4.
122.3, 121.8, 121.3, 117.2 (d, J ¼ 22 Hz), 98.6, 71.1, 38.3, 18.5. 19F NMR
(CDCl3):
d
ꢀ 112.9.
4.2.13. Methyl 2-(1-methoxyethylideneamino)nicotinate (24)
4.2.10. N-Methyl-N-phenyl-2-[2-(pyrimidin-2-yl)quinolin-4-yl]
oxypropanamide (21)
Methyl 2-aminopyridine-3-carboxylate (5.17 g, 34.0 mmol) was
heated to 110 ꢁC for 1 h in a mixture of trimethyl orthoacetate
(50 mL) and acetic anhydride (20 mL), whereupon the colorless
solution turned yellow. After 1 h, methyl acetate began to distill off
and heating was continued for 5 h. The excess reagents were
removed in vacuo leaving a red oil and white syrup. This was taken
up in Et2O (100 mL) and washed with aq Na2CO3 (2 M, 50 mL ꢂ 2),
water (50 mL), and brine (50 mL ꢂ 2), and then dried. The residue
was purified by Kugelrohr distillation (140e160 ꢁC at 1.3 mmHg) to
yield 24 as a yellow oil which smelled like sugar snap peas (2.00 g,
28%). HRMS-ESI (m/z): [MþH]þ calcd for C10H13N2O3, 209.0926;
2-Bromo-N-methyl-N-phenylpropanamide
(216
mg,
0.89 mmol), 15 (179 mg, 0.80 mmol), and K2CO3 (1.04 g, 7.53 mmol)
in dry MeCN (10 mL) were heated to 55 ꢁC for 4.5 h, whereupon the
colorless slurry turned yellow. The mixture was cooled to rt and
filtered through diatomaceous earth. The solvent was removed and
the residue taken up in CH2Cl2 (100 mL), washed with water
(50 mL ꢂ 2) followed by brine (50 mL), dried, and recrystallized
(cyclohexaneedioxane) to give 21 as a colorless solid (263 mg, 86%).
mp 211e212 ꢁC. HRMS-ESI (m/z): [MþH]þ calcd for C23H21N4O2,
385.1665; found, 385.1665. 1H NMR (CDCl3):
d
9.02 (d, J ¼ 4.8 Hz,
found, 209.0930. 1H NMR (CDCl3):
d
8.51 (dd, J ¼ 4.8, 1.6 Hz, 1H),
2H), 8.32 (d, J ¼ 8.4 Hz, 1H), 8.26 (d, J ¼ 8.4 Hz, 1H), 7.87 (s, 1H), 7.72
(dt, J ¼ 6.8, 1.2 Hz, 1H), 7.55e7.51 (3H), 7.41e7.37 (3H), 7.32 (t,
J ¼ 7.2 Hz,1H), 5.09 (q, J ¼ 6.8 Hz,1H), 3.34 (s, 3H),1.69 (d, J ¼ 6.4 Hz,
8.21 (dd, J ¼ 7.6, 2.0 Hz, 1H), 7.06 (dd, J ¼ 8.0, 4.8 Hz, 1H), 3.87 (s,
3H), 3.86 (s, 3H), 1.87 (s, 3H). 13C NMR (CDCl3):
152.3, 140.0, 118.3, 117.6, 53.9, 52.1, 17.2.
d 165.9, 163.1, 160.7,
3H). 13C NMR (CDCl3):
d 170.0, 163.7, 161.0, 157.6, 155.3, 149.4, 142.6,
130.2, 130.1, 130.0, 128.6, 127.7, 126.6, 122.1, 121.7, 120.6, 100.6, 71.5,
38.2, 18.3.
4.2.14. 2-Methoxy-1,8-naphthyridin-4(1H)-one (25)
A solution of 24 (1.9 g, 9.1 mmol) in dry THF (10 mL) was added
dropwise to a slurry of LDA (10 mmol) in THFehexane (10 mL)
at ꢀ50 ꢁC, whereupon the solution turned bright orange. After 1 h,
the temperature was raised to 0 ꢁC. The solution was stirred for
another 30 min and quenched with cold satd. NH4Cl (25 mL) fol-
lowed by aq Na2CO3 (2 M, 50 mL). The aqueous layer was separated
off, washed with Et2O (50 mL ꢂ 2), and carefully neutralized with
hydrochloric acid (1 M) to give a white precipitate. Product
remaining in the mother liquor was extracted into BuOH
(50 mL ꢂ 4), which was washed with brine (50 mL ꢂ 2), and dried.
The product was isolated with HPLC [PFP column;
MeOHeNH4HCO2 buffer (25 mM, pH 4); 35:65] to give 25 as a
cream solid (1.2 g, 76%). mp ~160 ꢁC (dec; if ramping was omitted:
~200 ꢁC dec). HRMS-ESI (m/z): [MþH]þ calcd for C9H9N2O2,
4.2.11. N-Isopropyl-N-methyl-2-[2-(pyridin-2-yl)quinolin-4-yloxy]
propanamide (22)
The method for 21 was applied to 2-(pyridin-2-yl)-quinolin-
4(1H)-one (222 mg, 1.00 mmol) and 16 (170 mL, 1.12 mmol). The
crude product was recrystallized (aq EtOH) to give 22 as white sea
urchin-shaped clusters (219 mg, 63%). mp 175e176 ꢁC. HRMS-ESI
(m/z): [MþH]þ calcd for C21H24N3O2, 350.1869; found, 350.1868.
1H NMR (CDCl3, cisetrans; 1.1:1.0):
d
8.67e8.64 (4H, cis þ trans),
8.31 (d, J ¼ 8.4 Hz, 2H, cis þ trans), 8.10 (d, J ¼ 8.4 Hz, 2H, cis þ trans),
7.95 (s, 1H, trans), 7.90 (s, 1H, cis), 7.85 (tt, J ¼ 7.6, 2.0 Hz, 2H,
cis þ trans), 7.72 (dd, J ¼ 6.8, 1.6 Hz, 1H, cis), 7.71 (dd, J ¼ 6.8, 1.6 Hz,
1H, trans), 7.53 (dd, J ¼ 6.8, 1.2 Hz, 1H, cis), 7.51 (dd, J ¼ 6.8, 1.2 Hz,
1H, trans), 7.35e7.31 (2H, cis þ trans), 5.55 (q, J ¼ 6.8 Hz, 1H, trans),
5.41 (q, J ¼ 6.4 Hz, 1H, cis), 4.91 (sept, J ¼ 6.8 Hz, 1H, cis), 4.35 (sept,
177.0664; found, 177.0665. 1H NMR (HFIP-d2):
1.2 Hz,1H), 8.64 (dd, J ¼ 4.8, 1.6 Hz, 1H), 7.57 (dd, J ¼ 8.0, 5.2 Hz, 1H),
d
8.82 (dd, J ¼ 8.0,

C. Brouwer et al. / European Journal of Medicinal Chemistry 124 (2016) 677e688
685
6.27 (s, 1H), 4.11 (s, 3H). 13C NMR (HFIP-d2):
136.9, 119.5, 116.7, 90.6, 55.3.
d
164.2, 149.9, 146.8,
(CDCl3):
d
7.83 (d, J ¼ 8.0 Hz, 1H), 7.54 (q, J ¼ 8.0 Hz,1H), 7.54 (s, 1H),
7.30e7.28 (3H), 7.19e7.14 (3H), 5.67 (s, 1H), 4.90 (q, J ¼ 6.4 Hz, 1H),
4.67 (s, 2H), 3.31 (s, 3H), 1.60 (d, J ¼ 6.4 Hz, 3H). 13C NMR (CDCl3):
4.2.15. 4-Chloro-2-methoxy-1,8-naphthyridine (26)
d 169.7, 161.0, 158.0, 148.6, 142.4, 130.2, 129.9, 128.4, 127.2, 125.3,
Method A: A slurry of 2,4-dichloro-1,8-naphthyridine (4.95 g,
24.9 mmol) in dry toluene (50 mL) was added to a slurry of NaOMe
(5.0 g, 93 mmol) in toluene (50 mL) at rt. The temperature rose to
32 ꢁC as the yellow solid dissolved to give a brown solution [Note-
NaOMe should be broken up periodically if needed]. After 17 h, the
mixture was filtered through diatomaceous earth, and washed with
toluene. The solvent was removed and the residue recrystallized
(aq EtOH) to give 26 as fine, light yellow needles (3.87 g). Con-
centration of the mother liquor yielded more product (250 mg;
4.12 g total, 85%). mp 134e135 ꢁC. HRMS-ESI (m/z): [MþH]þ calcd
122.3, 121.9, 117.7, 90.5, 71.9, 38.4, 17.8.
4.2.18. 2-[2-(1H-Pyrrol-1-yl)quinolin-4-yloxy]-N-methyl-N-
phenylpropanamide (29)
2,5-Dimethoxytetrahydrofuran (195 mL, 1.50 mmol) was added
to a pale yellow solution of 28 (321 mg, 1.00 mmol) in acetic acid
(4 mL) at 120 ꢁC (bath temp.), causing the solution to turn red. This
solution was heated for 25 min, cooled to rt, poured into CH2Cl2
(100 mL), and washed with NH4OH (50 mL ꢂ 2), water (50 mL), and
brine (50 mL ꢂ 2), and finally dried. The product was isolated with
HPLC [PFP column; MeCNeNH4CH3CO2 buffer (25 mM, pH 7);
70:30] to give 29 as a pale pink solid (90 mg, 24%). mp 130 ꢁC (dec).
HRMS-ESI (m/z): [MþH]þ calcd for C23H22N3O2, 372.1712; found,
for C9H8ClN2O, 195.0325; found, 195.0327. 1H NMR (CDCl3):
d
8.99
(dd, J ¼ 4.4, 2.0 Hz,1H), 8.48 (dd, J ¼ 8.0, 2.0 Hz,1H), 7.45 (dd, J ¼ 8.0,
4.4 Hz, 1H), 7.12 (s, 1H), 4.16 (s, 3H). 13C NMR (CDCl3):
164.5, 155.4,
d
153.6, 143.7, 133.6, 120.5, 118.2, 114.0, 54.5. Method B: Compound 25
(1.20 g, 6.82 mmol) in POCl3 (10 mL) was heated to 110 ꢁC for 19 h to
give an orange solution. The solution was cooled and carefully
quenched by slow addition to water (100 mL) and neutralized with
NH4OH. This product was extracted into CHCl3 (100 mL), washed
with water (100 mL) followed by brine (100 mL ꢂ 2), and then
dried. The product was isolated by HPLC [Gemini column;
MeOHeNH4HCO2 buffer (25 mM, pH 7); 65:35] to give 26 as a pale
yellow solid (41 mg, 3%). mp 132e133 ꢁC.
372.1713. 1H NMR (CDCl3):
d
8.07 (dd, J ¼ 8.0, 0.4 Hz, 1H), 7.88 (d,
J ¼ 8.4 Hz, 1H), 7.88 (ddd, J ¼ 8.4, 7.2, 1.6 Hz, 1H), 7.52 (d, J ¼ 4.4 Hz,
1H), 7.52 (s, 1H), 7.40 (ddd, J ¼ 8.0, 6.8, 1.2 Hz, 1H), 7.37e7.31 (3H),
7.23e7.21 (2H), 6.41 (s, 1H), 6.40 (t, J ¼ 2.0 Hz, 2H), 5.04 (q,
J ¼ 6.4 Hz, 1H), 3.32 (s, 3H), 1.67 (d, J ¼ 6.4 Hz, 3H). 13C NMR (CDCl3):
d
169.4, 161.9, 150.7, 148.0, 142.3, 130.7, 130.2, 128.7, 127.9, 127.3,
124.7, 122.4, 120.0, 118.6, 111.4, 91.8, 71.8, 38.3, 18.1.
4.2.19. 3-Chloro-1,5,5-trimethylpyrrolidin-2-one (31)
t-Butyl lithium (1.64 M; 17 mmol) in pentane (2.3 mL) was
added dropwise to a solution of 2-bromomesitylene (fractionally
distilled from CaH2) (2.8 mL, 18 mmol) in dry THF (30 mL) held
between ꢀ70 ꢁC and ꢀ60 ꢁC, whereupon a white precipitate
formed. The suspension was warmed to ꢀ20 ꢁC and then cooled
back to ꢀ70 ꢁC. A solution of 1,5,5-trimethylpyrrolidin-2-one
(fractionally distilled from BaO) (2.3 mL, 17 mmol) in dry THF
(25 mL) was added dropwise. The reaction mixture was warmed to
0 ꢁC for 30 min, and then transferred dropwise via a cannula into a
solution of hexachloroethane (4.17 g, 17.6 mmol) in dry THF (40 mL)
held between ꢀ78 ꢁC and ꢀ70 ꢁC. After 1 h, the solution was
warmed to rt and stirred for another 1 h. The solvent was removed
and the residue taken up in CH2Cl2 (200 mL), and then filtered
through diatomaceous earth several times until the solution was no
longer cloudy. The solvent was removed and the residual yellow oil
shaken with water (100 mL), causing separation of a white oil. The
aqueous layer was decanted from this oil, filtered through diato-
maceous earth, and extracted with CHCl3 (100 mL ꢂ 2). The extracts
were washed with brine (100 mL ꢂ 2) and dried to give a white
waxy solid that was contaminated with 3-bromo-1,5,5-
trimethylpyrrolidin-2-one (~13%). This byproduct could not be
removed either by recrystallization (hexanes) or by sublimation
(65 ꢁC at 0.2 mmHg). Therefore, the solid was taken up in DMF
(2 mL) and stirred with LiCl (510 mg, 12 mmol) for several hours.
The mixture was diluted with water (10 mL), extracted into CHCl3
(10 mL ꢂ 2), washed with brine (10 mL ꢂ 2), and dried to give white
crystals which were triturated with cold hexanes to give 31 (1.12 g,
41%). mp 74e75 ꢁC. HRMS-ESI (m/z): [MþH]þ calcd for C7H13ClNO,
4.2.16. 2-(2-Methoxy-1,8-naphthyridin-4-yloxy)-N-methyl-N-
phenylpropanamide (27)
2-Hydroxy-N-methyl-N-phenylpropanamide
(197
mg,
1.10 mmol) and NaH (44 mg, 1.10 mmol; 60%) were stirred in dry
DMF (1.0 mL) at rt for 4 h. The red solution was transferred via
cannula into a mixture of 26 (195 mg, 1.00 mmol) in DMSOeDMF
(1:1; 2.0 mL). The reaction stalled after 30 min so the product was
isolated with HPLC [PFP column; MeOHeNH4HCO2 buffer (25 mM,
pH 7); 75:25, followed by Gemini column; MeOHeNH4HCO2 buffer
(25 mM, pH 7); 70:30] to yield 27 as small, colorless needles
(112 mg, 33%). mp 151e152 ꢁC. HRMS-ESI (m/z): [MþH]þ calcd for
C
19H20N3O3, 338.1505; found, 338.1499. 1H NMR (CDCl3):
d 8.88 (dd,
J ¼ 4.4, 2.0 Hz, 1H), 8.39 (dd, J ¼ 8.4, 2.0 Hz, 1H), 7.43e7.34 (3H),
7.28e7.25 (3H), 5.94 (s, 1H), 4.88 (q, J ¼ 6.8 Hz, 1H), 4.11 (s, 3H), 3.33
(s, 3H),1.61 (d, J ¼ 6.8 Hz, 3H). 13C NMR (CDCl3):
d 169.3,165.9,161.5,
156.0, 152.9, 142.3, 132.0, 130.2, 128.7, 127.2, 119.0, 113.8, 92.5, 71.9,
54.1, 38.2, 17.9.
4.2.17. 2-(2-Aminoquinolin-4-yloxy)-N-methyl-N-
phenylpropanamide (28)
2-Amino-4-hydroxyquinoline hydrate was dried in an Abder-
halden pistol under high vacuum (T ¼ 110 ꢁC) in the presence of
P2O5. A slurry of this quinoline (3.70 g, 23.1 mmol) with sodium
ethoxide (24 mmol) in ethanol (9.1 mL) was heated to 70 ꢁC,
whereupon a red solution formed. The solvent was distilled off,
leaving a white solid, which was dried under vacuum until the
internal temperature returned to 70 ꢁC. Dry DMF (20 mL) was
added to the solid and the slurry heated to 90 ꢁC. 2-Bromo-N-
methyl-N-phenylpropanamide (5.87 g, 24.3 mmol) was added in
one portion. Heating was continued for 30 min, whereupon NaBr
gradually precipitated. The mixture was cooled to rt, poured into
water (200 mL), and extracted into CH2Cl2 (150 mL ꢂ 3). The
organic extracts were washed with water (40 mL ꢂ 5), brine
(250 mL ꢂ 2), and finally dried (K2CO3). Recrystallization (aq EtOH)
of the white solid gave light yellow crystals, which were dried in an
Abderhalden pistol under high vacuum (T ¼ 110 ꢁC) in the presence
of P2O5 to give 28 (4.50 g, 61%). mp 182e184 ꢁC. HRMS-ESI (m/z):
[MþH]þ calcd for C19H20N3O2, 322.1556; found, 322.1552. 1H NMR
162.0686; found,162.0691. 1H NMR (CDCl3):
d
4.45 (ddd, J ¼ 8.4, 5.6,
0.4 Hz, 1H), 2.81 (s, 3H), 2.48 (dd, J ¼ 14.0, 8.4 Hz, 1H), 2.17 (dd,
J ¼ 14.0, 5.6 Hz, 1H), 1.37 (s, 3H), 1.24 (s, 3H). 13C NMR (CDCl3):
d
169.2, 58.8, 53.9, 44.3, 26.5, 26.3, 24.9.
4.2.20. 3-Chloro-1-isopropylpyrrolidin-2-one (32)
This compound was synthesized in the same manner as 31 using
1-isopropylpyrrolidin-2-one (9.3 g, 73 mmol). The workup was
modified slightly in that the reaction mixture was quenched with
satd. NaHCO3 (100 mL) and filtered through diatomaceous earth.
The organic layer was separated off, and the aqueous layer

686
C. Brouwer et al. / European Journal of Medicinal Chemistry 124 (2016) 677e688
extracted with CHCl3 (100 mL ꢂ 3). The combined extracts were
washed with brine (300 mL ꢂ 2) and dried. The residual oil was
worked up as for 31. No brominated byproduct was present so the
product was fractionally distilled (bp 68e70 ꢁC at 0.9 mmHg) to
give 32 as a colorless oil (8.2 g, 70%). HRMS-ESI (m/z): [MþH]þ calcd
neutralized with NH4OH. The resultant precipitate was recrystal-
lized twice (aq EtOH) to yield 35 as tiny golden needles (862 mg,
8%). mp 92e93 ꢁC. HRMS-ESI (m/z): [MþH]þ calcd for C12H12N3O2,
230.0930; found, 230.0937. 1H NMR (CDCl3):
d 8.87 (s, 1H), 8.64
(ddd, J ¼ 4.8, 1.6, 0.8 Hz, 1H), 8.21 (dd, J ¼ 8.4, 1.6 Hz, 1H), 7.68 (ddd,
J ¼ 7.6, 7.6, 1.6 Hz, 1H), 7.40 (ddd, J ¼ 8.4, 6.8 Hz, 1H), 7.32 (d,
J ¼ 8.0 Hz, 1H), 7.23 (ddd, J ¼ 7.6, 4.8, 0.8 Hz, 1H), 6.83 (dd, J ¼ 8.4,
0.8 Hz,1H), 6.68 (ddd, J ¼ 8.4, 6.8,1.2 Hz,1H), 4.68 (d, J ¼ 5.2 Hz,1H).
for C7H13ClNO, 162.0686; found, 162.0690. 1H NMR (CDCl3):
d 4.38
(dd, J ¼ 7.6, 4.4 Hz, 1H), 4.36 (sept, J ¼ 6.8 Hz, 1H), 3.48 (ddd,
J ¼ ꢀ13.6, 6.8, 6.4 Hz, 1H), 3.32 (ddd, J ¼ ꢀ13.6, 7.6, 4.0 Hz, 1H), 2.53
(dddd, J ¼ ꢀ14.0, 7.6, 7.6, 6.4 Hz, 1H), 2.23 (dddd, J ¼ ꢀ14.0, 7.6, 4.0,
4.0 Hz, 1H), 1.18 (d, J ¼ 6.8 Hz, 3H), 1.16 (d, J ¼ 6.8 Hz, 3H). 13C NMR
13C NMR (CDCl3):
d 156.6, 149.6, 145.0, 136.9, 136.2, 132.4, 126.9,
122.6, 121.2, 115.8, 114.3, 48.4.
(CDCl3):
d 169.4, 55.7, 43.4, 39.4, 29.8, 19.8, 19.4.
4.2.24. N-Isopropyl-N-methyl-2-(2-(pyridin-2-yl)-1H-benzo[d]
imidazol-1-yloxy)propanamide (36)
4.2.21. 1,5,5-Trimethyl-3-[2-(pyridin-2-yl)quinolin-4-yloxy]
pyrrolidin-2-one (33)
NaH (480 mg, 2.00 mmol) in mineral oil (60% w/v) was added to
a solution of 35 (229 mg, 1.00 mmol) in DMF (20 mL) at rt, which
turned from bright yellow to deep red-brown. The solution was
2-(Pyridin-2-yl)-quinolin-4(1H)-one (222 mg, 1.00 mmol), 31
(210 mg, 1.40 mmol), and K2CO3 (866 mg, 6.28 mmol) in dry MeCN
(35 mL) were heated to 80 ꢁC for 70 h. The mixture was cooled to rt,
poured into water (300 mL) and kept at 4 ꢁC overnight. The pre-
cipitate that formed was recrystallized (hexaneseEt2O) to give 33
as large white prisms (201 mg, 58%). mp 131e133 ꢁC. HRMS-ESI (m/
z): [MþH]þ calcd for C21H22N3O2, 348.1712; found, 348.1712. 1H
heated to 55 ꢁC for 1 h. A solution of 16 (315
mL, 2.00 mmol) in DMF
(5 mL) was then added and heating continued for another 1 h. The
solution was then cooled and the solvent removed under reduced
pressure. The residue was taken up into water (100 mL) and
extracted with CHCl3 (50 mL ꢂ 3). The combined extracts were then
washed with water (50 mL) followed by brine (50 mL ꢂ 2), and then
dried. The product was isolated with HPLC [PFP column;
MeOHeNH4HCO2 buffer (25 mM, pH 7); 70:30] to give 36 as an
orange syrup (240 mg, 71%). HRMS-ESI (m/z): [MþH]þ calcd for
NMR (CDCl3):
d
8.72 (ddd, J ¼ 4.8, 1,6, 0.8 Hz, 1H), 8.66 (d, J ¼ 8.0 Hz,
1H), 8.26 (dd, J ¼ 8.4, 0.8 Hz, 1H), 8.12 (s, 1H), 8.12 (d, J ¼ 8.4 Hz, 1H),
7.90 (dt, J ¼ 8.0, 2.0 Hz, 1H), 7.71 (ddd, J ¼ 8.4, 6.8, 1.6 Hz, 1H), 7.50
(ddd, J ¼ 8.4, 6.8, 1.2 Hz, 1H), 7.36 (ddd, J ¼ 7.6, 4.8, 1.2 Hz, 1H), 5.40
(dd, J ¼ 7.6, 5.6 Hz, 1H), 2.90 (s, 3H), 2.67 (dd, J ¼ 13.6, 8.0 Hz, 1H),
2.19 (dd, J ¼ 13.6, 8.0 Hz, 1H), 1.40 (s, 3H), 1.37 (s, 3H). 13C NMR
C
19H23N4O2, 339.1821; found, 339.1825. 1H NMR (CDCl3, cisetrans;
1.3:1.0):
d
8.76e8.73 (m, 2H, cis þ trans), 8.32 (s, 1H, cis), 8.30 (s, 1H,
(CDCl3):
d
169.3, 161.1, 157.2, 156.3, 149.1, 149.0, 136.9, 130.0, 129.2,
trans), 7.90e7.85 (m, 2H, cis þ trans), 7.76e7.72 (m, 3H, 2
cis þ trans), 7.68 (m, 1H, trans) 7.41 (ddd, J ¼ 7.6, 4.0, 1.2 Hz, 1H,
trans), 7.40 (ddd, J ¼ 7.6, 4.0, 1.2 Hz, 1H, cis), 7.35e7.28 (4H, 2 cis þ 2
trans), 6.06 (q, J ¼ 6.4 Hz, 1H, trans), 5.99 (q, J ¼ 6.4 Hz, 1H, cis), 4.83
(sept, J ¼ 6.8 Hz, 1H, cis), 4.13 (sept, J ¼ 6.8 Hz, 1H, trans), 2.74 (s, 3H,
trans), 2.60 (s, 3H, cis), 1.63 (d, J ¼ 6.0 Hz, 3H, cis), 1.62 (d, J ¼ 6.0 Hz,
3H, trans), 1.10 (d, J ¼ 6.4 Hz, 3H, trans), 1.04 (d, J ¼ 6.8 Hz, 3H, cis),
0.78 (d, J ¼ 6.8 Hz, 3H, cis), 0.63 (d, J ¼ 6.4 Hz, 3H, trans). 13C NMR
125.9, 124.1, 122.2, 121.9, 121.5, 99.5, 75.0, 58.3, 41.3, 27.3, 26.6, 24.6.
4.2..22. 1-Isopropyl-3-[2-(pyridin-2-yl)quinolin-4-yloxy]
pyrrolidin-2-one (34)
2-(Pyridin-2-yl)-quinolin-4(1H)-one (222 mg, 1.00 mmol), 32
(160 mL, 1.12 mmol), and K2CO3 (834 mg, 6.03 mmol) in dry MeCN
(35 mL) were heated to 80 ꢁC for 7 d. The red mixture was cooled to
rt, filtered through diatomaceous earth, taken up into CH2Cl2
(100 mL), washed with water (50 mL), and brine (50 mL ꢂ 2), and
then dried. The product was isolated with HPLC [PFP column;
MeOHeNH4CH3CO2 buffer (25 mM, pH 7); 75:25] followed by
trituration with aq MeOH (50% v/v) to give 34 as a white solid
(167 mg, 48%). mp 108e110 ꢁC. HRMS-ESI (m/z): [MþH]þ calcd for
(CDCl3):
d 169.5 (cis), 169.4 (trans), 149.2 (cis), 148.7 (trans), 147.3
(trans), 147.2 (cis), 137.8 (cis þ trans), 136.9 (cis þ trans), 133.7
(trans), 133.5 (cis), 124.7 (trans), 124.6 (cis), 124.3 (cis þ trans), 123.3
(cis þ trans), 120.2 (trans), 120.1 (cis), 111.1 (cis), 111.0 (trans), 80.8
(cis), 80.1 (trans), 47.7 (trans), 44.5 (cis), 27.8 (cis), 26.6 (trans), 20.8
(trans), 19.9 (trans), 19.3 (cis), 18.7 (cis), 16.8 (trans), 16.6 (cis).
C
21H22N3O2, 348.1712; found, 348.1712. 1H NMR (CDCl3):
d 8.70
(ddd, J ¼ 4.8, 1,6, 0.8 Hz, 1H), 8.66 (ddd, J ¼ 8.0, 1.2, 1.2 Hz, 1H), 8.28
(dd, J ¼ 8.4, 1.2 Hz, 1H), 8.14 (s, 1H), 8.10 (d, J ¼ 8.4 Hz, 1H), 7.86 (dt,
J ¼ 8.0, 2.0 Hz, 1H), 7.71 (ddd, J ¼ 8.4, 6.8, 1.2 Hz, 1H), 7.50 (ddd,
J ¼ 8.0, 6.8, 0.8 Hz, 1H), 7.34 (ddd, J ¼ 7.2, 4.8, 1.2 Hz, 1H), 5.38 (dd,
J ¼ 7.6, 7.2 Hz, 1H), 4.50 (sept, J ¼ 6.8 Hz, 1H), 3.48 (ddd, J ¼ ꢀ13.2,
10.0, 3.6 Hz, 1H), 3.42 (ddd, J ¼ ꢀ13.6, 7.2, 2.4 Hz, 1H), 2.83 (dddd,
J ¼ ꢀ13.6, 10.0, 7.6, 3.6 Hz, 1H), 2.26 (dddd, J ¼ ꢀ13.6, 8.8, 7.2, 1.6 Hz,
1H),1.26 (d, J ¼ 6.8 Hz, 3H),1.22 (d, J ¼ 6.8 Hz, 3H). 13C NMR (CDCl3):
4.3.. Radiochemistry
4.3.1. [11C]Methyl triflate
[
11C]Methyl triflate was prepared from cyclotron-produced [11C]
carbon dioxide, via conversion into [11C]methane by reduction with
hydrogen over palladium, direct iodination of [11C]methane, and
passage of the generated [11C]methyl iodide over heated silver
triflate [46].
d
169.1, 161.2, 157.3, 156.3, 149.2, 148.9, 136.9, 130.0, 129.2, 125.9,
124.1, 122.3, 121.8, 121.5, 99.5, 75.9, 43.2, 38.5, 26.2, 19.8, 19.5.
4.3.2. [11C]11
Compound 19 (0.55 mg, 1.5
triflate in the presence of aq NaOH (0.5 M, 1 eq.) in dry MeCN
(300 L) at rt for 5 min. The radioligand was isolated with HPLC
m
mol) was treated with [11C]methyl
4.2.23. 2-Nitro-N-(pyridin-2-ylmethyl)aniline (35)
2-(Aminomethyl)pyridine (4.9 mL, 48 mmol), 1-fluoro-2-
nitrobenzene (5.5 mL, 52 mmol), and K2CO3 (13 g, 95 mmol) in
DMF (35 mL) were heated for 19 h (90 ꢁC, bath temp.). The blood
red mixture was then allowed to cool, poured into citrate buffer (pH
7; 450 mL), and extracted with CH2Cl2 (250 mL ꢂ 2). The organic
layer was then washed with water (350 mL ꢂ 5), followed by brine
(350 mL), and then dried. The residue was sonicated in Et2O
(100 mL ꢂ 2), filtered through diatomaceous earth, and taken up
into rapidly stirring hydrochloric acid (0.1 M; 500 mL). The biphasic
mixture was filtered through diatomaceous earth, and the aqueous
layer was separated off, washed with Et2O (200 mL ꢂ 2), and
m
[XBridge column; MeCNeNH4OH buffer (1 mM, pH 7.7); 65:35] at
6 mL/min (tR ¼ 11 min). The isolated product was taken up in
ethanolesaline (10:90, v/v) containing polysorbate 80 (12 mg) and
sterile filtered (Millex-MP 0.22
mm, 25 mm). The radiochemical
purity of [11C]11 was >99% as established by HPLC [XBridge col-
umn; MeCNeNH4HCO2 buffer (0.1 M, pH 6.3); 50:50] at 2 mL/min
(tR ¼ 9.2 min). Product identity was also confirmed by LC-MS of
associated carrier. The average decay-corrected radiochemical yield
of [11C]11 was 17% from cyclotron-produced [11C]carbon dioxide
and the average specific activity was 244 GBq/mmol at the end of

C. Brouwer et al. / European Journal of Medicinal Chemistry 124 (2016) 677e688
687
synthesis (n ¼ 7), corresponding to about 40 min from the end of
All authors approved the final version of the manuscript. CB syn-
thesized and characterized the compounds; KJJ tested the ligands
in vitro; SSZ measured the lipophilicities; CLM prepared the radi-
oligands; RBI supervised in vitro tests; VWP initiated and super-
vised the project.
radionuclide production.
4.3.3. [11C]20
Compound 18 (0.80 mg, 2.2
m
mol) was treated with [11C]MeOTf
L) at rt for 5 min
in the presence of excess NaH in dry MeCN (300
m
The radioligand was isolated by HPLC [Luna column;
MeCNeNH4HCO2 (0.1 mM); 45:55] at 6 mL/min (tR ¼ 12 min). The
isolated product was formulated as for [11C]11. HPLC analysis of
Acknowledgments
This study was supported by the Intramural Research Program
of the National Institutes of Health (NIH), specifically the NIMH
(projects ZIA MH0023793 and ZIAMH002852). We thank the NIH
Clinical Center PET Department (Chief: Dr. Peter Herscovitch) for
radioisotope production.
[
11C]20, as for 11C]11, showed
[ > 99% radiochemical purity
(tR ¼ 6.1 min). LC-MS of associated carrier confirmed product
identity. The average decay-corrected radiochemical yield of [11C]
20 was 21% from cyclotron-produced [11C]carbon dioxide and the
average specific activity was 126 GBq/mmol at the end of radio-
synthesis (n ¼ 4).
Appendix A. Supplementary data
4.4. LogD determinations
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ejmech.2016.08.046.
These were performed on [11C]11 and [11C]20 of high radio-
chemical purity (>99.4%) by distribution between n-octanol and
sodium phosphate buffer (pH 7.4, 0.15 M) at rt with the method-
ology that we have described previously [14]. Chemical stabilities of
these radioligands in buffer were verified with HPLC. Neither
radioligand was lost to adsorption during measurements.
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Each author declares there were not any actual or potential
conflicts of interest that could have influenced this study.
Author contributions
This paper was composed with contributions from all authors.

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