Efficient syntheses of dual radioisotope-labelled PF-00217830, a D 2-partial agonist for the treatment of schizophrenia disorder

Article abstract of DOI:10.3184/174751912X13359587895460

PF-00217830 is a D₂-partial agonist developed for the treatment of schizophrenia. The dual ¹⁴C/¹⁴C and ¹⁴C/³H-labelled PF-00217830 were required for animal and human absorption, distribution, metabolism and elimination (ADME) studies. This report describes the development of the syntheses of dual ¹⁴C/ ¹⁴C- and dual ¹⁴C/³H-labelled PF-00217830. [¹⁴C]PF-00217830-L labelled at the naphthalene ring was prepared in three steps starting with [¹⁴C]naphthalene, while [ ³H]PF-00217830-R was synthesised by Pd-catalysed tritium de-bromination of brominated PF-00217830 in a single radiosynthetic step. The dual [¹⁴C/³H]PF-00217830 was made by mixing 1:10 radioactivity ratio of [¹⁴C]PF-00217830-L and [³H]PF- 00217830-R. The chemistry for [¹⁴C]PF-00217830-R labelled at the pyridine ring was also described.

Full text of DOI:10.3184/174751912X13359587895460

JOURNAL OF CHEMICAL RESEARCH 2012
RESEARCH PAPER 351
JUNE, 351–355
Efficient syntheses of dual radioisotope-labelled PF-00217830, a D2-partial
agonist for the treatment of schizophrenia disorder
Yinsheng Zhanga, Keith T. Garnesb and Donna Brownc
aRadiochemistry Group, Chemical RandD, Groton Labs, Pfizer Inc. Groton, CT, USA
bSynthesis Services, Radiosynthesis, ABC Laboratories Inc. Columbia, MO, USA
cDonna Brown Consulting, Groton, CT, USA
PF-00217830 is a D2-partial agonist developed for the treatment of schizophrenia. The dual 14C/14C and 14C/3H-labelled
PF-00217830 were required for animal and human absorption, distribution, metabolism and elimination (ADME)
studies. This report describes the development of the syntheses of dual 14C/14C- and dual 14C/3H-labelled PF-00217830.
[14C]PF-00217830-L labelled at the naphthalene ring was prepared in three steps starting with [14C]naphthalene, while
[3H]PF-00217830-R was synthesised by Pd-catalysed tritium de-bromination of brominated PF-00217830 in a single
radiosynthetic step. The dual [14C/3H]PF-00217830 was made by mixing 1:10 radioactivity ratio of [14C]PF-00217830-L
and [3H]PF-00217830-R. The chemistry for [14C]PF-00217830-R labelled at the pyridine ring was also described.
Keywords: radiochemistry, 14C/14C and 14C/3H labelling, pyridine formation, N-alkylation, tritium de-bromination,schizophrenia
Schizophrenia is a mental disorder that is characterised by
positive symptoms such as delusions, hallucinations and disor-
ganised speech/behaviour and negative symptoms including
apathy, withdrawal, lack of pleasure and impaired attention.
PF-00217830, a D2-partial agonist, was developed at phase II
for the treatment of schizophrenia and bipolar disorder.1
R indicates RHS)was synthesised in eight steps starting with
[14C]KCN by modifying the literature procedures. Our devel-
opment of a palladium-catalysed N-arylation reaction of
unprotected piperazine with aryl halide allowed us to prepare
a key radio-labelled intermediate 27 efficiently. Therefore,
[14C]PF-00217830-L labelled at the naphthalene ring was pre-
pared in three radiosynthetic steps starting with [14C]naphthalene.
Aselective bromination at the pyridine ring was achieved and
resulted in a quick access to the bromo-substituted analogue
of PF-00217830. [3H]PF-00217830-L (where L indicates LHS)
labelled at the 5-position of pyridine ring was then synthesised
in one radiosynthetic step using a Pd-catalysed tritium de-
bromination process. The dual 14C/14C and 14C/3H-labelled PF-
00217830 were prepared by combining mono-radiolabelled
versions 18 and 23, 23 and 26 based on 1:1 and 1:10 ratio of
radioactivity, respectively.
The initial in vitro biotransformation studies using non-
radiolabelled drug suggested that PF-00217830 underwent
significant scission as well as oxidation at various sites of the
intact pentacyclic compound (Scheme 1). The complex nature
of this metabolic situation limited the utilisation of mono-
radiolabelled PF-00217830 for animal and human absorption,
distribution, mechanism and elimination (ADME) studies.
In order to investigate both scission products, 14C/14C-labelled
PF-00217830 at the naphthalene ring (left hand side=LHS or
L) and the pyridine ring (right hand side = RHS or R) and dual
14C/3H-labelled PF-00217830 were required. A mixture of two
3
radioisoforms (14C and H) can be monitored simultaneously
Results and discussion
without the confusion of change in specific activity of
mono-radiolabelled metabolites that occurs using dual 14C/14C
labelled PF-00217830.
This report describes the syntheses of mono-radiolabelled
(14C, 3H) and dual-radiolabelled (14C/14C, 14C/3H) PF-00217830
for use in pre-clinical studies. The synthesis of unlabelled
PF-00217830 has been previously reported (Scheme 2).1
[14C]PF-0021783-R (18) labelled at the pyridine ring (where
A dual 14C/14C-labelled PF-00217830 was the first target, with
the labelling located at the naphthalene and pyridine ring
systems. The synthesis of unlabelled PF-00217830 involves
a total of eight steps from commercially available starting
materials (Scheme 2).1 Since the radio-labelled starting mate-
rials 1 and 8 were not commercially available, the synthesis of
each compound was devised as described below.
Scheme 1 Metabolism of PF-00217830.
* Correspondent. E-mail: yinsheng.zhang@yahoo.com

352 JOURNAL OF CHEMICAL RESEARCH 2012
Scheme 2 Synthesis of unlabelled PF-00217830.
Synthesis of [14C]PF-00217830-R labelled at the pyridine ring
formed from 16 was found to be unstable and the yield of the
subsequent reductive amination was low. In our radiosynthe-
sis, treatment of alcohol 16 with methanesulfonyl chloride first
and then with lithium chloride in one pot produced chloride
17 in 80% yield. The final N-alkylation of the chloride 17
with 1-napthylenyl-piperazine hyrochloride in basic aqueous
solution provided the desired 18, [14C]PF-00217830-R, in 80%
yield with a radiochemical purity of 98.8% and a specific
activity of 53 mCi mmol−1 after purification.
The radiosynthetic route (Scheme 3) was developed using
[14C]KCN as radio-labelled source. The cyanide substitution of
epichlorohydrin in an aqueous solution buffered with MgSO4
gave the dinitrile alcohol 11 in a good yield (62%). Treatment
of the dinitrile alcohol 11 with ammonia in MeOH (7 N)
instead of ammonia gas1 in a pressure tube formed the desired
2,6-diamino[2,6-14C2]pyridine 12 in 72% yield. Cyclisation of
12 with D,L-malic acid in concentrated H2SO4 provided ami-
nonapthyridinone 13 in 75% yield. Diazotisation of 13 with
NaNO2 in the presence of HF in pyridine gave fluoronapthy-
ridinone 14 in 83% yield. Coupling reaction of 14 with
4-(benzyloxy)butan-1-ol using potassium t-butoxide as a base
afforded 15. The yield was improved by addition of a catalytic
amount of Bu4NBr (68% versus 42%). Palladium-catalysed
hydrogenation of 15 resulted in reduction to the dihydronap-
thyridinone with concomitant removal of the benzyl group to
form alcohol 16 in 90% yield following purification.
Synthesis of [14C]PF-00217830-L labelled at the naphthalene ring
[14C]1-Napthylenyl-piperazine is a key intermediate for the
synthesis of [14C]PF-00217830-L labelled at the naphthalene
ring. The synthesis of unlabelled 1-napthylenyl-piperazine
8 was reported previously.2,3 One possible way was to use
1-amino-naphthalene, an OEB 5 compound, to combine with
bis(chloroethyl)amine-HCl in dichlorobenzene at 140 °C to
give 1-napthylenyl-piperazine. Alternatively, 1-bromonaph-
thalene could be aminated with mono-N protected piperazine
using Buchwald-Hartwig conditions following by conc. HCl
hydrolysis to furnish 1-napthylenyl-piperazine. Both known
According to the original synthesis (Scheme 2), the unla-
belled alcohol 16 was oxidised with IBX to aldehyde 7 fol-
lowed by reductive amination with aryl piperazine 8 to furnish
the target, PF-00217830. However, the radiolabelled aldehyde
Scheme 3 Synthesis of [14C]PF-00217830 labelled at the pyridine ring.

JOURNAL OF CHEMICAL RESEARCH 2012 353
approaches could be adapted to prepare 14C labelled 8, how-
ever, both labelled starting materials, 1-aminonaphthalene and
1-bromonaphthalene were not commercially available, and the
radiosynthesis would involve in multiple radiolabelling steps
would give poor overall yields.
not offer only mono substituted bromide 25 since the naphtha-
lene ring is more electron-rich than pyridine ring. Therefore,
the bromination of 22 with NBS in CHCl3 was carried out and
indeed afforded the desired bromide 24 in 88% yield. The
N-alkylation of 1-napthylenyl-piperazine 8 with the bromide
24 under the same conditions as the synthesis of [14C]PF-
00217830 gave the desired brominated precursor 25 in
62% yield after purification. In order to ensure that tritiation
occurred only at the desired position, Pd(PPh3)4 was utilised as
a reduction catalyst instead of 10% Pd/C. The latter might
cause the undesired T/H exchange at other positions according
to previous studies. So, the bromide 25 was hydrogenated with
T2 gas in the presence of Pd(PPh3)4 and KOAc at 100 °C for
2 h to give the desired tritiated PF-00217830 (26) in high
yields. [3H]PF-00217830-R (26) was obtained with a specific
activity of 25.3 Ci mmol−1 and a >99% radiochemical purity.
Our interest has been in seeking a mild, selective method for
mono-N-arylation of piperazine with aryl halides to cut the
reaction steps and make the synthesis more environmentally
friendly, which can be applied to the synthesis of [14C]PF-
00217830 and other potential drug candidates. After extensive
study on the Pd-catalysed amination of 1-halo-naphthalene
with unprotected piperazine,4 we found that PXPd, dichloro-
bis(chlorodi-t-butylphosphine) palladium (II), was the most
efficient catalyst for C–N bond coupling reaction of halonaph-
thalene and piperazine compared with other catalysts reported
in the literature.5 Therefore, [1-14C]naphthalene, a commer-
cially available compound, was brominated with CuBr2/Al2O3
in CCl4 to produce the desired [14C]1-bromonaphthalene 20 in
65% yield. Since four positions (1,4,5,8) of the naphthalene
ring are chemically equivalent, after the bromination carbon-
14 is equally distributed in the 4 positions. Then, 20 was
aminated with piperazine in the presence of PXPd to furnish
21 in 84% yield as a free base after purification (Scheme 4).
In the synthesis of [14C]PF-00217830-R (Scheme 3), the
HCl salt of 1-napthylenyl-piperazine was used, and there was
no solubility issue of N-alkylation reaction in an aqueous
solution. However, by addition of 20% CH3CN to the aqueous
reaction mixture, we were able to get the free base 21 into
the reaction solution and react with chloride 22 to achieve
a similar yield (86%) to the original reaction (80%). The
[14C]PF-00217830-L labelled at the naphthalene ring was
prepared in only three radiosynthetic steps in a 47% overall
yield with a radiochemical purity of 99.5% and a specific
activity of 54.5 mCi mmol−1.
Preparation of [14C/3H]PF-00217830 and [14C/14C]PF-00217830
Two different methods have been utilised to prepare dual
14C/14C and 14C/3H-labelled compounds. Method 1 is to synthe-
sise two radiolabelled intermediates first and then couple them
together to make a dual labelled target. Method 2 is to prepare
single-labelled final compounds separately and then combine
them in the desired ratio. Among these, method 2 is used more
often and is more convenient for obtaining the desired ratio
of two isotopomers. In addition, the single-isotope labelled
compounds were needed for in vitro and vivo studies.
In our case, 14C and 3H-labelled PF-00217830 were synthe-
sised separately, and combined together as a 1:10 ratio of the
total activity of 14C to 3H and a 1:1 ratio of the activity of 14
C
to 14C, respectively to support the dual-radiolabelled studies
(Scheme 6). The final [14C/3H]PF-00217830 (27) was prepared
as a methanol solution with 3H-radiochemical purity of 99.3%
and 14C radiochemical purity of 99.5%. The final specific
activity was determined to be 13.9 µC mg−1 for 14C and 140 µCi
Synthesis of [3H]PF-00217830-R labelled at the pyridine ring
3
mg−1 for H after radiochemical dilution. Similarly, the final
Tritium-labelled PF-00217830 was required for receptor bind-
ing and animal ADME studies. Based on the original synthetic
route for PF-00217830, the shortest way to prepare the required
[3H]PF-00217830-R is to reduce the halide precursor of PF-
00217830 with tritium gas (Scheme 5). We could either bromi-
nate the final molecule PF-00217830 or the intermediate 22.
However, the bromination of the final API, PF-00217830, did
[14C/14C]PF-00217830 (28) was obtained as a solid with a
radiochemical purity of 98.8% and a specific activity of
53.6 mCi mmol−1.
In conclusion, we have reported the efficient three-step
radiosynthesis of [14C] PF-00217830 labelled at the naphtha-
lene ring and one-step radiosynthesis of [3H]PF-00217830
Scheme 4 Synthesis of [14C]PF-00217830 labelled at the naphthalene ring.
Scheme 5 Synthesis of [3H]PF-00217830.

354 JOURNAL OF CHEMICAL RESEARCH 2012
Scheme 6 Preparation of [14C/3H]PF-0217830 and [14C/14C]PF-00217830.
1
75%, 53 mCi mmol−1). H NMR and 13C NMR were consistent with
labelled at the pyridine ring. The key radiolabelled intermedi-
ate 27 was prepared in two steps in 55% chemical and radio-
chemical yields using the N-aryl amination reaction with
unprotected piperazine in the presence of a new palladium
catalyst, PXPd. The bromide precursor of PF-00217830 was
also synthesised in two steps in 75% overall yield. We also
described the synthesis of [14C2] PF-00217830 labelled at the
pyridine ring. The modified 2-step approach to [2, 6-14C2] 2,6-
diaminopyridine 12 was developed and will be useful for other
labelled drug candidates having a pyridine core. In addition, a
new chlorination and N-alkylation sequence was developed to
avoid the stability issue of the labelled aldehyde intermediate 7.
published spectra.1
[14C2]7-Fluoro-1H-[1,8]naphthyridin-2-one (14): To a mixture of
[14C2]7-amino-1H-[1,8]naphthyridin-2-one (260 mg, 1.61 mmol,
53 mCi mmol−1) and HF-pyridine (70%, 15 mL) was added sodium
nitrite (260 mg, 3.7 mmol) at 0 °C. The resulting solution was stirred
vigorously with ice bath cooling for 3 h. To this cold solution was
added ice water (3 mL) to form a suspension. The precipitate was
collected by filtration, and then resuspended in ethyl acetate (5 mL).
The resulting suspension was filtered to give the title product as a
yellow solid (221.8 mg, 84%, 53 mCi mmol−1). 1H NMR and 13C NMR
were consistent with published spectra.1
[14C2]7-(4-Benzyloxy-butoxy)-1H-[1,8]naphthyridin-2-ol (15): A
suspension of [14C2]7-fluoro-1H-[1,8]naphthyridin-2-one (220 mg,
1.34 mmol, 53 mCi mmol−1), tetrabutylammonium bromide (218 mg,
0.7 mmol), 4-benzyloxybutanol (0.26 mL, 1.48 mmol) and THF
(4 mL) was stirred at 25 °C for 30 min and cooled to 0 °C. To this cold
mixture was added a solution of potassium t-butoxide in THF (1M,
3 mL, 3 mmol) at 0 °C. After completion of addition, the thick slurry
became a solution and was stirred at room temperature for 4 h. HCl
solution (1N, 3.2 mL) was slowly added to the reaction mixture at
25 °C and the resulting mixture was stirred at room temperature for
30 min. THF in the reaction mixture was evaporated under vacuum
and the residue was treated with ethyl acetate (10 mL). The organic
layer was separated and the aqueous layer was extracted with ethyl
acetate. The combined organic layers were dried over MgSO4
and evaporated under vacuum to give the product as a yellow solid
Experimental
All reactions were carried out under an atmosphere of nitrogen unless
otherwise stated. Microwave reactions were carried out using CEM-
discover microwave reactor. LC-MS data were obtained on a Waters
Micromass LCT mass spectrometer with flow injection analysis and
electrospray ionisation (ESI). 1H and 13C NMR spectra were recorded
on a Varian Gemini 400 MHz instrument. Chemical purity of all com-
pounds was determined by HPLC and LC-MS. Purifications were
done by flash column chromatography on Biotage Flash 40 system.
3
Quantitation of radioactivity of 14C and H labelled compounds was
performed using a Packard 2200CA liquid scintillation analyser, with
Scintiverse BD cocktail used throughout. Commercial reagents and
solvents were purchased from Aldrich and used as-received unless
otherwise noted. [14C]naphthalene (200 mCi, 54 mCi mmol−1) and
[14C]KCN (200 mCi, 54 mCi mmol−1) were purchased from American
Radiolabelled Chemicals, Inc. Intermediates 4, 8 and 22 and final
API 1 were provided by Chemical RandD, Groton Lab, Pfizer Inc.
All known compounds were identified by comparison of their NMR
spectra with those reported in the literature.
(295.2 mg, 68%, 53 mCi mmol−1). H NMR and 13C NMR were
1
consistent with published spectra.1
[14C2]7-(4-Hydroxy-butoxy)-3,4-dihydro-1H-[1,8]naphthyridin-2-
one (16): [14C2]7-(4-benzyloxy-butoxy)-[1,8]naphthyridin-2-ol (290 mg,
0.89 mol, 53 mCi mmol−1) and MeOH (5 mL) were charged to a pres-
sure reactor with 20% palladium on carbon (522 mg) and hydroge-
nated for 48 h at 45 °C and 50 psi. Upon completion, the palladium
catalyst was filtered and the filtrate was concentrated to give the prod-
uct as an off-white solid (189 mg, 90%, 53 mCi mmol−1). 1H NMR and
13C NMR were consistent with published spectra.1
3-Hydroxy-[1,5-14C2]glutaronitrile (11): This compound was pre-
pared according to a literature procedure.3 From [14C]KCN (200 mCi,
54 mCi mmol−1), the title compound was obtained as an oil (162 mCi,
[14C2]7-(4-Chloro-butoxy)-3,4-dihydro-1H-[1,8]naphthyridin-2-
one (17): To a solution of [14C2]7-(4-hydroxy-butoxy)-3,4-dihydro-
1H-[1,8]naphthyridin-2-one (180 mg, 0.76 mmol, 53 mCi mmol−1) in
THF (2 mL) was added methanesulfonyl chloride (0.1 mL, 1.29 mmol)
at –10 °C with an acetone-ice bath. Triethylamine (0.2 mL) was added
at a rate to keep the internal temperature below 0 °C. Following com-
pletion of addition, the reaction was warmed to room temperature.
LiCl (65 mg, 1.55 mmol) was added to the reaction suspension. The
resulting mixture was refluxed for 12 h. THF was removed by vacuum
distillation and then ethyl acetate (10 mL) was added to the residue.
The organic layer was washed with water (2 mL), sat. NaHCO3 (2 mL)
and brine (2 mL), and dried over MgSO4. Concentration under vacuum
gave the crude product, which was purified by flash column chroma-
tography (10% EtOAc/hexane). The title compound was obtained as
a white solid (153.8 mg, 80%, 53 mCi mmol−1) with a radiochemical
1
105 mCi mmol−1, 81%). H NMR was consistent with that of the
authentic sample.
2,6-Diamino-[2,6-14C2]pyridine (12): A mixture of 3-hydroxy-[1,5-
14C2]glutaronitrile (162 mCi, 1.54 mmol), 3-hydroxy-glutaronitrile
(170 mg, 1.54 mmol), and copper(I) chloride (40 mg, 0.4 mmol) in
ammonia solution in MeOH (7 M, 10 mL) was heated at 150 °C in an
autoclave for 4 h. After the reaction mixture had been cooled down
and filtered, the solvent was evaporated under vacuum to give a crude
material, which was purified by flash column chromatography (20%
MeOH/CH2Cl2). The title compound was obtained as a brown solid
(117.5 mCi, 72%, 53 mCi mmol−1, 241.7 mg). 1H NMR was consistent
with that of the authentic sample.
[14C2]7-Amino-1H-[1,8]naphthyridin-2-one (13): To a cold mixture
of 2,6-diamino-[2,6-14C2]pyridine (240 mg, 2.2 mmol) and D/L-malic
acid (324.5 mg, 2.42 mmol) was added dropwise conc. H2SO4
(3.0 mL) at 0 °C. The resulting mixture was heated at 110 °C for 4 h
and then cooled to 0 °C. To the cold reaction mixture was added ice
water (5 mL) slowly. The pH of the mixture was adjusted to >10 by
gradual addition of 6N NaOH with ice-bath cooling. The suspension
was stirred at room temperature for 30 min. The solid in the suspect-
ion was collected by filtration, washed with water (5 mL × 3) and
dried under vacuum at 45 °C to give the title compound (265 mg,
purity (RCP) of 98.2% and a specific activity of 53 mCi mmol−1. H
1
NMR and 13C NMR were consistent with published spectra.5
[14C2]7-[4-(4-Naphthalen-1-yl-piperazin-1-yl)-butoxy]-3,4-dihydro-
1H-[1,8]naphthyridin-2-one (18, [14C]F-00217830-R): A mixture of
[14C2]7-(4-chloro-butoxy)-3,4-dihydro-1H-[1,8]naphthyridin-2-one
(150 mg, 0.59 mmol, 53 mCi mmol−1), 1-naphthalen-1-yl-piperazine
hydrochloride (161. 2 mg, 0.65 mmol) and potassium carbonate
(284 mg, 2.0 mmol) in water (1.5 mL) was heated at 110 °C for 20 h.

JOURNAL OF CHEMICAL RESEARCH 2012 355
The reaction mixture was cooled to room temperature, then diluted
with ethyl acetate (20 mL) and stirred for 20 min. The organic layer
was separated, and the aqueous layer was extracted with additional
ethyl acetate (10 mL). The combined organic layers were washed with
brine (15 mL) water (15 mL), and dried over sodium sulfate. Concen-
tration under vacuum gave the crude product, which was purified
by flask column chromatography (ethyl acetate:hexane, 1:1). The title
compound was obtained as a white solid (152 mg, 60%, 53 mCi
mmol−1, 18.7 mCi, radiochemical purity (RCP) and chemical purity
(CP): 98.8% and 98.5% by HPLC assay). 1H NMR and 13C NMR were
consistent with published spectra.1 HPLC condition for purity: Water
X-Bridge C18, 4.6 × 50, 5 µm, Mobile Phase A: 0.1% Formic acid;
Mobile Phase B: MeCN, 0% B linear gradient to 80% over 8 min,
hold A:B 20:80 to 10 min. Flow rate = 1.5 mL min−1, UV detection:
210 nm.
67.4(1C), 44.6(1C), 30.1 (1C), 29.1(1C), 26.8(1C), 23.5(1C). HRMS
(ESI) m/z Found 331.9924; calcd for C12H14BrClN2O2: 331.9927.
7-[4-(4-Naphthalen-1-yl-piperazin-1-yl)-butoxy]-3,4-dihydro-1H-6-
bromo-[1,8]naphthyridin-2-one(25):Amixtureof7-(4-chloro-butoxy)-
3,4-dihydro-1H-6-bromo-[1,8]naphthyridin-2-one
24
(140 mg,
0.55 mmol), 1-naphthalen-1-yl-piperazine hydrochloride 8 (160 mg,
0.65 mmol) and potassium carbonate (284 mg, 2.0 mmol) in water
(1.5 mL) was heated at 110 °C for 20 h. The reaction mixture was
cooled to room temperature, then diluted with ethyl acetate (20 mL)
and stirred for 20 min. The organics were separated, and the aqueous
layer was extracted with additional ethyl acetate (10 mL). The com-
bined organics were washed with brine (10 mL) and = water (15 mL),
and dried over sodium sulfate. Concentration under vacuum gave the
crude product, which was purified by flash chromatography (ethyl
acetate:hexane, 1:1). The tilted compound was obtained as a white
solid (173.5 mg, 62%). 1H NMR (CDCl3): δ 8.17 (d, 1H), 7.80 (d, 1H),
7.56 (s, 1H), 7.54 (d, 1H), 7.45 (m, 1H), 7.42 (m, 1H), 7.38 (t, 1H),
7.08 (d, 1H), 4.3 (t, 2H), 3.2 (b, 2H), 2.82 (t, 2H), 2.6 (t, 2H), 1.85 (m,
4H). 13C NMR (CDCl3): δ 170.2, 160.1, 151.2, 150.1, 140.3, 134.9,
129.3, 128.1, 126.4, 125.8, 125.6, 125.0, 123.5, 116.7, 115.2, 98.9,
67.3, 57.5, 53.4 (2C), 50.9 (2C), 30.1, 27.8, 26.0, 23.7. HRMS (ESI)
m/z Found 508.1474, calcd for C26H29BrN4O2: 508.1473.
[14C]1-Bromonaphthalene (20): To a solution of copper (II) bromide
(17.22 g, 0.077 mol) in HPLC grade water (60 mL) was added neutral
alumina (40.0 g, Brockmann I neutral, Aldrich product # 199974) at
room temperature. The water was removed by rotary evaporation at
80 °C under reduced pressure. The resulting dark brown reagent was
further dried under vacuum at 100 °C for 15 h. The mass obtained was
56.867 g (theoretical yield) after drying.
To a 125 mL round bottom flask was added [1-14C]naphthalene
(484 mg, 200 mCi, 3.7 mmol, 54 mCi mmol−1), carbon tetrachloride
(33 mL) and copper (II) bromide-aluminum oxide reagent (6.2 g)
described above. The reaction mixture was heated at 80 °C for 1.5 h.
The reaction was cooled to room temperature and the solids were
removed by filtration. The filtrate was concentrated to give the crude
product as a yellow oil. Purification of the crude material by flash
chromatography (hexane) offered the title compound 20 (498.3 mg,
130.0 mCi, 65%, 54.5 mCi mmol−1, 98.3% RCP by HPLC). 1H NMR
was consistent with that of the authentic sample.
7-[4-(4-Naphthalen-1-yl-piperazin-1-yl)-butoxy]-3,4-dihydro-1H-
[6-3H]-[1,8]naphthyridin-2-one (26, [3H]PF-00217830-R): A flask
was charged with 7-[4-(4-naphthalen-1-yl-piperazin-1-yl)-butoxy]-
3,4-dihydro-1H-6-bromo-[1,8]naphthyridin-2-one (25, 5 mg, 0.0098
mmol), KOAc (3 mg) and Pd(PPh3)4 (5 mg) in DMF (0.5 mL). The
reaction mixture was purged with nitrogen gas for three times and
connected to tritium gas (2 Ci). The resulting mixture was heated at
100 °C for 2 h under tritium gas atmosphere. The solvent was removed
under vacuum. The crude material was purified by prep HPLC to give
[3H]PF-00217830 (50 mCi, RCP and CP>98%, specific activity:
25.3 Ci mmol−1). 1H NMR (CD3OD) δ 7.93 (d, 1H), 7.75 (d, 1H), 7.52
(d, 1H), 7.41 (m, 3H), 7.31 (t, 1H), 7.04 (d, 1H), 4.08 (t, 2H), 3.48 (m,
2H), 3.30 (m, 2H), 3.13 (m, 4H), 2.96 (m, 2H), 2.66 (t, 2H), 2.41 (t,
2H), 1.72 (m, 4H); 3H NMR (CD3OD): δ 6.41. HPLC conditions for
purity: Water X-Bridge C18, 4.6 × 50, 5 µm, Mobile Phase A: 0.1%
formic acid; Mobile Phase B: MeCN, 0% B linear gradient to 80%
over 8 min, hold A:B 20:80 to 10 min. Flow rate = 1.5 mL min−1, UV
detection: 210 nm.
[14C]N-naphthenyl-piperazine (21): In a 125 mL flask was charged
with [14C]1-bromonaphthalene (492.0 mg, 2.37 mmol, 128 mCi,
54 mCi mmol−1) in THF (27 mL), piperazine (304.0 mg, 3.53 mmol),
PXPd catalyst [Dichlorobis(chlorodi-tert-butylphosphine) palladium
(II), 38 mg, 0.071 mmol] and sodium t-butoxide (346 mg, 3.6 mmol).
The reaction mixture was heated at reflux for 60 min. After comple-
tion of the reaction (checked by HPLC), the reaction mixture was
cooled to room temperature, diluted with ethyl acetate (40 mL) and
stirred at room temperature for 20 min. The mixture was filtered and
the filtrate was concentrated under vacuum to give the crude product
as a brown oil. Purification of the crude material by flash chromato-
graphy (ethyl acetate and ethyl acetate:methanol, 4:3) gave the title
compound 21 (326.5 mg, 83.2 mCi, 65%, 54.5 mCi mmol−1). 1H NMR
was consistent with that of the authentic sample.
[14C/3H] PF-00217830 (27): A solution of [14C]PF-00217830 (23,
250 µCi) in MeOH (2.5 mL) was combined with a solution of [3H]PF-
00217830 (26, 2500 µCi) in MeOH (7.5 mL) to form a solution of
[3H/14C]PF-00217830 (250 µCi of 14C and 2500 µCi of 3H in 10 mL of
MeOH) with 3H radiochemical purity of 99.3% and 14C radiochemical
purity of 99.5%. The final specific activity was determined to be
3
13.9 µCi mg−1 for 14C and 140 µCi mg−1 for H. HPLC condition for
[14C] 7-[4-(4-Naphthalen-1-yl-piperazin-1-yl)-butoxy]-3,4-dihydro-
1H-[1,8]naphthyridin-2-one (23, [14C]PF-00217830-L): To a mixture
of [14C]N-naphthenyl-piperazine (21, 184 mg, 0.859 mmol, 46.4 mCi,
54 mCi mmol−1), 7-(4-chloro-butoxy)-3,4-dihydro-1H-[1,8]naphthy-
ridin-2-one (22, 227 mg, 0.893 mmol) and acetonitrile (2.5 mL) was
added a solution of potassium carbonate (356 mg, 2.58 mmol) in
water (9.5 mL) at room temperature. The resulting mixture was heated
at 115 °C for 21 h, and then cooled to room temperature and diluted
with ethyl acetate (35 mL). After the mixture was stirred for 20 min,
the organic layer was separated, and the aqueous layer was extracted
with additional ethyl acetate (15 mL). The combined organic layers
were washed with brine (15 mL) and water (15 mL) and dried over
sodium sulfate. Concentration gave the crude product, which was
purified by a 40 g Redi-Sep silica column (ethyl acetate:hexane, 1:1)
to give the title compound 23 (316.3 mg, 39.9 mCi, 86%, 54.5 mCi
mmol−1) at 99.5% RCP by HPLC (HPLC conditions for purity: Water
X-Bridge C18, 4.6 × 50, 5 µm, Mobile Phase A: 0.1% Formic acid;
Mobile Phase B: MeCN, 0% B linear gradient to 80% over 8 min, hold
A:B 20:80 to 10 min. Flow rate = 1.5 mL min−1, UV detection: 210 nm).
1H NMR and 13C NMR were consistent with published spectra.1
7-(4-Chloro-butoxy)-3,4-dihydro-1H-6-bromo-[1,8]naphthyridin-
2-one (24): A solution of 7-(4-chloro-butoxy)-3,4-dihydro-1H-
[1,8]naphthyridin-2-one 22 (124 mg, 0.486 mmol) and NBS (96 mg,
0.54 mmol) in CHCl3 (3 mL) was heated at 74 °C for 18 h. Upon
completion, the reaction mixture was diluted with CHCl3 (5 mL) and
washed with water (5 mL × 3). Concentration under vacuum gave the
crude product, which was purified by a flash column (50% ethyl ace-
tate in hexane). The tilted compound 24 was obtained as a brown solid
(142 mg, 89%). 1H NMR (CDCl3): δ 7.56 (s, 1H), 4.30 (t, 2H), 3.63 (t,
2H), 2.84 (t, 2H), 2.63 (t, 2H), 1.95 (m, 4H); 13C NMR (CDCl3): δ
170.2 (1C), 160.3(1C), 150.3(1C), 140.6 (1C), 115.3 (1C), 98.93(1C),
purity: Water X-Bridge C18, 4.6 × 50, 5 µm, Mobile Phase A: 0.1%
Formic acid; Mobile Phase B: CH3CN, 0% B linear gradient to 80%
over 8 min, hold A:B 20:80 to 10 min. Flow rate = 1.5 mL min−1, UV
detection: 210 nm.
[14C/14C] PF-00217830 (28): A solution of compound 18 (50.1 mg,
6.17 mCi) and compound 23 (49.8 mg, 6.31 mCi) in MeOH (10 mL)
was stirred at 30 °C for 30 min and then filtered through a 0.2 µm
syringe filter. The filtrate was evaporated to dryness and dried to con-
stant weight under high vacuum to give the dual 14C/14C radiolabelled
compound (99.2 mg, 12.36 mCi, 99.3%) with a RCP of 98.8% and a
specific activity of 53.6 mCi mmol−1.
The authors would like to thank the Groton Radiochemistry
team for their support.
Received 27 January 2012; accepted 24 March 2012
Paper 1201132 doi: 10.3184/174751912X13359587895460
Published online: 4 June 2012
References
1
D. Johnson, C. Choi, L. Fay, D. Favour, J. Repine and A, White, Bioorg.
Med. Chem. Lett., 2011, 21, 2621.
2
F. Kerrigan, C. Martin and G. H. Thomas, Tetrahedron Lett., 1998, 39
2219.
3
4
E. Brenner, R. Schneider and Y. Fort, Tetrahedron, 2002, 58, 6913.
Y. Zhang, Oral presentation at 26th Northeast US Chapter Symposium of
International Isotope Society, NJ, Oct. 2011.
5
K. Banno, T. Fujioka, T. Kikuchi, Y. Oshiro, T. Hiyama and K. Nakagawa,
Chem. Pharm. Bull., 1988, 36, 4377.

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