Stereoselective synthesis of an active metabolite of the potent PI3 kinase inhibitor PKI-179

Article abstract of DOI:10.1021/jo9026269

Chicical Equation Represented The synthesis and stereochemical determination of 1-(4-(4-((1R,5R,6R)-6-hydroxy-3-oxa-8-azabicyclo[3.2.1]octan-8- yl)-6-morpholino-1,3,5-triazin-2-yl)phenyl)-3-(pyridin-4-yl)urea(2), an active metabolite of the potent PI3 kinase inhibitor PKI-179 (1), is described. Stereospecific hydroboration of the double bond of 2,5-dihydro-1H-pyrrole 8 gave the 2,3-trans alcohol 9 exclusively. The configuration of the 3-hydroxyl group in 9 was inverted by an oxidation and stereoselective reduction sequence to give the corresponding 2,3-cis isomer 23. Both exo (21) and endo (27) isomers of the metabolite 2 were prepared via a practical synthetic route from 9 and 23, respectively, and the stereochemistry of 2 was determined to be endo. The endo isomer (27) was separated into two enantiomers 28 and 29 by chiral HPLC. Compound 2 was found to be enantiomerically pure and identical to the enantiomer 28. The absolute stereochemistry of the enantiomer 28 was determined by Mosher's method, thus establishing the stereochemistry of the active metabolite 2.

Full text of DOI:10.1021/jo9026269

pubs.acs.org/joc
Stereoselective Synthesis of an Active Metabolite of the Potent PI3 Kinase
Inhibitor PKI-179
Zecheng Chen,* Aranapakam M. Venkatesan, Osvaldo Dos Santos, Efren Delos Santos,
Christoph M. Dehnhardt, Semiramis Ayral-Kaloustian, Joseph Ashcroft,
Leonard A. McDonald, and Tarek S. Mansour
Chemical Sciences, Wyeth Research, 401 North Middletown Road, Pearl River, New York 10956
chenz1@wyeth.com; chenzecheng@hotmail.com
Received December 11, 2009
The synthesis and stereochemical determination of 1-(4-(4-((1R,5R,6R)-6-hydroxy-3-oxa-8-
azabicyclo[3.2.1]octan-8-yl)-6-morpholino-1,3,5-triazin-2-yl)phenyl)-3-(pyridin-4-yl)urea (2), an ac-
tive metabolite of the potent PI3 kinase inhibitor PKI-179 (1), is described. Stereospecific hydro-
boration of the double bond of 2,5-dihydro-1H-pyrrole 8 gave the 2,3-trans alcohol 9 exclusively. The
configuration of the 3-hydroxyl group in 9 was inverted by an oxidation and stereoselective reduction
sequence to give the corresponding 2,3-cis isomer 23. Both exo (21) and endo (27) isomers of the
metabolite 2 were prepared via a practical synthetic route from 9 and 23, respectively, and the
stereochemistry of 2 was determined to be endo. The endo isomer (27) was separated into two
enantiomers 28 and 29 by chiral HPLC. Compound 2 was found to be enantiomerically pure and
identical to the enantiomer 28. The absolute stereochemistry of the enantiomer 28 was determined by
Mosher’s method, thus establishing the stereochemistry of the active metabolite 2.
Introduction
drugs.2 Many active metabolites, such as atorvastatin
(Liptor),3 simavastatin (Zocor),4 fluoxetine (Prozac),5 fexo-
fenadine (Allegra),6 and cetirizine (Zyrtec),7 have been de-
veloped as drugs in their own right. To support further
studies of metabolite 2, including its in vivo, PK, and
toxicology, we developed a practical route for the prepara-
tion of 2 in gram quantities. We now wish to report on the
synthesis of the active metabolite 2 with high stereoselectivity
and the determination of its absolute stereochemistry.
A potent PI3KR/mTOR dual inhibitor, 1-(4-(4-(3-oxa-8-
azabicyclo[3.2.1]octan-8-yl)-6-morpholino-1,3,5-triazin-2-yl)-
phenyl)-3-(pyridin-4-yl)urea (PKI-179, 1), is currently being
advanced into development for the treatment of solid tumors.1
In vitro studies showed that 1 was metabolically stable in nude
mouse, rat, and dog microsomes, but a major metabolite 2
(Figure 1) was observed when 1 was incubated with human or
monkey liver microsomes.1 Metabolite 2 was isolated and its
molecular structure was determined by 1H NMR and LCMS.
Biological assays showed that compound 2 is an active
metabolite, with in vitro potency comparable to that of the
parent compound 1.1
Results and Discussion
To assemble the whole molecule of 2, the 6-hydroxyl
bridged-morpholine (3) was considered a key building block,
because it could be converted to 2 via a route similar to that
used for the synthesis of 1. Since the configuration and
Pharmacologically active metabolites can contribute sig-
nificantly to the overall therapeutic or adverse effects of
*To whom correspondence should be addressed. Phone: 845-602-8341.
Fax: 845-602-5561.
(1) Mallon, R.; Hollander, I.; Feldberg, L.; Lucas, J.; Soloveva, V.; Gibbons,
J.; Venkatesan, A. M.; Chen, Z.; Dos Santos, O.; Dehnhardt, C. M.; Delos
Santos, E.; Ayral-Kaloustian, S.; Mansour, T. S. Cancer Res. 2009, in press.
(2) For a review, see: Fura, A. Drug Discovery Today 2006, 11, 133–142.
(3) Lennernas, H. Clin. Pharmacokinet. 2003, 42, 1141–1160.
(4) Williams, D.; Feely, J. Clin. Pharmacokinet. 2002, 41, 343–370.
(5) Cheer, S. M.; Goa, K. L. Drugs 2001, 61, 81–110.
(6) Meeves, S. G.; Appajosyula, S. J. Allergy Clin. Immunol. 2003, 112,
S69–S77.
(7) Golightly, L. K.; Greos, L. S. Drugs 2005, 65, 341–384.
DOI: 10.1021/jo9026269
r
Published on Web 01/29/2010
J. Org. Chem. 2010, 75, 1643–1651 1643
2010 American Chemical Society

Chen et al.
JOCArticle
FIGURE 1. Metabolism of PKI-179 (1).
SCHEME 1. Retrosynthetic Analysis of the 6-Hydroxy-Bridged
Morpholine 3
SCHEME 2
SCHEME 3
chirality of the hydroxyl group in 2 was not yet determined,
we devised a synthetic route to the racemic form of 3.
A retrosynthetic analysis of compound 3 (Scheme 1)
suggested that N-Boc-protected cis-2,5-diester 6, available
by a modified Birch reduction under “ammonia-free” con-
ditions,8 could serve as a starting material. Conversion of the
cis diester 6 to the corresponding cis diol 5 by reduction,
followed by ring closure of the cis diol 5 could provide the
bridged morpholine 4. Introduction of the hydroxyl group
through hydroboration of the double bond in 4 should result
in the desired 6-hydroxyl-bridged morpholine 3.
In our initial approach to the bridged morpholine 4,
following Donohoe’s procedure,9 cis diester 6 was prepared
in reasonable yield in two steps from N-Boc-pyrrole. Con-
version of 6 to the corresponding N-Bn-protected cis diol 5
was achieved by replacement of the Boc group in 6 with a
benzyl group followed by reduction with LiAlH4. Heating
the cis diol 5 with 66% H2SO4, followed by basic workup,10
gave the desired bridged morpholine 4 in low yield (Scheme 2).
To improve yield of the cyclization of diol 5 to 4, we screened a
variety of known ring-closure methods,11-16 such as using
SOCl2 (1 equiv) in DMF,11 heating with concd HCl,12 reflux-
ing with triphenylphosphine in CCl4,13 heating with TsCl
(3 equiv) in pyridine,14 or treating with TsCl (1 equiv) in
pyridine followed by NaH workup.15 The desired cyclized
product 4 was not under any of these conditions. The observed
difficulty of the ring closure for diol 5 may be due to the
presence of the double bond in the 5-membered ring, as it has
been shown that the saturated analogue of 5 formed the
corresponding bicyclic product in high yield under the same
conditions.10
(8) Donohoe, T. J.; Rigby, C. L.; Thomas, R. E.; Nieuwenhuys, W. F.;
Bhatti, F. L.; Cowley, A. R.; Bhalay, G.; Linney, I. D. J. Org. Chem. 2006, 71,
6298–6301.
(9) Donohoe, T. J.; Freestone, G. C.; Headley, C. E.; Rigby, C. L.;
Cousins, R. P. C.; Bhalay, G. Org. Lett. 2004, 6, 3055–3058.
(10) Feurer, A.; Flubacher, D.; Weigand, S.; Stasch, J.; Stahl, E.;
Schenke, T.; Alonso-alija, C.; Wunder, F.; Lang, D.; Dembowsky, K.;
Straub, A.; Perzborn, E. PCT WO 2003/004503.
(11) Revesz, L.; Blum, E.; Wicki, R. Tetrahedron Lett. 2005, 46, 5577–
5580.
(12) Newkome, G. R.; Arai, S.; Gupta, V. K.; Griffin, R. W. J. Org.
Chem. 1987, 52, 5480–5482.
Manipulation of the double bond in the 5-membered ring
prior to ring closure and the synthesis of the advanced diol 10
is shown in Scheme 3. Reduction of the cis diester 6 using
LiBH4 in THF resulted in the cis diol 7 in good yield.
(13) Erickson, G. W.; Fry, J. L. J. Org. Chem. 1980, 45, 970–972.
(14) Tadano, K.-I.; Ishihara, J.; Yamada, H.; Ogawa, S. J. Org. Chem.
1989, 54, 1223–1227.
(15) Barhoumi-Slimi, T.; Crousse, B.; Ourevitch, M.; El Gaied, M.;
Begue, J.-P.; Bonnet-Delpon, D. J. Fluorine Chem. 2002, 117, 137–141.
(16) Huck, B. R.; Gellman, S. H. J. Org. Chem. 2005, 70, 3353–3362.
1644 J. Org. Chem. Vol. 75, No. 5, 2010

Chen et al.
JOCArticle
SCHEME 4
SCHEME 5
derived from its precursor diol 10 with the large 3-OBn group
in the trans position relative to the 2-CH2OH group. Hence,
the hydroboration of the double bond in 8 must have
occurred in a stereoselective manner, exclusively from the
less crowded face to form 2,3-trans alcohol 9.
Subsequent protection of both hydroxyl groups with tert-
butyldimethylsilyl chloride (TBSCl) provided the bis-TBS
silyl ether 8,16 and introduction of the 3-hydroxyl group was
achieved by hydroboration of the double bond using
BH3 DMS, followed by oxidation using H2O2/NaOH com-
3
bination.17 Benzyl protection of the resulting 3-hydroxyl
group of 9, followed by removal of both TBS protecting
groups using TBAF, provided the cis diol 10, which served as
a precursor for the ring-closure step.
With the desired diol 10 in hand, we then explored
cyclization conditions (Scheme 4). Heating the diol 10 with
H2SO4 did not yield the desired bicyclic product 11, and the
starting material was destroyed under the harsh conditions.
Similarly, no cyclized products were detected under condi-
tions, such as heating with PPh3/CCl4.13 Replacement the
N-Boc group in 10 with N-Bn (an acid-stable group) gave the
corresponding diol 12. The desired ring-closure product 14
was not observed when compound 12 was heated with
sulfuric acid or with a catalytic amount of pTsOH in toluene,
although a small portion of byproduct 13 was observed
under the later conditions.
FIGURE 2. Determination of the stereochemistry of the cycliza-
tion product 16. The arrow denotes an observed proton-proton
NOE correlation of importance in the exo/endo determination.
With the exo bridged morpholine 16 prepared, the stage
was set for the synthesis of the exo metabolite 21 (Scheme 6).
Compound 17, readily prepared by reaction of 1 equiv of
morpholine with cyanuric chloride, was reacted with 16 at
room temperature to give product 18 in good yield. Suzuki
coupling of 18 with 4-aminophenylboronic acid pinacol ester
under microwave conditions gave aniline 19, which was
converted to 4-pyridylurea 20 by reaction with triphosgene,
followed by addition of 4-aminopyridine. Removal of the
benzyl group in 20 by catalytic hydrogenation in acidic
conditions gave the target compound 21. Compound 21
displayed the same HRMS and UV spectra as that of the
Treatment of diol 10 with TsCl (1 equiv) in pyridine15
resulted in recovered starting material without any tosylate
formation. However, deprotonation of both hydroxyl
groups using NaH (2 equiv) at low temperature, followed
by addition of 1 equiv of TsCl, produced the corresponding
mixture of monotosylate intermediates 15. The mixture of
tosylates was concentrated to remove the solvent (THF), and
the residue was dissolved in DMF and treated with excess
KH. The resulting mixture was heated at 100 °C for 30 min
under microwave conditions, and the desired bicyclic com-
pound 11 was isolated as a single product in good yield.
Through optimization of the reaction conditions, we found
that 15 could be completely converted to the cyclized product
11 after stirring overnight at room temperature without the
need for microwave irradiation. Removal of the Boc group in
11 using TFA gave the corresponding bridged morpholine 16
in quantitative yield (Scheme 5).
1
isolated metabolite 2; however, its H NMR spectrum was
not identical to that of 2. Compounds 2 and 21 were
coinjected in HPLC to give two close but distinct peaks.
On the basis of these findings, it was clear that the synthetic
exo isomer 21, prepared from the exo bridged morpholine 16,
was a stereoisomer of the isolated metabolite 2, suggesting
that 2 had an endo hydroxyl configuration.
Attempted inversion of the configuration of the hydroxyl
group in 21, using Mitsunobu reaction conditions (PPh3,
DEAD and benzoic acid),18,19 failed to give the desired endo
isomer. Tosylation or triflation of the hydroxyl group fol-
lowed by replacement of the resulting TsO or TfO group
The exo configuration of the 6-OBn group in 16 was
confirmed by its NMR studies including 1H-1H COSY,
1H-13C HMBC, and NOE, in particular, the observed NOE
correlation between H-6 (δ = 4.36 ppm) and H-4 (δ = 3.83
ppm) as shown in Figure 2. This exo selectivity must be
(18) Rabiczko, J.; Urbanczyk-Lipkowska, Z.; Chmielewski, M. Tetrahe-
dron 2002, 58, 1433–1441.
(19) Wyrick, S. D.; Booth, R. G.; Myers, A. M.; Owens, C. E.; Kula,
N. S.; Baldessarini, R. J.; McPhail, A. T.; Mailman, R. B. J. Med. Chem.
1993, 36, 2542–2552.
(17) Brown, H. C.; Prasad, J. V. N. V.; Gupta, A. K. J. Org. Chem. 1986,
51, 4296–4298.
J. Org. Chem. Vol. 75, No. 5, 2010 1645

Chen et al.
JOCArticle
SCHEME 6
SCHEME 7. Conversion of the 2,3-Trans Alcohol 9 to the 2,3-
Cis Alcohol 23
SCHEME 8
using KOAc in DMF20,21 or KO2/DMSO22 failed to yield the
desired inversion product. Since attempts to invert the
hydroxyl group of 21 were unsuccessful, attention was
focused on making the endo isomer of the key building block
16. Mitsunobu reaction of the 2,3-trans alcohol 9 failed to
give the desired product. Epoxidation of the double bond in 8
using m-CPBA23 gave an inseparable 1:1 mixture of syn and
anti epoxides. Finally, the desired 2,3-cis alcohol 23 was
readily prepared from the 2,3-trans alcohol 9 by Swern
oxidation24 to ketone 22 followed by stereoselctive reduction
of the ketone from the anti position, using NaBH4, to afford
the desired 2,3-cis alcohol 23, exclusively (Scheme 7).
yield (Scheme 8). The endo configuration of 26 was con-
firmed by comparing 1H-1H COSY, 1H-1H NOE, and
1H-13C HMBC NMR data for 26 to the corresponding
data for the exo bridged-morpholine 16.
From the desired endo bridged-morpholine 26, the endo
isomer 27 was prepared in four steps (26% total yield) by the
same synthetic route that was used for the synthesis of the
exo analogue 21 (Scheme 9).
All the spectral data of the synthetic endo isomer 27 (1H
NMR, 13C NMR, HRMS, and LCMS) werefoundto be iden-
tical with those of the isolated metabolite 2, thus confirming
that 2 bears the endo hydroxyl configuration (Scheme 10).
Therefore, synthesis of the racemic endo metabolite 27 was
accomplished in 13 steps with 4.3% total yield from the
known cis diester 6.
With the desired 2,3-cis alcohol 23 in hand, the endo
bridged-morpholine 26 was prepared in four steps in good
To determine the absolute stereochemistry and enantio-
meric purity of 2, a chiral HPLC method was developed to
separate the enantiomers of the synthetic racemic endo
isomer 27. The enantiomers 28 and 29 were well separated
under chiral HPLC conditions (1:1 ratio), with retention
times of 10 and 13.2 min, respectively. Under the same
conditions, injection of the isolated sample of metabolite 2
(20) Kocalka, P.; Pohl, R.; Rejman, D.; Rosenberg, I. Tetrahedron 2006,
62, 5763–5774.
(21) Shing, T. K.M.; Tai, V. W.-F. J. Org. Chem. 1995, 60, 5332–5334.
(22) Giumanini, A. G.; Roveri, S.; Mazza, D. D. J. Org. Chem. 1975, 40,
1678–1680.
(23) Garcia, A. L. L.; Correia, C. R. D. Tetrahedron Lett. 2003, 44, 1553–
1557.
(24) Degnan, A. P.; Meyers, A. I. J. Org. Chem. 2000, 65, 3503–3512.
1646 J. Org. Chem. Vol. 75, No. 5, 2010

Chen et al.
JOCArticle
SCHEME 9. Synthesis of the Endo Isomer 27 from the Endo-
Bridged Morpholine 26
FIGURE 3. Δδ values (δS-δR, ppm) for the MTPA esters 28a and
28b in ppm (proton NMR spectra measured in DMSO-d6).
SCHEME 10
chemistry of 2 was determined to be endo. The enantiomers
(28 and 29) of the racemic endo isomer 27 were separated by
chiral HPLC, and 2 was confirmed to be a single enantiomer
and identical to the enantiomer 28. The absolute stereochem-
istry of 28 (2) was determined to be (1R,5R,6R) by Mosher’s
method. Further studies including in vivo assessment of the
active metabolite 2 (28) and its PK properties are in progress.
Experimental Section
General Methods. All solvents and reagents were used as
received. 1H NMR and 13C NMR spectra were recorded in the
solvent indicated at 400 and 100 MHz, respectively. Chemical
shifts are reported in parts per million (δ) using tetramethylsi-
lane as the internal standard with coupling constants (J) re-
ported in hertz (Hz). The peak shapes are denoted as follows: s,
singlet; d, doublet; t, triplet; q, quartet; m, multiplet; br, broad;
sp, split peak in 13C NMR. Mass spectra (MS) and high-
resolution mass spectra (HRMS) were measured with a TOF 2
spectrometer. The purity of final compounds was determined by
analytical HPLC. Conditions: ACN/H2O eluent at 1 mL/min
flow (containing 0.05% TFA) at 40 °C, 20 min, gradient 5%
ACN to 95% ACN, monitored by UV absorption at 215 nm. All
final compounds were found to be g95% pure unless otherwise
specified. Reversed-phase HPLC purifications were performed
on a preparative HPLC system (100 mm ꢀ 30 mm column).
Thin-layer chromatography (TLC) was performed on TLC
silica gel 60F254 aluminum sheets and visualized under UV light
or developed by immersion in the solution of 20% phosphomo-
lybdic acid in ethanol or in solution of 0.6% KMnO4 and 6%
K2CO3 in water. Microwave irradiation was performed by using
a Biotage Microwave Reactor (model: Initiator EXP60). The
terms “concentrated” and “evaporated” refer to removal of
solvents using a rotary evaporator at water aspirator pressure
with a bath temperature equal to or less than 40 °C.
in chiral HPLC gave only one peak with retention time at 10
min, consistent with that of 28, suggesting that 2 is an
enantiomerically pure compound.
The absolute stereochemistry of 28 was determined
by Mosher’s method. After converting 28 to the (R)- and
(S)-MTPA esters25 (28a and 28b, respectively) with the
corresponding MTPACl, the protons were assigned and
their Δδ values (δS-δR, ppm) were obtained as shown in
Figure 3. Model projection and determination of the con-
figuration of the R center were done according to the
empirical method outlined by Kusumi et al.25 and provided
the chirality for C6 and subsequently C1 and C5 which
followed from relative stereochemical determination. Com-
bined, these studies indicate that the stereochemistry of 28
should be (1R,5R,6R).
Conclusions
1-tert-Butyl 2,5-dimethyl 1H-pyrrole-1,2,5(2H,5H)-tricar-
boxylate (6, cis diester) was synthesized by following the proce-
dure described in the literature:9 1H NMR (CDCl3, 300 MHz) δ
5.97-5.88 (m, 2H), 5.12 (m, 1H), 5.05 (m, 1H), 3.78 (s, 6H), 1.45
(s, 9H); MS (ESI) m/z 186 (M þ H - Boc).
Hydroboration of the double bond in 2,5-dihydro-1H-
pyrrole 8 gave the 2,3-trans alcohol 9 exclusively. Oxidation
of the 3-OH group in 9 to the corresponding ketone, was
followed by stereospecific reduction by NaBH4 to give the
2,3-cis alcohol 23 as the sole product. Both exo (21) and endo
(27) isomers of the metabolite 2 were prepared via a practical
synthetic route from 9 and 23, respectively, and the stereo-
Synthesis of (1-Benzyl-2,5-dihydro-1H-pyrrole-2,5-diyl)-
dimethanol (5). Step 1. Dimethyl 1-benzyl-2,5-dihydro-1H-pyr-
role-2,5-dicarboxylate (6a, N-Bn-protected cis diester): To a
solution of cis diester 6 (2.0 g, 7 mmol) in CH2Cl2 (20 mL)
was added TFA (4 mL), and the resulting mixture was stirred at
room temperature for 2 h. The mixture was concentrated in
vacuo to remove as much excess TFA as possible, and the
(25) Ohtani, I.; Kusumi, T.; Kashman, Y.; Kakisawa, H. J. Am. Chem.
Soc. 1991, 113, 4092–4096.
J. Org. Chem. Vol. 75, No. 5, 2010 1647

Chen et al.
JOCArticle
residue was dissolved in EtOH (10 mL) and toluene (10 mL).
The solvent was removed again, and the residue was dissolved in
acetone (10 mL), followed by addition of benzyl bromide (1.66
mL, 14 mmol) and K2CO3 (2.9 g, 21 mmol). The mixture was
heated in 60 °C for 5 h and cooled to room temperature. The
solid was filtered off and washed with acetone, and the resulting
filtrate was concentrated to give a residue which was treated
with EtOAc and water. The two phases were separated, and the
aqueous phase was extracted with EtOAc. The combined or-
ganic phases were washed with water and brine and dried
(MgSO4). Removal of solvent gave the crude product 6a (1.92 g,
99%), which was used for next step without purification: MS
(ESI) m/z 276 (M þ H).
aqueous phase was extracted with EtOAc. The combined or-
ganic phases were washed with water and brine and dried
(MgSO4). The organic solvent was removed in vacuo to give
the crude product, which was purified by flash chromatography
on silica gel with EtOAc/Hex (10:90) to give the 8 (7.12 g, 98%):
1H NMR (CDCl3, 300 MHz) δ 5.85 (d, 2H, J = 3.0 Hz), 4.50
(s, 1H), 4.36 (s, 1H), 3.85 (m, 2H), 3.47 (t, 1H, J = 8.0 Hz), 3.35
(t, 1H, J = 8.7 Hz), 1.43 (s, 9H), 0.85 (s, 9H), 0.84 (s, 9H), 0.00 (s,
12H); MS (ESI) m/z 458 (M þ H); HRMS calcd for C23H47NO4-
Si2 (M þ H) 458.3116, obsd 458.3118.
Synthesis of tert-Butyl 2,5-Bis((tert-butyldimethylsilyloxy)-
methyl)-3-hydroxypyrrolidine-1-carboxylate (9). To a solution
of 8 (4.8 g, 10.5 mmol) in THF (50 mL) was added slowly
Step 2. Synthesis of 1-benzyl-2,5-dihydro-1H-pyrrole-2,5-
diyl)dimethanol (5): To a solution of 6a (1.92 g, 7 mmol) in
THF (50 mL) was added slowly LiAlH4 solution (2 M in THF,
10.5 mL, 21 mmol) at 0 °C. The resulting mixture was stirred at
rt for 17 h and then cooled to 0 °C again. Water (1 mL) was
added dropwise to the reaction mixture, followed by addition of
15% NaOH aqueous solution (1 mL) and water (3 mL). The
mixture was stirred at room temperature for 2 h, and the
resulting solid was filtered off and washed with THF. The
organic solvent was removed in vacuo to give the crude product,
which was purified by flash chromatography on silica gel with
EtOAc/Hex/MeOH (50:50:5) to give the diol 5 (0.86 g, 57%) as a
white solid: mp 67-68 °C (EtOAc); 1H NMR (CDCl3, 300
MHz) δ 7.35-7.32 (m, 5H), 5.73 (s, 2H), 3.94 (s, 2H), 3.95-3.92
(m, 2H), 3.50 (dd, 2H, J = 10.9, 1.5 Hz), 3.36 (dd, 2H, J = 10.9,
3.0 Hz), 2.46 (br, 2H); MS (ESI) m/z 220 (M þ H); HRMS calcd
for C13H17NO2 (M þ H) 220.1332, obsd 220.1330.
BH3 DMS solution (2 M in THF, 6.97 mL, 13.9 mmol) at 0 °C.
3
The resulting mixture was stirred at room temperature for 3 h
and then cooled to 0 °C again. NaOH solution (5 M, 12.6 mL,
63.2 mmol) was added to the reaction mixture, followed by
addition of H2O2 (30%, 6.33 mL, 62.0 mmol). The resulting
mixture was stirred for 5 h before diluted with EtOAc. The
organic layer was separated, and the aqueous phase was ex-
tracted with EtOAc. The combined organic phases were washed
with water and brine and dried (MgSO4). The organic solvent
was removed in vacuo to give the crude product, which was
purified by flash chromatography on silica gel with EtOAc/Hex
(30:70) to give the 9 (3.8 g, 77%): 1H NMR (CDCl3, 300 MHz) δ
4.35 (s, 1H), 4.0-3.46 (m, 5H), 3.33 (m, 1H), 2.22-2.10 (m, 1H),
1.89-1.73 (m, 1H), 1.39 (s, 9H), 0.82 (s, 18H), -0.01 (s, 6H),
-0.03 (s, 6H); MS (ESI) m/z 476 (M þ H); HRMS calcd for
C23H49NO5Si2 (M þ H) 476.3222, obsd 476.3218.
Synthesis of tert-Butyl 3-(Benzyloxy)-2,5-bis(hydroxymethyl)-
pyrrolidine-1-carboxylate(10). Step 1. To a solution of 9 (2.515 g,
5.3 mmol) in THF (50 mL) was added NaH (60%, 0.423 g, 10.6
mmol). The mixture was stirred at rt for 30 min, and benzyl
bromide (1.085 g, 6.3 mmol) and TBAI (0.195 g, 0.5 mmol) were
added. The mixture was stirred at rt for 12 h and quenched by
addition of satd NH4Cl solution (20 mL). The mixture was
concentrated in vacuo, and the residue was taken up in water
and EtOAc. The organic layer was separated, and the aqueous
phase was extracted with EtOAc. The combined organic phases
were washed with water and brine and dried (MgSO4). The
organic solvent was removed in vacuo to give the crude product,
which was purified by flash chromatography on silica gel with
EtOAc/Hex (10:90) to give the 3-OBn intermediate 9a (3.0 g,
100%) as colorless oil: 1H NMR (CDCl3, 300 MHz) δ 7.38-7.27
(m, 5H), 4.52 (m, 2H), 4.07 (d, 1H, J = 5.5 Hz), 4.03-3.86 (m,
3H), 3.73-3.55 (m, 2H), 3.45-3.29 (m, 1H), 2.21 (m, 1H), 2.02
(m, 1H), 1.46 (s, 9H), 0.88 (s, 18H), 0.02 (s, 12H); MS (ESI) m/z
566 (M þ H).
Synthesis of 8-Benzyl-3-oxa-8-azabicyclo[3.2.1]oct-6-ene (4).
A mixture of diol 5 (100 mg, 0.46 mmol) and 66% H2SO4 (1.5
mL) was heated at 175 °C for 4 h. The mixture was cooled to
room temperature and basified by addition of 5 M NaOH
aqueous solution at 0 °C. The mixture was extracted with EtOAc
(3 ꢀ 50 mL), and the combined organic phases were washed with
water and brine and dried (MgSO4). The organic solvent was
removed in vacuo, and the residue was purified by flash chro-
matography on silica gel with EtOAc/Hex (20:80) to give 4 (1
mg, 1%) along with 56 mg of recovered starting material.
Compound 4: 1H NMR (CDCl3, 300 MHz) δ 7.39-7.28 (m,
5H), 6.13 (s, 2H), 3.76 (d, 2H, J = 9.8 Hz), 3.53 (s, 2H), 3.41 (dd,
2H, J = 12.4, 2.3 Hz); MS (ESI) m/z 202 (M þ H); HRMS calcd
for C13H15NO (M þ H) 202.1226, obsd 202.1226.
Synthesis of tert-Butyl 2,5-Bis(hydroxymethyl)-2,5-dihydro-
1H-pyrrole-1-carboxylate (7). To a solution of cis diester 6 (6.6
g, 23.1 mmol) in THF (100 mL) was added slowly a LiBH4
solution (2 M in THF, 34.7 mL, 69.4 mmol) at 0 °C. The
resulting mixture was stirred at rt for 3 h and then cooled to
0 °C again. HCl solution (1 M, 30 mL) was added to the reaction
mixture and stirred for 10 min before being diluted with EtOAc.
The organic layer was separated, and the aqueous phase was
extracted with EtOAc. Combined organic phases were washed
with water and brine and dried (MgSO4). The organic solvent
was removed in vacuo to give the crude product, which was
purified by flash chromatography on silica gel with EtOAc/Hex/
MeOH (50:50:10) to give the cis diol 7 (3.8 g, 72%): 1H NMR
(CDCl3, 300 MHz) δ 5.77 (d, 2H, J = 3.0 Hz), 4.70 (s, 1H), 4.59
(s, 1H), 4.06 (d, 1H, J = 11.3 Hz), 3.97 (d, 1H, J = 11.3 Hz), 3.66
(m, 2H), 1.49 (s, 9H); MS (ESI) m/z 230 (M þ H); HRMS calcd
for C11H19NO4 (M þ H) 230.1386, obsd 230.1380.
Step 2. To a solution of the above intermediate 9a (3.0 g, 5.3
mmol) in THF (50 mL) was added slowly of TBAF solution
(1 M in THF, 21.8 mL, 21.8 mmol) at 0 °C. The resulting mixture
was stirred at rt for 6 h and quenched by addition of satd NH4Cl
solution (10 mL). The mixture was concentrated in vacuo, and
the residue was treated with water and EtOAc. The organic layer
was separated, and the aqueous phase was extracted with
EtOAc. The combined organic phases were washed with water
and brine and dried (MgSO4). The organic solvent was
removed in vacuo to give the crude product, which was purified
by flash chromatography on silica gel with EtOAc/Hex/MeOH
1
(50:50:5) to give 10 (1.15 g, 63%): H NMR (DMSO-d6, 300
MHz) δ 7.31 (m, 5H), 4.82 (t, 1H, J = 5.5 Hz), 4.71 (s, 1H), 4.47
(m, 2H), 4.02 (s, 1H), 3.89-3.73 (m, 2H), 3.54-3.40 (m, 3H),
3.22 (m, 1H), 2.04 (m, 2H), 1.39 (s, 9H); MS (ESI) m/z 338 (M þ
H); HRMS calcd for C18H27NO5 (M þ H) 338.1962, obsd
338.1961.
Synthesis of tert-Butyl 6-(Benzyloxy)-3-oxa-8-azabicyclo-
[3.2.1]octane-8-carboxylate (11). To a solution of diol 10 (1.15 g,
3.4 mmol) in THF (50 mL) was added NaH (60%, 0.409 g,
tert-Butyl 2,5-Bis((tert-butyldimethylsilyloxy)methyl)-2,5-di-
hydro-1H-pyrrole-1-carboxylate (8). To a solution of diol 7
(3.57 g, 15.6 mmol) in DMF (15 mL) were added TBSCl (5.16
g, 34.3 mmol) and imidazole (3.18 g, 46.7 mmol). The mixture
was heated at 80 °C for 30 min in a microwave oven (150 w).
Cooled to rt, the mixture was taken up in water (50 mL) and
EtOAc (50 mL). The organic layer was separated, and the
1648 J. Org. Chem. Vol. 75, No. 5, 2010

Chen et al.
JOCArticle
10.2 mmol). The mixture was stirred at rt for 30 min and cooled
to 0 °C. A solution of p-TsCl (0.65 g, 3.4 mmol) in THF (5 mL)
was slowly added to the mixture. The reaction mixture was then
stirred at rt for 12 h and quenched by addition of satd NH4Cl
solution (20 mL). The mixture was concentrated in vacuo, and
the residue was taken up with water and EtOAc. The organic
layer was separated, and the aqueous phase was extracted with
EtOAc. The combined organic phases were washed with water
and brine and dried (MgSO4). The organic solvent was removed
in vacuo to give the crude product, which was purified by flash
chromatography on silica gel with EtOAc/Hex (20:80) to give 11
(716 mg, 66%) as an off-white solid: 1H NMR (CDCl3, 300
MHz) δ 7.35-7.27 (m, 5H), 4.59-4.43 (m, 2H), 4.38-4.07 (m,
3H), 3.73-3.56 (m, 3H), 3.47 (t, 1H, J = 9.8 Hz), 2.27 (m, 1H),
1.96 (m, 1H), 1.48 (s, 4.5H), 1.44 (s, 4.5H); 13C NMR (DMSO-
d6, 100 MHz) δ 153.5 (sp), 138.2, 128.1 (2C), 127.5, 127.4, 127.3,
80.4 (sp), 78.8 (sp), 70.0 (sp), 69.8 (sp), 68.4 (sp), 59.3 (sp),
55.3 (sp), 35.8 (sp), 27.9 (sp, 3C); MS (ESI, m/z) 342 (M þ Na);
HRMS calcd for C18H25NO4 (M þ Na) 342.1676, obsd 342.1670.
MS (ESI) m/z 320 (M þ H); HRMS calcd for C18H25-
NO4 (M þ H) 320.1857, obsd 320.1856. *The observed split
peaks in 13C NMR would be due to the presence of N-Boc group,
which gives rise to rotamers.
11.7, 7.6 Hz), 4.39 (td, 1H, J = 7.1, 2.7 Hz), 3.84-3.62 (m, 11H),
3.55 (d, 1H, J = 13.2 Hz), 2.39 (dd, 1H, J = 13.2, 7.1 Hz), 2.37
(m, 1H); MS (ESI) m/z 418 (M þ H); HRMS calcd for
C20H24ClN5O3 (M þ H) 418.1640, obsd 418.1646.
Synthesis of 4-(4-(6-(Benzyloxy)-3-oxa-8-azabicyclo[3.2.1]-
octan-8-yl)-6-morpholino-1,3,5-triazin-2-yl)aniline (19). To a
vial were charged 18 (200 mg, 0.48 mmol), boronic ester (157
mg, 0.7 mmol), Pd(PPh3)4 (28 mg, 5 mol %), Na2CO3 (2M,
1 mL, 2 mmol), and DME (3 mL). The resulting mixture was
heated at 120 °C for 60 min in a microwave oven (150 W).
Cooled to rt, the mixture was taken up in water and EtOAc. The
organic layer was separated, and the aqueous phase was ex-
tracted with EtOAc. The combined organic phases were washed
with water and brine and dried (MgSO4). The organic solvent
was removed in vacuo to give the crude product, which was
purified by flash chromatography on silica gel with EtOAc/Hex
1
(50:50) to give the 19 (173 mg, 76%) as yellow oil: H NMR
(CDCl3, 300 MHz) δ 8.22 (d, 2H, J = 8.3 Hz), 7.33-7.21 (m,
5H), 6.67 (d, 2H, J = 8.0 Hz), 5.07 (d, 1/2H, J = 7.2 Hz), 4.97 (s,
1/2H), 4.85 (d, 1/2H, J = 7.6 Hz), 4.70 (s, 1/2H), 4.55 (dd, 2H,
J = 18.1, 11.6 Hz), 4.43 (dd, 1H, J = 6.8, 2.3 Hz), 4.01-3.81 (m,
5H), 3.79-3.68 (m, 6H), 2.40 (dd, 1H, J = 13.2, 7.1 Hz), 2.10 (m,
1H); MS (ESI) m/z 475 (M þ H); HRMS calcd for C26H30N6O3
(M þ H) 475.2452, obsd 475.2451.
Synthesis of 6-(Benzyloxy)-3-oxa-8-azabicyclo[3.2.1]octane
(16). To a solution of 11 (400 mg, 1.2 mmol) in CH2Cl2
(10 mL) was added TFA (2 mL). The mixture was stirred at
room temperature for 6 h. The mixture was concentrated in
vacuo, and the residue was taken up in water and CH2Cl2. The
mixture was basified by addition of a solution of Na2CO3. The
organic layer was separated, and the aqueous phase was ex-
tracted with CH2Cl2. The combined organic phases were washed
with water and brine and dried (MgSO4). The organic solvent
Synthesis of 1-(4-(4-(6-(Benzyloxy)-3-oxa-8-azabicyclo[3.2.1]-
octan-8-yl)-6-morpholino-1,3,5-triazin-2-yl)phenyl)-3-(pyridin-
4-yl)urea (20). To a solution of triphosgene (91 mg, 0.3 mmol) in
CH2Cl2 (3 mL) was added a solution of 19 (292 mg, 0.6 mmol) in
CH2Cl2 (3 mL) at rt, followed by addition of triethylamine (0.26
mL, 1.8 mmol). The mixture was stirred at rt for 15 min, and
4-aminopyridine (290 mg, 3.1 mmol) was added. The mixture was
stirred at rt for 6 h and concentrated to give the crude. The crude
was subjected to HPLC separation to give 20 (150 mg, 41%) as a
white solid: 1H NMR (CD3OD, 300 MHz) δ 8.37 (m, 4H), 7.58
(m, 4H), 7.34-7.20 (m, 5H), 5.10 (d, 1H, J = 7.1 Hz), 5.01 (s, 1/
2H), 4.82 (s, 1/2H), 4.59 (q, 2H, J = 12.1 Hz), 4.47 (d, 1H, J = 6.8
Hz), 3.96-3.56 (m, 12H), 2.43 (m, 1H), 2.00 (m, 1H); 13C NMR
(DMSO-d6, 100 MHz) δ 169.1 (sp), 164.6 (sp), 163.9, 151.9, 150.2
(2C), 146.3, 142.3 (sp), 138.2 (sp), 130.4, 129.0 (2C), 128.2, 128.1,
127.9, 127.6, 127.4, 117.5 (2C), 112.3 (2C), 80.6 (sp), 70.1 (sp), 70.0
(sp), 68.5 (sp), 66.0 (2C), 58.6 (sp), 54.6 (sp), 43.2 (2C), 35.9 (sp);
MS (ESI) m/z 595 (M þ H); HRMS calcd for C32H34N8O4 (M þ
H) 595.2775, obsd 595.2764.
1
was removed in vacuo to give 16 (275 mg, 100%): H NMR
(CDCl3, 300 MHz) δ 7.37-7.28 (m, 5H), 4.52 (d, 1H, JAB = 11.7
Hz), 4.46 (d, 1H, JAB = 11.7 Hz), 4.33 (dd, 2H, J = 6.8, 2.3 Hz),
3.77-3.60 (m, 4H), 3.49-3.41 (m, 2H), 3.26 (s, 1H), 2.35 (dd,
1H, J = 13.2, 6.8 Hz), 1.81 (m, 1H); 13C NMR (DMSO-d6, 125
MHz) δ 137.6, 128.1 (2C), 127.8 (2C), 127.5, 77.7, 70.3, 67.3,
66.1, 59.4, 55.6, 34.5; MS (ESI) m/z 220 (M þ H); HRMS calcd
for C13H17NO2 (M þ H) 220.1331, obsd 220.1331.
Synthesis of 4-(4,6-Dichloro-1,3,5-triazin-2-yl)morpholine
(17). To a solution of cyanuric chloride (51.7 g, 0.28 mol) in
CH2Cl2 (200 mL) was added dropwise of a solution of morpho-
line (23.2 g, 0.27 mol) and triethylamine (58.6 mL, 0.42 mol) in
CH2Cl2 (50 mL) at -78 °C during a period of 2 h. After being
stirred for additional 30 min at -78 °C, the reaction mixture was
quenched by addition of water (50 mL) and allowed to reach
at rt. The organic phase was washed with water three times (100
mL each) and dried over MgSO4. The organic solvent was
removed in vacuo to give dichloride 17 (42 g, 64%) as white
solid: 1H NMR (CDCl3, 300 MHz) δ 3.90 (d, 4H, J = 5.3 Hz),
3.76 (d, 4H, J = 5.3 Hz); 13C NMR (DMSO-d6, 75 MHz) δ 169.1
(2C), 163.4, 65.3 (2C), 44.2 (2C); MS (ESI) m/z 235 (M þ H);
HRMS calcd for C7H8Cl2N4O (M þ H) 235.0148, obsd
235.0145.
Synthesis of 1-(4-(4-(6-Hydroxy-3-oxa-8-azabicyclo[3.2.1]-
octan-8-yl)-6-morpholino-1,3,5-triazin-2-yl)phenyl)-3-(pyridin-
4-yl)urea (21, Exo Isomer). A mixture of 20 (150 mg, 0.25 mmol),
Pearlman’s catalyst (66 mg), concd HCl (30%, 0.2 mL, 2 mmol),
and MeOH (10 mL) was taken for hydrogenation under 50 psi at
rt for 12 h. Upon completion, the mixture was filtered through a
pad of Celite and washed with MeOH. The resulting filtrate was
concentrated in vacuo, and the residue was treated with ether. The
resulting white solid was collected by filtration to give 21 (80 mg,
63%): 1H NMR (DMSO-d6, 600 MHz) δ 9.20 (s, 2H), 8.38 (d, 2H,
J = 6.4 Hz), 8.29 (dd, 2H, J = 8.6, 7.1 Hz), 7.57 (d, 2H, J = 8.6
Hz), 7.45 (d, 2H, J = 6.4 Hz), 4.96 (d, 1H, J = 3.7 Hz), 4.93 (d, 1/
2H, J = 8.3 Hz), 4.75 (d, 1/2H, J = 7.1 Hz), 4.52 (s, 1/2H), 4.47
(m, 1H), 4.33 (s, 1/2H), 3.90-3.63 (m, 8H), 3.57-3.45 (m, 3H),
2.31 (m, 1H), 1.75 (m, 1H); MS (ESI) m/z 505 (M þ H); HRMS
calcd for C25H29N8O4 (M þ H) 505.2306, obsd 505.2297.
Synthesis of tert-Butyl 2,5-Bis((tert-butyldimethylsilyloxy)-
methyl)-3-oxopyrrolidine-1-carboxylate (22). To a stirred solu-
tion of oxalyl dichloride (2 M in CH2Cl2, 10.72 mL, 21.4 mmol)
in CH2Cl2 (80 mL) was added dropwise DMSO (3.04 mL, 42.9
mmol) at -78 °C. After being stirred for 15 min, a solution of 9
(3.4 g, 7.1 mmol) in CH2Cl2 (20 mL) was added dropwise to the
reaction mixture. The mixture was stirred at -78 °C for 1 h, and
triethylamine (9.96 mL, 71.5 mmol) was added. The mixture was
stirred for 5 min before the ice bath was removed, and the
Synthesis of 6-(Benzyloxy)-8-(4-chloro-6-morpholino-1,3,5-
triazin-2-yl)-3-oxa-8-azabicyclo[3.2.1]octane (18). To a solution
of dichloride 17 (295 mg, 1.3 mmol) in CH2Cl2 (10 mL) was
added a solution of bridged morpholine 16 (275 mg, 1.3 mmol)
and Et3N (0.5 mL, 3.8 mmol) in CH2Cl2 (2 mL) at 0 °C. After
being stirred at rt for 3 h, the reaction mixture was diluted with
CH2Cl2. The organic phase was washed with water and brine
and dried over MgSO4. The organic solvent was removed in
vacuo to give the crude, which was purified by flash chroma-
tography on silica gel with EtOAc/Hex (30:70) to give 18 (369
mg, 68%) as a yellow oil: 1H NMR (CDCl3, 300 MHz) δ
7.38-7.28 (m, 5H), 4.86 (d, 1/2H, J = 7.5 Hz), 4.80 (d, 1/2H,
J = 7.5 Hz), 4.78 (s, 1/2H), 4.60 (s, 1/2H), 4.52 (dd, 2H, J =
J. Org. Chem. Vol. 75, No. 5, 2010 1649

Chen et al.
JOCArticle
mixture was allowed to reach at rt. The mixture was poured into
satd NaHCO3 solution and extracted with CH2Cl2. The com-
bined organic phases were washed with water and brine and
dried (MgSO4). The organic solvent was removed in vacuo to
give the crude product, which was purified by flash chromatog-
raphy on silica gel with EtOAc/Hex (10:90) to give the ketone 22
(3.02 g, 89%): 1H NMR (CDCl3, 300 MHz) δ 4.24 (s, 1/2H), 4.13
(s, 1/2H), 4.01-3.75 (m, 4H), 3.42 (t, 1H, J = 9.4 Hz), 2.69-2.46
(m, 2H), 1.46 (s, 9H), 0.83 (s, 18H), 0.01 (s, 12H); MS (ESI) m/z
474 (M þ H); HRMS calcd for C23H47NO5Si2 (M þ H)
474.3066, obsd 474.3070.
Synthesis of tert-Butyl 2,5-bis((tert-butyldimethylsilyloxy)-
methyl)-3-hydroxypyrrolidine-1-carboxylate (23, 2,3-Cis Alcohol).
To a solution of ketone 22 (3.02 g, 6.4 mmol) in MeOH (50 mL)
was added NaBH4 (0.482 g, 12.8 mmol) in several potions. The
mixture was stirred at room temperature for 6 h and quenched by
addition of satd NH4Cl solution (10 mL). The mixture was con-
centrated in vacuo, and the residue was treated with water and
EtOAc. The organic layer was separated, and the aqueous phase
was extracted with EtOAc. The combined organic phases were
washed with water and brine and dried (MgSO4). The organic
solvent was removed in vacuo to give the crude product, which was
purified by flash chromatography on silica gel with EtOAc/Hex
(10:90) to give the 2,3-cis alcohol 23 (2.54 g, 84%) as a colorless oil:
1H NMR (CDCl3, 300 MHz) δ 4.33 (m, 1H), 4.04-3.54 (m, 6H),
2.11 (m, 1H), 2.04-1.95 (m, 1H), 1.40 (s, 9H), 0.84 (s, 18H), 0.03
(s, 6H), 0.02 (s, 6H); MS (ESI) m/z 476 (M þ H); HRMS calcd for
C23H49NO5Si2 (M þ H) 476.3222, obsd 476.3220.
g, 3.4 mmol) in THF (50 mL) was added NaH (60%, 0.409 g,
10.2 mmol). The mixture was stirred at room temperature for 30
min and cooled down to 0 °C. A solution of p-TsCl (0.65 g, 3.4
mmol) in THF (5 mL) was slowly added to the mixture. The
reaction mixture was then stirred at room temperature for 12 h
and quenched by addition of satd NH4Cl solution (20 mL). The
mixture was concentrated in vacuo, and the residue was taken
up in water and EtOAc. The organic layer was separated, and
the aqueous phase was extracted with EtOAc. Combined or-
ganic phases were washed with water and brine and dried
(MgSO4). The organic solvent was removed to give the crude
product, which was purified by flash chromatography on silica
gel with EtOAc/Hex (20:80) to give 25 (716 mg, 66%) as an off-
white solid: 1H NMR (CDCl3, 300 MHz) δ 7.39-7.27 (m, 5H),
4.67-4.54 (m, 2H), 4.17-3.87 (m, 4H), 3.82-3.62 (m, 2H), 3.58
(d, 1H, J = 11.0 Hz), 2.39 (m, 1H), 1.88 (dd, 1H, J = 12.8, 4.2
Hz), 1.46 (s, 9H); 13C NMR (CDCl3, 75 MHz) δ 153.5, 138.0,
128.4 (2C), 127.7 (3C), 80.1, 77.2, 72.3, 70.8, 66.1, 56.2 (sp), 54.3
(sp), 33.7 (sp), 28.4 (3C); MS (ESI) m/z 342 (M þ Na); HRMS
calcd for C18H25NO4 (M þ Na) 342.1676, obsd 342.1676.
Synthesis of 6-(Benzyloxy)-3-oxa-8-azabicyclo[3.2.1]octane
(26, Endo-Bridged Morpholine). To a solution of 25 (716 mg,
2.2 mmol) in CH2Cl2 (10 mL) was added TFA (2 mL). The
mixture was stirred at room temperature for 6 h. The mixture
was diluted with CH2Cl2 and basified by addition of a solution
of Na2CO3. The organic layer was separated, and the aqueous
phase was extracted with CH2Cl2. The combined organic phases
were washed with water and brine and dried (MgSO4). The
organic solvent was removed in vacuo to give the product endo-
bridged morpholine 26 (492 mg, 100%): 1H NMR (CDCl3, 300
MHz) δ 7.39-7.27 (m, 5H), 4.68 (d, 1H, JAB = 11.7 Hz), 4.56 (d,
1H, JAB = 11.7 Hz), 4.43 (m, 1H), 4.15-3.96 (m, 3H), 3.68 (d,
2H, J = 9.8 Hz), 3.53 (d, 1H, J = 6.0 Hz), 2.58-2.48 (m, 1H),
2.0 (dd, 1H, J = 13.6, 4.1 Hz); 13C NMR (CDCl3, 75 MHz) δ
137.7, 128.5 (2C), 127.9, 127.8 (2C), 77.3, 72.9, 70.7, 65.9, 56.4,
54.8, 32.8; MS (ESI) m/z 220 (M þ H); HRMS calcd for
C13H17NO2 (M þ H) 220.1332, obsd 220.1329.
Synthesis of tert-Butyl 3-(benzyloxy)-2,5-bis(hydroxymethyl)-
pyrrolidine-1-carboxylate (24). Step 1. To a solution of 23
(2.515 g, 5.3 mmol) in THF (50 mL) was added NaH (60%,
0.423 g, 10.6 mmol). The mixture was stirred at rt for 30 min, and
benzyl bromide (1.085 g, 6.3 mmol) and TBAI (0.195 g, 0.5
mmol) were added. The mixture was stirred at rt for 12 h and
quenched by addition of satd NH4Cl solution (20 mL). The
mixture was concentrated in vacuo, and the residue was taken
up in water and EtOAc. The organic layer was separated, and
the aqueous phase was extracted with EtOAc. Combined or-
ganic phases were washed with water and brine and dried
(MgSO4). The organic solvent was removed in vacuo to give
the crude product, which was purified by flash chromatography
on silica gel with EtOAc/Hex (10:90) to give 23a (3.0 g, 100%) as
colorless oil: 1H NMR (CDCl3, 300 MHz) δ 7.40-7.28 (m, 5H),
4.69-4.60 (m, 1H), 4.55 (d, 1H, J = 11.7 Hz), 4.12-3.92 (m,
2H), 3.88-3.66 (m, 5H), 2.13 (m, 2H), 1.47 (s, 9H), 0.88 (s, 18H),
0.02 (s, 12H); MS (ESI) m/z 566 (M þ H); HRMS calcd for
C30H55NO5Si2 (M þ H) 566.3692, obsd 566.3687.
Synthesis of 1-(4-(4-(6-Hydroxy-3-oxa-8-azabicyclo[3.2.1]-
octan-8-yl)-6-morpholino-1,3,5-triazin-2-yl)phenyl)-3-(pyridin-
4-yl)urea (27, Endo Isomer). Step 1. Synthesis of 6-(benzyloxy)-
8-(4-chloro-6-morpholino-1,3,5-triazin-2-yl)-3-oxa-8-azabicy-
clo[3.2.1]octane (26a, endo): To a solution of dichloride 17
(1.055 g, 4.5 mmol) in CH2Cl2 (50 mL) was added a solution
of endo-bridged morpholine 26 (0.492 g, 2.2 mmol) and Hunig’s
base (1.2 mL, 6.7 mmol) in CH2Cl2 (5 mL) at 0 °C. After being
stirred at rt for 3 h, the reaction mixture was diluted with CH2Cl2.
The organic phase was washed with water and brine and dried over
MgSO4. The organic solvent was removed in vacuo to give the
crude, which was purified by flash chromatography on silica gel
with EtOAc/Hex (30:70) to give 26a (856 mg, 91%) as a yellow oil:
1H NMR (CDCl3, 300 MHz) δ 7.40-7.28 (m, 5H), 4.72-4.43 (m,
4H), 4.16 (m, 1H), 4.10 (d, 1H, J = 10.9 Hz), 3.86-3.63 (m, 11H),
2.42 (m, 1H), 2.0 (dd, 1H, J = 12.8, 4.5 Hz); 13C NMR (CDCl3, 75
MHz) δ 169.8, 164.4, 162.5, 137.9, 128.5 (2C), 127.8, 127.7, 127.6,
76.4, 72.3 (sp), 70.9 (sp), 66.5 (2C), 65.9, 55.9 (sp), 54.0 (sp), 43.8
(2C), 33.4; MS (ESI) m/z 418 (M þ H); HRMS calcd for
C20H24ClN5O3 (M þ H) 418.1640, obsd 418.1638.
Step 2. To a solution of 23a (3.0 g, 5.3 mmol) in THF (50 mL)
was added slowly a TBAF solution (1 M in THF, 21.8 mL, 21.8
mmol) at 0 °C. The resulting mixture was stirred at rt for 6 h and
quenched by addition of satd NH4Cl solution (10 mL). The
mixture was concentrated in vacuo, and the residue was treated
with water and EtOAc. The organic layer was separated, and the
aqueous phase was extracted with EtOAc. The combined or-
ganic phases were washed with water and brine and dried
(MgSO4). The organic solvent was removed in vacuo to give
the crude product, which was purified by flash chromatography
on silica gel with EtOAc/Hex/MeOH (50:50:5) to give 24 (1.15 g,
62%): 1H NMR (CDCl3, 300 MHz) δ 7.39-7.27 (m, 5H), 4.63
(d, 1H, JAB = 11.7 Hz), 4.50 (d, 1H, JAB = 11.7 Hz), 4.18 (m,
1H), 4.04 (br, 1H), 3.98-3.70 (m, 4H), 3.61 (dd, 1H, J = 11.7,
5.7 Hz), 2.37-2.13 (m, 1H), 1.85 (br, 1H), 1.47 (s, 9H); 13C
NMR (CDCl3, 75 MHz) δ 136.1, 128.6 (2C), 128. 1, 127.7 (2C),
81.1, 77.2, 72.1, 66.9, 62.2, 60.4, 58.5, 32.2, 28.4 (3C); MS (ESI)
m/z 338 (M þ H); HRMS calcd for C18H27NO5 (M þ Na)
360.1781, obsd 360.1779.
Step 2. Synthesis of 4-(4-(6-(benzyloxy)-3-oxa-8-azabicyclo-
[3.2.1]octan-8-yl)-6-morpholino-1,3,5-triazin-2-yl)aniline (26b,
endo): To a vial were charged with 26a (856 mg, 2.0 mmol),
boronic ester (673 mg, 3.1 mmol), Pd(PPh3)4 (118 mg, 0.1
mmol), Na2CO3 (2M, 4.1 mL, 8.2 mmol), and DME (8 mL).
The resulting mixture was heated at 120 °C for 30 min in a
microwave oven (150 w). Cooled to rt, the mixture was taken up in
water and EtOAc. The organic layer was separated, and the
aqueous phase was extracted with EtOAc. The combined organic
phases were washed with water and brine and dried (MgSO4). The
organic solvent was removed in vacuo to give the crude product,
Synthesis of tert-Butyl 6-(benzyloxy)-3-oxa-8-azabicyclo-
[3.2.1]octane-8-carboxylate (25). To a solution of diol 24 (1.15
1650 J. Org. Chem. Vol. 75, No. 5, 2010

Chen et al.
JOCArticle
which was purified by flash chromatography on silica gel with
EtOAc/Hex (50:50) to give the 26b (800 mg, 82%) as a yellow oil:
1HNMR(CDCl3, 300 MHz) δ8.18 (d, 2H, J= 8.6 Hz), 7.43-7.28
(m, 5H), 6.68 (d, 2H, J = 8.7 Hz), 4.86-4.51 (m, 4H), 4.20 (m,
1H), 4.10 (d, 1H, J = 10.6 Hz), 3.98-3.61 (m, 13H), 2.46 (m, 1H),
2.01 (m, 1H); 13C NMR (CDCl3, 75 MHz) δ 170.4, 165.2, 164.0,
149.6, 138.2, 130.0 (2C), 128.4 (2C), 127.7 (3C), 127.1, 114.1 (2C),
77.2, 72.3, 70.6 (sp), 66.8 (2C), 65.9 (sp), 55.7 (sp), 53.7 (sp), 43.6
(2C), 33.6; MS (ESI) m/z 475 (M þ H); HRMS calcd for
C26H30N6O3 (M þ H) 475.2452, obsd 475.2448.
The endo isomer 27 (100 mg) was subjected to chiral HPLC
separation under the above conditions to give enantiomers 28
(27 mg) and 29 (24 mg) and a mixture (35 mg) containing both 28
and 29.
1-(4-(4-((1R,5R,6R)-6-Hydroxy-3-oxa-8-azabicyclo[3.2.1]octan-
8-yl)-6-morpholino-1,3,5-triazin-2-yl)phenyl)-3-(pyridin-4-yl)-
urea (28, identical with the metabolite 2): HPLC purity 98.0%;
chiral purity 96.0% ee; [R]25D = -7.5 (DMSO, 1%); 1H NMR
(DMSO-d6, 600 MHz) δ 9.23 (s, 1H), 9.22 (s, 1H), 8.37 (d, 2H,
J = 6.6 Hz), 8.27 (t, 2H, J = 7.7 Hz), 7.56 (d, 2H, J = 8.8 Hz),
7.45 (d, 2H, J = 6.6 Hz), 5.17 (br, 1H), 4.73 (d, 1/2H, J = 7.3
Hz), 4.54 (d, 1/2H, J = 7.3 Hz), 4.51 (d, 1/2H, J = 5.5 Hz), 4.33
(d, 1/2H, J = 5.5 Hz), 4.28 (m, 1H), 4.0 (dd, 1H, J = 18.7, 11.0
Hz), 3.90-3.54 (m, 11H), 2.39 (m, 1H), 1.72 (m, 1H); MS (ESI)
m/z 505 (M þ H); HRMS calcd for C25H28N8O4 (M þ H)
505.2306, obsd 505.2313.
1-(4-(4-((1S,5S,6S)-6-Hydroxy-3-oxa-8-azabicyclo[3.2.1]octan-
8-yl)-6-morpholino-1,3,5-triazin-2-yl)phenyl)-3-(pyridin-4-yl)urea
(29): HPLC purity 97.8%; chiral purity 97.8% ee; 1H NMR
(DMSO-d6, 600 MHz) δ 9.18 (s, 1H), 9.16 (s, 1H), 8.37 (d, 2H,
J = 6.6 Hz), 8.27 (t, 2H, J =7.7 Hz), 7.56 (d, 2H, J = 8.8 Hz), 7.45
(d, 2H, J = 6.6 Hz), 5.17 (br, 1H), 4.73 (d, 1/2H, J = 7.3 Hz), 4.54
(d, 1/2H, J = 7.0 Hz), 4.51 (d, 1/2H, J = 5.5 Hz), 4.33 (d, 1/2H,
J = 5.9 Hz), 4.28 (br, 1H), 4.0 (dd, 1H, J = 18.7, 10.6 Hz),
3.90-3.54 (m, 11H), 2.39 (m, 1H), 1.72 (m, 1H); MS (ESI) m/z 505
(M þ H); HRMS calcd for C25H28N8O4 (M þ H) 505.2306, obsd
505.2315.
Step 3. Synthesis of 1-(4-(4-(6-(benzyloxy)-3-oxa-8-aza-
bicyclo[3.2.1]octan-8-yl)-6-morpholino-1,3,5-triazin-2-yl)phenyl)-
3-(pyridin-4-yl)urea (26c, endo): To a solution of triphosgene
(144 mg, 0.5 mmol) in CH2Cl2 (5 mL) was added a solution of
26b (385 mg, 0.8 mmol) in CH2Cl2 (3 mL) at rt, followed by
addition of triethylamine (0.11 mL, 0.8 mmol). The mixture was
stirred at rt for 15 min, and 4-aminopyridine (382 mg, 4.1 mmol)
was added, followed by addition of triethylamine (0.22 mL, 1.6
mmol). The mixture was stirred at rt for 6 h and concentrated to
give the crude. The crude was subjected to HPLC separation to
give 26c (288 mg, 60%) as a white solid: 1H NMR (DMSO-d6,
300 MHz) δ 9.20 (s, 1H), 9.18 (s, 1H), 8.38 (d, 2H, J = 6.5 Hz),
8.29 (d, 2H, J = 7.9 Hz), 7.57 (d, 2H, J = 9.4 Hz), 7.45 (d, 2H,
J = 6.1 Hz), 7.43-7.28 (m, 5H), 4.78 (m, 1H), 4.61 (m, 2H), 4.18
(m, 1H), 4.0-3.55 (m, 13H), 2.43 (m, 1H), 1.86 (d, 1H, J = 12.5
Hz); 13C NMR (DMSO-d6, 75 MHz) δ 169.2, 164.6, 163.3,
151.9, 150.2 (2C), 146.2, 142.3, 138.4, 130.3, 129.1, 128.2 (2C),
127.7 (2C), 127.5 (2C), 117.5 (2C), 112.3 (2C), 76.7, 71.2 (2C),
66.0 (2C), 65.0, 55.1 (2C), 43.2 (2C), 33.0; MS (ESI) m/z 595
(M þ H); HRMS calcd for C32H34N8O4 (M þ H) 595.2775, obsd
595.2768.
Preparation of (R)- and (S)-MTPA Esters 28a and 28b.25 To a
suspension of 28 (3 mg, 5.95 μmol) in CH2Cl2 (0.5 mL) was
added pyridine (5 μL, 0.06 mmol) to form a solution. The
mixture was cooled to 0 °C, and (S)-3,3,3-trifluoro-2-meth-
oxy-2-phenylpropanoyl chloride (3 mg, 0.012 mmol) was added,
followed by DMAP (0.8 mg, 5.95 μmol). The mixture was stirred
at 0 °C for 5 h and concentrated to remove the solvent, and the
residue was subjected to HPLC separation to give (R)-MTPA
ester (28a) as a white solid (2.6 mg, 61%).
(S)-MTPA ester (28b) was prepared by the method used for
the preparation of 28a by using (R)-3,3,3-trifluoro-2-methoxy-
2-phenylpropanoyl chloride to give a white solid (1.5 mg, 35%).
For the proton chemical shifts and NMR assignments for both
28a and 28b, see Table S1 in the Supporting Information.
Step 4. Synthesis of 1-(4-(4-(6-hydroxy-3-oxa-8-azabicyclo-
[3.2.1]octan-8-yl)-6-morpholino-1,3,5-triazin-2-yl)phenyl)-
3-(pyridin-4-yl)urea (27, endo isomer): A mixture of 26c (288 mg,
0.5 mmol), 10% Pd/C (188 mg), concd HCl (30%, 0.5 mL, 4.9
mmol), and MeOH (20 mL) was hydrogenated under 50 psi at rt
for 24 h. Upon completion, the mixture was filtered through a
pad of Celite and washed with MeOH. The resulting filtrate was
concentrated in vacuo, and the residue was treated with ether.
The resulting white solid was collected by filtration to give 27
(HCl salt, 150 mg, 57%): mp 195-198 °C (MeOH); HPLC
purity 96.4%; 1H NMR (DMSO-d6, 600 MHz) δ 11.05 (s, 1H),
10.25 (s, 1H), 8.63 (d, 2H, J = 7.0 Hz), 8.31 (t, 2H, J = 8.4 Hz),
7.97 (d, 2H, J = 6.3 Hz), 7.64 (d, 2H, J = 8.8 Hz), 4.74 (d, 1/2H,
J = 7.4 Hz), 4.55 (d, 1/2H, J = 7.3 Hz), 4.52 (d, 1/2H, J = 5.8
Hz), 4.34 (d, 1/2H, J = 5.9 Hz), 4.29 (m, 1H), 4.01 (dd, 1H, J =
16.8, 11.0 Hz), 3.95-3.55 (m, 11H), 2.39 (m, 1H), 1.72 (m, 1H);
13C NMR (DMSO-d6, 100 MHz) δ 168.7, 164.4, 162.8, 153.8,
151.2, 141.8 (2C), 141.5, 130.9 (sp), 129.2 (2C), 118.0 (2C), 113.0
(2C), 70.1 (sp), 68.9 (sp), 66.0 (2C), 65.1 (sp), 57.0 (sp), 53.7 (sp),
43.3 (2C), 35.3 (sp); MS (ESI) m/z 505 (M þ H); HRMS calcd for
C25H28N8O4 (M þ H) 505.2306, obsd 505.2300.
Acknowledgment. We thank Dr. Karthick Vishwanathan
for isolation and identification of the active metabolite 2. We
thank GVK-Bioscience for the scale up of compound 6 and
11 and the Pearl River Chemical Technologies group for
compound identification and purity analysis. In particular,
we thank Ms. Mairead N. Young and Mr. Murthy Damarla
for chiral separation of enantiomers of compound 27.
We also thank Drs. Derek Cole, Fangming Kong, and
Xidong Feng for their helpful discussions regarding this
manuscript.
Separation of Racemic Endo Isomer 27 by Chiral HPLC To
Give Enantiomers 28 and 29. Separation conditions: column:
Chiralpak IA 250 ꢀ 4.6 mm; mobile phase: 70% heptane/DEA
30% ethanol; flow: 1.0 mL/min; column temperature: 40°C; UV
wavelength: 215 and 254 nm; retention time: 10.4 min for ent-1
(28) and 13.6 min for ent-2 (29).
Supporting Information Available: 1H and 13C NMR spec-
tra of most of the reported compounds. This material is avail-
able free of charge via the Internet at http://pubs.acs.org.
J. Org. Chem. Vol. 75, No. 5, 2010 1651