Synthesis and pharmacological activity of metabolites of the 5-HT4 receptor antagonist SB-207266

Article abstract of DOI:10.1016/S0968-0896(01)00120-1

Three metabolites of N-[(1-butyl-4-piperidinyl)methyl]-3,4-dihydro-2H-[1,3]-oxazino[3,2_a]indole- 10-carboxamide (SB-207266) (1) were synthesised and their pharmacological activity determined.

Full text of DOI:10.1016/S0968-0896(01)00120-1

Bioorganic & Medicinal Chemistry 9 (2001) 2119–2128
Synthesis and Pharmacological Activity of Metabolites of the
5-HT4 Receptor Antagonist SB-207266
Michael Fedouloff,a,* Frank Hossner,a Martyn Voyle,a Jennie Ranson,b
Jenifer Powles,b Graham Rileyb and Gareth Sangerb
aDepartment of Synthetic Chemistry, Smithkline Beecham Pharmaceuticals, New Frontiers Science Park, Third Avenue,
Harlow, Essex CM19 5AW, UK
bDepartment of Neuroscience Research, Smithkline Beecham Pharmaceuticals, New Frontiers Science Park, Third Avenue,
Harlow, Essex CM19 5AW, UK
Received 18 December 2000; accepted 9 April 2001
Abstract—Three metabolites of N-[(1-butyl-4-piperidinyl)methyl]-3,4-dihydro-2H-[1,3]-oxazino[3,2_a]indole-10-carboxamide (SB-
207266) (1) were synthesised and their pharmacological activity determined. # 2001 Elsevier Science Ltd. All rights reserved.
Introduction
SB-207266, N-[(1-butyl-4-piperidinyl)methyl]-3,4-dihy-
dro-2H-[1,3]-oxazino[3,2-a]indole-10-carboxamide (1) is
a potent and selective 5-HT4 receptor antagonist with
possible therapeutic indications proposed in a number
of areas such as control of atrial arrhythmia or gastro-
intestinal disorders.1,2a This compound is metabolised
by de-alkylation (loss of butyl side-chain), gamma
hydroxylation on the butyl-side chain and hydroxyla-
tion at positions C-7 and C-4 to give 2, 3, 4 and 5
respectively. The synthesis and pharmacological activity
of the ‘des-butyl’ metabolite 2 has been described pre-
viously.2a We now wish to report the synthesis of 3, 4
and 5 together with their pharmacological activity as
selective 5-HT4 receptor antagonists.2b,c
*Correspondingauthor. Tel.:+44-1279-622-346; fax: +44-1279-622-
348; e-mail: Michael_fedouloff@sbphrd.com
0968-0896/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(01)00120-1

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M. Fedouloff et al. / Bioorg. Med. Chem. 9 (2001) 2119–2128
Results and Discussion
intermediate, the oxazinoindole ester 12, was prepared
by treatment of methyl indole-3-carboxylate 10 with
NCS in dichloromethane in the presence of 1,4-diazabi-
cyclo[2.2.2]octane (DABCO) followed by chloro-
propanol to give the intermediate 11 (64% yield) which
was cyclised by treatment with aqueous sodium
hydroxide solution in toluene giving 12 in 96% isolated
yield.
Synthesis
Synthesis of (Æ)-3,4-dihydro-N-[1-(3-hydroxybutyl)-4-
piperidinyl]methyl] - 2H - [1,3]oxazino[3,2_a]indole - 10-
carboxamide (3)
The synthesis of the metabolite 3 is shown in Scheme 1.
The synthesis of the 3-benzyloxyiodobutane 6 has been
described elsewhere.3 Treatment of 6 with iso-nipecota-
mide 7 in toluene in the presence of potassium carbon-
ate gave the amide 8 in 64% yield. This was reduced
with lithium aluminium hydride in THF to furnish the
key intermediate 9 in a yield of 96%. The second key
The couplingof 9 and 12 was achieved by treatment of a
mixture of the two intermediates in toluene with tri-
methylaluminium at reflux. The benzyl protected meta-
bolite 13 was isolated in 60% yield after purification by
column chromatography. Final de-protection was
achieved by hydrogenation of the oxalate salt of 13 in
Scheme 1. Reagents and conditions: (i) NCS, DABCO, CH2Cl2, 0 ꢀC, 10 min; (ii) HO(CH2)3Cl, CH2Cl2, MeSO3H, 0 ꢀC, 68%; (iii) NaOH(aq),
toluene, 99%; (iv) K2CO3, toluene, 64%; (v) LiAlH4, THF, 5 ꢀC, 1 h, 96%; (vi) AlMe3, toluene, reflux, 5 h, 60%; (vii) EtOH, AcOH, (CO2H)2, H2,
10% Pd/C, 50 psi, 45 ꢀC, 30 h, 40%.

M. Fedouloff et al. / Bioorg. Med. Chem. 9 (2001) 2119–2128
2121
ethanol/acetic acid at 45 ꢀC and 50 psi for 30 h giving 3
in a yield of 40% after crystallisation from iso-propa-
nol/diethyl ether.
the acetoxy protecting group was thought unsuitable for
the rest of the synthesis, benzyloxy protection was
restored by hydrolysis to give 20 in 58%, followed by
treatment with benzylchloride in DMF/K2CO3 to give
21 in 70%. Couplingwith the amine 22 was achieved as
described in part (a) above to give 23 in 54% yield after
crystallisation from ethyl acetate. Final deprotection
was achieved by hydrogenolysis over 10% Pd/C in
methanol, giving the final product 4 in 84% after
recrystallisation from ethyl acetate.
Synthesis of N-[1-butyl-4-piperidinyl)methyl]-3,4-dihydro-
7-hydroxy-2H-[1,3]oxazino[3,2-a]indole-10-carboxamide
(4)
The synthesis of the 7-hydroxylated metabolite 4 is
shown in Scheme 2.
The synthesis of 6-benzyloxyindole 14, followingLeim-
gruber chemistry has been described elsewhere.4 Treat-
ment of 14 with trichloroacetyl chloride in THF in the
presence of pyridine gave the 3-trichloroacetylindole 15,
which was converted to the correspondingbenzyloxy
protected methyl ester 16 in 81% overall yield by treat-
ment in methanol with a catalytic amount of potassium
hydroxide.5 Attempted formation of the oxazino ring
from 16 failed, presumably due to the electron rich nat-
ure of the indole nucleus. However, exchanging the
protecting group to acetoxy by hydrogenolysis in
methanol/THF giving 17 in 81% followed by acety-
lation with acetic anhydride in THF/pyridine giving 18
in 99%, allowed successful construction of the oxazino
ring. Thus, treatment of 18 with NCS in dichloro-
methane in the presence of DABCO followed by
addition to a solution of 3-chloropropanol and metha-
nesulphonic acid in dichloromethane gave 19, con-
taminated with 3-chloropropanol, in a yield of 92%. As
Synthesis of (Æ)-N-[1-butyl-4-piperidinyl)methyl]-3,4-
dihydro - 4 - hydroxy - 2H - [1,3]oxazino[3,2 - a]indole - 10-
carboxamide (5)
The synthesis of the 4-hydroxylated metabolite 5 is
shown in Scheme 3.
Thus, treatment of methyl indole-3-carboxylate 10 with
NCS in dichloromethane in the presence of 1,4-diazabi-
cyclo[2.2.2]octane (DABCO) followed by propane-1,3-
diol gave the required 2-alkoxylated indole derivative 24
in 45% yield together with 12% of the dimeric indole
25. TPAP6 oxidation, which is usually selective for pri-
mary alcohols, gave the aminal 26 in 28% yield, but also
resulted in some over-oxidation to give 11% of the
amidoester 27. The secondary hydroxyl function in 26
was protected as the TBDMS ether by treatment with
TBDMS triflate and 2,6-lutidine in 69% yield, and the
resultingester 28 was coupled with the amine 22 to give
Scheme 2. Reagents and conditions: (i) ClCOCCl3, pyridine, THF, 2 ꢀC, 1 h, 20 ꢀC, 16 h, 100%; (ii) KOH(aq), MeOH, reflux, 5 h, 20 ꢀC, 16 h, 81%;
(iii) MeOH, H2, 10% Pd/C, 5 days, 81%; (iv) THF, pyridine, Ac2O, 4 ꢀC, 16 h, 99%; (v) NCS, DABCO, CH2Cl2, 0 ꢀC, 10 min; (vi) HO(CH2)3Cl,
CH2Cl2, MeSO3H, 0 ꢀC; (vii) Acetone, K2CO3, 20 ꢀC, 16 h, 92%; (viii) MeOH, H2O, NaHCO3, 40 ꢀC, 2 h, 58%; (ix) BnCl, K2CO3, DMF, 100 ꢀC,
6 h, 70%; (x) AlMe3, toluene, reflux, 5 h, 54%; (xi) MeOH, H2, 10% Pd/C, 28 h, 84%.

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M. Fedouloff et al. / Bioorg. Med. Chem. 9 (2001) 2119–2128
Scheme 3. Reagents and conditions: (i) NCS, DABCO, CHCl3, 5 ꢀC, 30 min; (ii) HO(CH2)3OH, MeSO3H, 16 ꢀC, 30 min, 45% (24), 12% (25); (iii)
CH2Cl2/MeCN, N-methylmorpholine-N-oxide, TPAP, 4 A mol sieves, 20 ꢀC, 16 h, 28% (26), 11% (27); (iv) CH2Cl2, lutidine, TBDMS triflate,
À70 ꢀC, warm to 20 ꢀC; (v) MeOH, À70 ꢀC, 69%; (vi) AlMe3, toluene, reflux, 4 h, 89%; (vii) AcOH, H2O, 75 ꢀC, 6.5 h, 83%.
the TBDMS protected metabolite 29 in 89% yield.
Havingtried and failed to de-protect 29 usingtetra-
butylammomium fluoride in THF, final de-protection
was achieved by treatment of 29 with aqueous acetic
acid giving 5 in a yield of 94%.
ried out at 37 ꢀC and Ki were derived from the IC50 as
11
described by Chengand Prusoff.
pKi was defined as
Àog10[Ki]. The results are shown in Table 1.
The rank-order affinities of SB-207266-A and its meta-
bolites for the 5-HT4 receptor were: 5=3=SB-207266-
A>2>4. SB-207266-A and metabolites 5, 4 and 3 dis-
played low affinity (pKi <6.0) for a range of human
recombinant 5-HT receptors. For metabolite 2 the pKi
was <6.0 for all of the tested receptors except 5-HT2A
[6.02], 5-HT2B [6.27] and 5-HT2C [6.11]).
Pharmacological Activity
Radioligand binding affinity and selectivity for the 5-HT4
receptor
The affinity of each metabolite for recombinant human
5-HT1A, 5-HT2A, 5-HT2B, 5-HT2C, 5-HT7 receptors
expressed in human embryo kidney (HEK) 293 cells; 5-
HT1B, 5-HT1D, 5-HT1E, 5-HT1F, 5-HT7 and adrenergic
alpha1B receptors expressed in Chinese hamster ovary
(CHO) cells; and 5-HT6 receptors expressed in HeLa
cells, together with the 5-HT4 receptor from guinea pig
was determined usingradioliagnd bindingassays, as
described by Kennett et al.10 All incubations were car-
5-HT4 receptor antagonism
To evaluate their functional activity as 5-HT4 receptor
antagonists, we used the guinea-pig isolated distal colon
longitudinal muscle-myenteric plexus (LMMP) pre-
paration,7 in which SB-207266-A has previously been
shown to be a highly potent 5-HT4 receptor antagonist.1
In this model, 5-HT4 receptor antagonism is assessed by
measuringthe ability of a compound to inhibit 5-HT-

M. Fedouloff et al. / Bioorg. Med. Chem. 9 (2001) 2119–2128
2123
Table 1. pKi values obtained from inhibition of radioligand binding to human recombinant and guinea pig brain receptors. mean ÆSEM (n)
Receptor
SB-207266-A
2
3
4
5
5-HT1A
5-HT1B
5-HT1D
5-HT1E
5-HT1F
5-HT2A
5-HT2B
5-HT2C
5-HT4
<5 (3)
<6 (3)
<6 (3)
<5 (3)
<5 (3)
<5 (3)
<5.2 (3)
<5.2 (3)
<5 (3)
<5 (3)
6.05Æ0.05 (4)
6.27Æ0.05 (3)
6.10Æ0.03 (6)
7.17Æ0.05 (3)
<5 (3)
<5 (3)
<6 (3)
<5.5 (3)
<5 (6)
<5 (6)
<6 (3)
<5 (3)
<6 (3)
<5.5 (3)
<5 (3)
<5 (3)
5.5 (3)
<5 (3)
5.5 (3)
<5 (3)
<5.5 (3)
<5.5 (3)
<5 (3)
<5 (3)
<5 (3)
<6 (6)
6.34Æ0.10 (5)
<6 (3)
<6 (3)
<5 (3)
<5 (3)
<6 (6)
8.75Æ0.05 (11)
<5 (3)
9.08Æ0.09 (3)
6.05Æ0.05 (3)
9.09Æ0.03 (6)
5-HT6
5-HT7
<5 (3)
<5 (3)
<5 (3)
<5 (3)
<5 (3)
<5 (3)
<5 (3)
<5 (3)
Adrenergic a1B
<5 (3)
<5 (3)
<5.5 (6)
<5 (3)
<5 (3)
evoked, neuronally-mediated contractions of the mus-
cle, in the presence of methiothepin and granisetron, to
block 5-HT2/5-HT1-like and 5-HT3 receptors, respec-
tively. The selectivity of such inhibition is then assessed
by determiningthe ability of the compound to inhibit
similar neuronally-mediated contractions evoked by the
nicotinic receptor agonist 1,1-dimethyl-4-phenyl-piper-
azinium iodide (DMPP).
to equilibrate with the tissue for 45 min before con-
struction of the second concentration–effect curve.
Responses were expressed as meanÆstandard error (n
animals). All agonist concentration–response curves
were fitted to the followingthree parameter loigstic
equation:
n
E ¼ ꢀ=1 ð½AŠ=EC50Þ
8
Male Dunkin Hartley guinea-pigs 200–300 g (Charles
River U.K. Ltd) were used. The colon was removed
approximately 10 cm from the anus and 2–3 cm long
sections of LMMP were dissected as described pre-
viously7. The preparations were suspended under 1 g
tension in tissue baths and were bathed in Krebs–Hen-
seleit solution (mM: NaCl 118, KCl 4.7, KH2PO4 1.2,
MgSO4.7H2O 1.2, glucose 11.1, NaHCO3 25,
CaCl2.6H2O 2.5) containingrganisetron (1 Â10À6 M)
and methiothepin (1Â10À7 M), bubbled with 5% CO2 in
oxygen and maintained at 37 ꢀC. Muscle contractions
were measured usingisotonic transducers connected to
Lectromed Multitrace 8 chart recorders. SB-207266-A
was dissolved in double distilled water to a stock con-
centration of 1Â10À3 M. Compound 2 was dissolved
in DMSO. Compounds 4 and 5 were dissolved in
methanol. Compound 3 was dissolved in 70% ethanol.
All metabolite stock solutions were at 1Â10À2 M.
Further dilutions were carried out in double distilled
water.
usingRobofit. a, [A] and n represent the maximum
response, agonist concentration and curve mid point
slope factor, respectively. The EC50 is the concentration
of agonist that produces 50% of the maximal response.
Where the maximum response to the agonist was not
significantly reduced by the antagonist, affinity esti-
mates for the antagonist were expressed as pKB values
calculated accordingto the method of Arunlakshana
and Schild.9 As previously reported,1 higher concentra-
tions of SB-207266-A reduced the maximum response to
5-HT and some of the metabolites had a similar effect.
For this reason, the term ‘apparent pKB’ is used.
In this model of the 5-HT4 receptor, none of the com-
pounds affected the tone of the tissue in the absence of
5-HT, indicatinga lack of intrinsic activity at the 5-HT 4
receptor, at the concentrations tested. Further, when
tested at the high concentration of 1Â10À8 M, each
compound had little-or-no ability to antagonise the
neuronally-mediated contractions evoked by the nico-
tinic receptor agonist DMPP (Table 2).
After preparation, tissues were left to stabilise for 30
min. Pargyline (1Â10À4 M), a monoamine oxidase inhi-
bitor, was added to the solution bathingthe tissues and
washed out 15 min later. Tissues were then sensitised to
5-HT as described,7 or to DMPP (1Â10À6 M, 30 s
exposure, repeated at intervals of 15 min until a con-
sistent response was obtained). Agonist concentration–
effect curves were constructed non-cumulatively by
addingincreasingconcentrations in half logunit incre-
ments at 15 min intervals. Each agonist was left in con-
tact with the tissue until a maximum response was
obtained but not for longer than 30 s. Two agonist
concentration–effect curves were constructed in each
tissue, the first in the absence of test compound and the
second in the presence of the appropriate concentration
of test compound (1Â10À8 M for all experiments with
DMPP). In all experiments, the test compound was left
However, low concentrations of SB-207266-A (1–
30 nM) caused a parallel rightward displacement of the
5-HT concentration–effect curve, with some depression
of the maximum response to 5-HT at the higher con-
centrations of SB-207266, as reported previously.1
A
Schild plot of these results yielded an apparent pA2 of
9.0. The slope of this plot was 1.42, the difference from 1
probably reflectingthe depression of maximum
response to 5-HT seen at the higher concentrations of
SB-207266-A. The apparent pKB determined from the
mean pEC50 values in the absence and presence of
1Â10À8 M SB-207266-A was 9.6 (Table 3).
Compounds 2 and 4 each produced no significant effect
on the 5-HT concentration–effect curve when tested at

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M. Fedouloff et al. / Bioorg. Med. Chem. 9 (2001) 2119–2128
Table 2. Effects of SB-207266-A and its metabolites on DMPP-
evoked contractions of guinea-pig LMMP preparationa
tor and as for SB-207266-A, acted as potent, surmoun-
table and selective antagonists at this receptor; in the
guinea-pig distal colon LMMP model of the receptor,
the pKi and apparent pKB values were, respectively, 9.09
and 9.08, and 8.5 and 9.3. These values are of the same
order as that of SB-207266-A. Two other metabolites
tested, 2 and 4, were weaker 5-HT4 receptor antagonists
(pKi and pKB values of 7.17 and 6.05, and 7.4 and 5.6,
respectively). None of the compounds exhibited high
affinity for a range of other 5-HT and non-5-HT recep-
tors. Further, when tested in the guinea-pig colon
LMMP preparation at 10À8 M, none of the compounds
markedly inhibited the contractions induced by the
nicotinic agonist DMPP, indicating that when 5-HT4
receptor antagonism was observed in this preparation,
this was unlikely to be mediated via a direct action on
the cholinergic neurones.
Compound
pEC50,
1st concentration–
effect curve
2nd concentration–
effect curve
SB-207266-A
5.5Æ0.1
5.9Æ0.1
5.5Æ0.3
5.8Æ0.1
5.9Æ0.1
5.4Æ0.3 (n=2)
5.3Æ0.1 (n=2)
5.2Æ0.3 (n=2)
5.6Æ0.2 (n=2)
5.7Æ0.1 (n=2)
2
4
5
3
apEC50 values for DMPP were derived from the first concentration–
effect curve in the absence of test compound, and the second curve in
the presence of test compound at 1Â10À8 M.
1Â10À8 M but caused small rightward shifts at 1Â10À6
M [surmountable for (2)], correspondingto an apparent
pKB of 7.4 and 5.7, respectively (Table 3). Compound 5
at 1Â10À8 M produced a rightward shift of the 5-HT
concentration–effect curve, with an apparent pKB of 8.5
(Table 3). Similar results were obtained using 3 1Â10À8
M, yieldingan apparent p KB of 9.3 (Table 3). Like SB-
207266-A tested in the same experiment, 3 produced
some depression of the maximum effect of 5-HT.
Experimental
All experiments were performed under dry nitrogen.
THF was dried by distillation from sodium/benzophe-
none. All other solvents were used without further pur-
ification. Separations by column chromatography were
12
The present results confirmed that SB-207266-A is a
potent antagonist of 5-HT4 receptor-mediated contrac-
tions in the LMMP preparation, which, at concentra-
tions up to 1Â10À8 M, acted in an apparently
competitive manner. Of the four human-derived meta-
bolites of SB-207266-A tested here, one (4) was a very
weak antagonist of 5-HT-evoked contractions in the
LMMP preparation and two (5 and 2) were only mod-
erately potent antagonists with apparent pKB’s 5–10-
fold lower than that of the parent compound. Com-
pound 3 was the most potent of the metabolites tested
with an apparent pKB similar to that of SB-207266-A
when tested in the same experiments. Like SB-207266-
A, 3 also caused some depression of the maximum
response to 5-HT when present at 10À8 M. This lack of
surmountability of the effect of SB-207266-A has been
ascribed to pseudoirreversible antagonism.1
achieved usingconditions described by Still.
Thin
layer chromatograms were run on 0.25 mm Merck pre-
coated plates of silica gel 60 F254. High performance
liquid chromatograms (HPLC) were obtained from C-
18 reverse phase Hichrom RPB columns. Analytical
samples were dried in a Buchi dryingpistol at 40 ꢀC.
NMR spectra were obtained from
a
Bruker
amx400 MHz spectrometer with tetramethylsilane as an
internal standard. Mass spectra were recorded on a
Micromass 70-VSEQ double focusingspectrometer.
Meltingpoints were obtained on a Mettler FP62 system.
Preparation of 1-[3-(phenylmethoxy)butyl]-4-piperidine-
carboxamide (8). A suspension of iso-nipecotamide 7
(18 g, 140 mmol), anhydrous potassium carbonate
(58.2 g, 420 mmol) and 3-benzyloxyiodobutane 6 (44.7 g,
150 mmol) was heated under nitrogen at reflux for 7 h.
The resultingmixture was subjected to hot filtration and
the filtrate evaporated under reduced pressure to give a
solid. This was re-dissolved in ethyl acetate, washed
with water and brine, and concentrated until crystal-
lisation occurred. The solid was collected by filtration
and washed with ethyl acetate and diethyl ether to give 8
Conclusions
Two human-derived metabolites of SB-207266-A, 5 and
3, were found to have high affinity for the 5-HT4 recep-
Table 3. Effects of SB-207266-A and its metabolites on 5-HT-evoked contractions in the guinea-pig LMMP preparationa
Compound (concentration)
pEC50
Apparent pKB
1st concentration–effect curve
2nd concentration–effect curve
None
SB-207266-A (1Â10À8 M)
8.8Æ0.1
8.7Æ0.1
8.6Æ0.2
8.2Æ0.2
8.4Æ0.1
8.4Æ0.2
8.2Æ0.1
9.0Æ0.1
na
7.2Æ0.1 (n=8)
7.2Æ0.1 (n=6)
7.9Æ0.1 (n=4)
7.7Æ0.1 (n=7)
7.4Æ0.2 (n=8)
6.9Æ0.1 (n=10)
9.6Æ0.1
7.4Æ0.1
5.7Æ0.4
8.5Æ0.2
9.0Æ0.1
9.3Æ0.1
2 (1Â10À6 M)
4 (1Â10À6 M)
5 (1Â10À8 M)
SB-207266-A (1Â10À8 M)
3 (1Â10À8 M)
aThe apparent pKB for each compound was calculated from the pEC50 values for 5-HT derived from the first concentration–effect curve, determined
in the absence of test compound, and the second curve in the presence of test compound at the concentration indicated. The effects of SB-207266-A
were tested twice, each in parallel with different metabolites (2, 4, 5 and again to compare with 3), as indicated.

M. Fedouloff et al. / Bioorg. Med. Chem. 9 (2001) 2119–2128
2125
26.1 g(64%). 1H NMR (CDCl3) d 7.4–7.2 (5H+CHCl3,
m), 5.5 (2H, NH2, br s), 4.5 (2H, AB quartet), 3.6ꢀ (1H,
m), 2.9 (2H, m), 2.4 (2H, m), 2.15 (1H, m), 2.0–1.6 (8H,
m), 1.2 (3H, d, J=6.6 Hz); m/z 291 (ES+, M+1).
ene, 18 mL) was diluted with toluene (18 mL) and the
solution cooled to 0 ꢀC. The amine 9 (9.2 g, 33 mmol)
dissolved in toluene (30 mL) was added over 3 min, fol-
lowed by the ester 12 (7.6 g, 33 mmol). The mixture was
heated at reflux for 5 h, then allowed to cool, and
sodium hydroxide solution (10% w/w, 80 mL) added
drop-wise. The toluene layer was washed with water,
brine and evaporated under reduced pressure to give an
oil (14.7 g). This was purified by flash chromatography
on silica (elutingwith 0–20% MeOH/CH 2Cl2) obtaining
13, 9.3 g(60%) as an oil. 1H NMR (CDCl3) d 8.3 (1H,
d, J=7.9 Hz), 7.4–7.1 (8H+CHCl3, m), 6.5 (1H, NH, t,
J=5.9 Hz), 4.5 (4H, t, J=5.3 Hz), 4.1 (2H, t,
J=6.6 Hz), 3.9 (1H, m), 3.3 (2H, t, J=6.6 Hz), 3.0 (2H,
m), 2.5–2.3 (4H, m), 2.1–1.6 (7H, m), 1.5–1.2 (5H, m);
Rf 0.7 (SiO2, 5:1:1 EtOAc/MeOH/NH4OH); m/z 476
(ES+, M+1).
Preparation of 1-[3-(phenylmethoxy)butyl]-4-piperidine-
methanamine (9). To a suspension of LiAlH4 (16.82 g,
440 mmol) in THF (250 mL) at 5 ꢀC was added the
amide 8 (10.34 g, 36 mmol) dissolved in THF (150 mL)
drop-wise over 1 h. The mixture was allowed to warm to
ambient temperature, then stirred for 18 h under nitro-
gen. The mixture was cooled to 5 ꢀC, aqueous sodium
hydroxide solution (7.8 M, 33 mL) added drop-wise over
30 min, and the mixture filtered through Celite. The
Celite was washed with THF (3Â50 mL) and the com-
bined filtrate and washings evaporated under reduced
pressure to give 9 as an oil, 9.4 g(96%). 1H NMR
(CDCl3) d 7.4–7.2 (5H+CHCl3, m), 4.5 (2H, AB quar-
tet), 3.6 (1H, m), 2.9 (2H, m), 2.6 (2H, d, J=6.6 Hz), 2.4
(2H, m), 2.0–1.2 (14H, m); m/z 277 (ES+, M+1).
Preparation of 3,4-dihydro-N-[1-(3-hydroxybutyl)-4-pi-
peridinyl]methyl]-2H-[1,3]oxazino[3,2-a]indole-10-carbox-
amide (3). To a solution of 13 (8.2 g, 17 mmol) in
ethanol (150 mL) and glacial acetic acid (8 mL) was
added oxalic acid dihydrate (2.4 g, 19 mmol), and the
mixture hydrogenated over 10% Pd/C (4.8 g) at 45 ꢀC,
50 psi for 30 h. The mixture was allowed to cool, filtered
through Celite, and the filtrate evaporated under
reduced pressure. The residue was partitioned between
chloroform (150 mL) and concentrated aqueous potas-
sium carbonate solution (100 mL). The chloroform layer
was washed with water (3Â25 mL), brine (25 mL), dried
over sodium sulphate, filtered, and the filtrate evapo-
rated under reduced pressure to give an oil (4.7 g). This
was crystallised from iso-propanol/diethyl ether obtain-
ing( 3), 3 g(40%). 1H NMR (CDCl3) d 8.3 (1H, d,
J=6.6 Hz), 7.26–7.08 (3H, m), 6.5 (1H, NH, t,
J=5.9 Hz), 4.5 (2H, t, J=5.3 Hz), 4.1 (2H, t,
J=5.9 Hz), 3.9 (1H, m), 3.3 (2H, m), 3.2 (1H, m), 2.9
(1H, m), 2.6–2.5 (2H, m), 2.4 (2H, m), 2.1 (1H, m), 1.8–
1.3 (10H, m), 1.26 (3H, d, J=6.6 Hz); 13C NMR
(CDCl3) d 165, 149, 131, 126, 122, 121, 120.7, 108, 89,
70, 67, 58, 55, 52, 44, 39, 36, 33, 30.4, 29.9, 23, 21; m/z
385.2353 (calcd 385.2365); Rf 0.5 (SiO2, 5:1:1 EtOAc/
MeOH/NH4OH); HPLC Retention time 15.8 min, pur-
ity 97.2% (PAR), column: Hichrome RPB 150Â4.6 mm
id, mobile phase: (A) 0.15M NH4OAc adjusted to
pH 4.0 with TFA, (B) methanol gradient: 25 to 60% B
over 40 min, then 5 min at 60% B, detection: UV at
265 nm.
Preparation of methyl 2-(3-chloropropoxy)indole-3-car-
boxylate (11). A suspension of methyl indole-3-carboxy-
late (240.4 g1.37 mol) and DABCO (84.5 g, 0.75 mol) in
dichloromethane (1200 mL) was cooled to 0 ꢀC, treated
in one portion with NCS (201.2 g, 1.51 mol) and the
mixture stirred for 10 min. The resultingsolution was
added to a solution of 3-chloropropan-1-ol (142.5 g,
1.51 mol) in dichloromethane (1200 mL) containing
methane sulphonic acid (10.6 mL) at such a rate as to
maintain the temperature at about 0 ꢀC. The resulting
suspension was stirred for 30 min and then washed with
10% aqueous sodium carbonate solution (3Â1250 mL).
The organic phase was dried over sodium sulphate,
filtered and concentrated on a rotary evaporator. The
ꢀ
resultingoil was triturated with toluene (400 mL) at 0 C
for 1 h and the solid filtered, washed with a small
amount of toluene and dried in vacuo to give 11 as an
off-white solid, 249.5 g(68%). 1H NMR (CDCl3) d: 9.6
(1H, NH, br s), 8.0 (1H, d, J=7.9 Hz), 7.24–7.11 (3H,
m), 4.43 (2H, t, J=5.9 Hz), 3.9 (3H, s), 3.6 (2H, t,
J=5.9 Hz), 2.1 (2H, m); 13C NMR (CDCl3) d 166, 157,
130, 126, 122.2, 121.9, 120.6, 111, 89, 69, 51, 41, 32; m/z
268 (ES+, M+1).
Preparation of methyl 3,4-dihydro-2H-[1,3]oxazino[3,2-
a]indole-10-carboxylate (12). A mixture of 11 (5.0 g
18.7 mmol) and aqueous sodium hydroxide solution
(3.8 mL, 5.4 M, 20.5 mmol) in toluene (50 mL) was
heated at 40 ꢀC for 4 h. The aqueous phase was sepa-
rated and the organic phase was washed with water
(3Â25 mL) while maintainingthe temperature at 60 ꢀC.
The organic solution was evaporated to dryness giving
12, 4.29 g(99%). 1H NMR (DMSO-d6) d 7.9 (1H, d,
J=7.1 Hz), 7.3 (1H, d, J=7.7 Hz), 7.2–7.05 (2H, m), 4.5
(2H, t, J=5.3 Hz), 4.1 (2H, t, J=6.2 Hz), 3.75 (3H, s),
2.2–2.3 (2H, m); 13C NMR (DMSO-d6) d: 164, 153, 131,
125, 121.7, 120.3, 119.2, 109, 85, 66, 50, 39, 20; m/z 232
(ES+, M+1).
Preparation of 6-benzyloxy-3-trichloroacetylindole (15).
To
a
solution of 6-benzyloxyindole 14 (32.6 g,
140 mmol) in THF (320 mL) was added pyridine
(14.8 mL, 180 mmol) and the solution cooled to 2 ꢀC.
Trichloroacetyl chloride (20.1 mL, 180 mmol) dissolved
in THF (320 mL) was added drop-wise over 1 h. The
mixture was allowed to warm to ambient temperature
over 16 h, then evaporated under reduced pressure and
the residue partitioned between ethyl acetate (1400 mL)
and hydrochloric acid (1M, 260 mL). The ethyl acetate
layer was washed with further hydrochloric acid (1M,
2Â260 mL), dried over sodium sulphate, filtered, and
the filtrate evaporated under reduced pressure to give 15
as a solid. This was used in the next stage without
Preparation of 3,4-dihydro-N-[1-[3-(phenylmethoxy)bu-
tyl]-4-piperidinyl]methyl]-2H-[1,3]oxazino[3,2-a]indole-
10-carboxamide (13). Trimethylaluminium (2 M in tolu-

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M. Fedouloff et al. / Bioorg. Med. Chem. 9 (2001) 2119–2128
1
purification. H NMR (CDCl3) d 8.65 (1H, br s), 8.22
(1H, d, J=9.2 Hz), 8.18 (1H, d, J=3.3 Hz), 7.4–7.1
(5H+CHCl3, m), 7.04 (1H, dd, J=11.8 Hz, 1.8 Hz), 6.9
(1H, d, J=2.6 Hz), 5.05 (2H, s); m/z 370 (M+1).
for 16 h. The reaction mixture was filtered, and the fil-
trate evaporated under reduced pressure to give 19,
20.9 g(92%) as a solid, contaminated with excess 3-
chloropropanol (1 equiv). This material was used in the
1
next stage without any further purification. H NMR
Preparation of methyl 6-benzyloxyindole-3-carboxylate
(16). To a solution of 15 (obtained above) in methanol
(2800 mL) was added aqueous potassium hydroxide
(50% w/w, 10 mL). The solution was heated at reflux
for 5 h, then stirred at ambient temperature for 16 h.
The reaction mixture was concentrated until crystal-
lisation occurred. The solid was collected by filtration,
and washed with methanol to give 16, 31.7 g(81%). A
sample recrystallised from methanol melted at 169–
(CDCl3) d 8.0 (1H, d, J=9 Hz), 7.0 (2H, m), 4.6 (2H, t,
J=5.2 Hz), 4.1 (2H, t, J=6.2 Hz), 3.9 (3H, s), 2.3 (5H,
m), plus resonances for 3-chloropropanol at d 3.8 (2H,
t, J=5.9 Hz), 3.7 (2H, t, J=6.4 Hz), 2.0 (2H, t,
J=6.2 Hz); m/z 290 (AP+, M+1).
Preparation of methyl 3,4-dihydro-7-hydroxy-2H-
[1,3]oxazino[3,2-a]indole-10-carboxylate (20). To a sus-
pension of crude 19 (20.9 g) obtained above, in metha-
nol (1000 mL) and water (500 mL) was added saturated
aqueous sodium bicarbonate solution (500 mL). The
mixture was stirred at 40 ꢀC for 2 h, concentrated to
about 300 mL by evaporation under reduced pressure,
diluted with ethyl acetate (400 mL) and cooled to 2 ꢀC.
The solution was neutralised with dilute hydrochloric
acid and the organic layer collected. The aqueous layer
was extracted with further ethyl acetate (2Â400 mL) and
the combined organic layers washed with water
(3Â250 mL), brine (250 mL), dried over sodium sul-
phate. The mixture was filtered and evaporated under
reduced pressure to give a solid which was recrystallised
from iso-propanol, obtaining 20, 11.3 g(58%). 1H
NMR (DMSO-d6) d 9.07 (1H, OH, s), 7.6 (1H, d,
J=8.3 Hz), 6.6 (2H, m), 4.45 (2H, t, J=5.3 Hz), 3.91
(2H, t, J=6.6 Hz), 3.7 (3H, s), 2.2 (2H, m); 13C NMR
(CDCl3) d: 169, 157.9, 157.6, 137, 125, 122, 116, 101, 89,
71, 55, 44, 26; m/z 248 (M+1).
170 ꢀC (dec.). H NMR (CDCl3) d 8.4 (1H, br s), 8.0
1
(1H, d, J=9 Hz), 7.7 (1H, d, J=3 Hz), 7.4–7.3 (5H, m),
5.0 (2H, s), 3.8 (3H, s); m/z 282 (M+1); Rf 0.42 (SiO2,
CHCl3).
Preparation of methyl 6-hydroxyindole-3-carboxylate
(17). A solution of 16 (31.7 g, 110 mmol) in THF
(250 mL) and methanol (200 mL) was hydrogenated at
atmospheric pressure over 10% Pd/C (15.9 g) for 5 days.
The reaction mixture was filtered through Celite, and
the filtrate was evaporated under reduced pressure to an
oil which was crystallised from chloroform, obtaining
17, 17.5 g(81%). 1H NMR (CD3OD) d 7.75 (0.5H, s),
7.70 (1.5H, m), 6.7 (1H, d, J=2 Hz), 6.6 (1H, dd,
J=2.2 Hz, 8.6 Hz), 3.8 (3H, s); m/z 192 (M+1).
Preparation of methyl 6-acetoxyindole-3-carboxylate
(18). To a solution of 17 (15.1 g, 79 mmol) in THF
(200 mL) at 3 ꢀC was added pyridine (5.5 mL, 68 mmol)
followed by acetic anhydride (22 mL, 230 mmol). The
solution was stood for 16 h at 4 ꢀC, then evaporated
under reduced pressure and the residue partitioned
between ethyl acetate (200 mL) and cold dilute hydro-
chloric acid (1 M, 100 mL). The organic layer was
washed with further cold dilute hydrochloric acid (1 M,
100 mL), water (3Â35 mL), brine (2Â50 mL), dried over
sodium sulphate, filtered and the filtrate evaporated
under reduced pressure to give 18, 8.4 g(99%) as a
solid. 1H NMR (CDCl3) d 8.82 (1H, NH, br s), 8.1 (1H,
d, J=8.7 Hz), 7.7 (1H, d, J=3.0 Hz), 7.1 (1H, d,
J=2.0 Hz), 6.95 (1H, dd, J=2.0 Hz, 8.7 Hz), 3.9 (3H, s),
2.35 (3H, s); 13C NMR (CDCl3) d 170, 165.3, 147, 136,
132, 124, 122, 116, 109, 105, 51, 21; m/z 234 (M+1).
Preparation of methyl 3,4-dihydro-7-benzyloxy-2H-
[1,3]oxazino[3,2-a]indole-10-carboxylate (21). A mixture
of 20 (11.2 g, 45.3 mmol), anhydrous potassium carbon-
ate (6.3 g, 46 mmol), benzyl chloride (6 mL, 52 mmol)
and DMF (125 mL) was heated at 100 ꢀC with stirring
under nitrogen for 6 h. The mixture was allowed to cool,
evaporated to dryness under reduced pressure and the
residue partitioned between ethyl acetate (600 mL) and
water (150 mL). The organic layer was washed with
further water (2Â100 mL) and brine (100 mL), then
dried over sodium sulphate. The mixture was filtered
and the filtrate evaporated under reduced pressure to
give a solid (16.5 g) which was recrystallised from
methanol (250 mL) obtaining 21, 11.9 g(70%) which
melted at 101–102 ꢀC. H NMR (CDCl3) d 7.9 (1H, d,
1
Preparation of methyl 3,4-dihydro-7-acetoxy-2H-[1,3]ox-
azino[3,2-a]indole-10-carboxylate (19). To a solution of
18 (18.4 g, 80 mmol) in chloroform (250 mL) at 2 ꢀC was
added DABCO (4.9 g, 43 mmol) followed by N-chloro-
succinimide (11.6 g, 87 mmol). The mixture was stirred
for 30 min, then 3-chloropropanol (14.9 g, 160 mmol)
was added, followed by methanesulphonic acid (1.1 g,
11 mmol). The mixture was stirred for 75 min, then
sodium carbonate solution (10% aqueous, 200 mL) was
added. The phases were separated and organic layer was
washed with water (2Â100 mL), brine (100 mL), dried
over sodium sulphate, filtered, and the filtrate evapo-
rated under reduced pressure. The residue was re-dis-
solved in acetone (570 mL), anhydrous potassium
carbonate (69 g, 0.5 mol) added and the mixture stirred
J=8.6 Hz), 7.5–7.3 (5H, m), 6.93 (1H, dd, J=2.3 Hz,
8.6 Hz), 6.7 (1H d, J=2.2 Hz), 5.1 (2H, s), 4.5 (2H, t,
J=5.2 Hz), 4.0 (2H, t, J=6.3 Hz), 3.9 (3H, s), 2.3 (2H,
m); 13C NMR (CDCl3) d: 165, 155, 153, 137, 132, 128.5,
128.3, 128, 127.5, 121, 119, 111, 95, 86, 71, 66, 51, 39,
30, 21; m/z 338 (M+1).
Preparation of N-[(1-butyl-4-piperidinyl)methyl]-3,4-di-
hydro-7-benzyloxy-2H-[1,3]oxazino[3,2-a]indole-10-car-
boxamide (23). Trimethylaluminium (19.3 mL, 2 M in
toluene) was diluted with toluene (18 mL) and the solu-
tion cooled to 2 ꢀC under nitrogen. Amine 22 (6.03 g,
35.4 mmol) dissolved in toluene (55 mL) was added drop
wise, allowingthe temperature to rise to 10 ꢀC. Com-
pound 21 (11.9 g, 35.3 mmol) was added as a solid and

M. Fedouloff et al. / Bioorg. Med. Chem. 9 (2001) 2119–2128
2127
the mixture heated at reflux for 5 h. The solution was
allowed to cool to 70 ꢀC, then sodium hydroxide solution
(10% w/w, 85 mL) was added cautiously over 10 min.
The phases were separated and the toluene layer was
washed with further sodium hydroxide solution (10%
w/w, 85 mL), water (2Â120 mL) and brine (120 mL).
The solution was dried over sodium sulphate, filtered,
and the filtrate evaporated under reduced pressure to
give a solid (16.9 g) which was recrystallised from ethyl
acetate, obtaining 23, 9.1 g(54%). 1H NMR (CDCl3) d
8.2 (1H, d, J=8.6 Hz), 7.5–7.3 (5H, m), 6.95 (1H, dd,
J=2.2 Hz, 8.7 Hz), 6.7 (1H, d, J=2.2 Hz), 6.51 (1H,
NH, t, J=5.9 Hz), 5.1 (2H, s), 4.5 (2H, t, J=5.1 Hz), 4.0
(2H, t, J=6.2 Hz), 3.3 (2H, t, J=6.1 Hz), 2.9 (2H, m),
2.3 (4H, m), 2.0–1.2 (11H, m), 0.9 (3H, t, J=7.1 Hz);
13C NMR (CDCl3) d 165, 155, 149, 143, 137, 132, 128,
127.8, 127.5, 122, 120, 111, 95, 94.5, 89, 71, 67, 59, 57,
54, 44, 39, 36, 30, 29, 21, 20.9, 14, 12; m/z 476 (M+1);
Rf 0.6 (SiO2, 5:1:1 EtOAc/MeOH/NH4OH).
Preparation of methyl 3,4-dihydro-5-hydroxy-4-oxo-2H-
[1,3]oxazino[3,2-a]indole-10-carboxylate (26). To a mix-
ture of 24 (4.8 g, 190 mmol), powdered 4 A molecular
sieves (9.6 g), N-methyl morpholine-N-oxide (3.39 g,
29 mmol) in dichloromethane/acetonitrile (10:1, 44 mL)
was added tetrapropylammonium perruthenate (TPAP)
(0.34 g, 0.97 mmol). The mixture was stirred for 18 h,
filtered through Celite, and the filtrate evaporated. The
residue was chromatographed (SiO2, 0–10% ethyl ace-
tate/dichloromethane) obtaining( 26), 1.23 g(28%) and
27, 0.52 g(11%). 1H NMR 26 (CDCl3) d 7.8 (1H, d,
J=7.2 Hz), 7.3–7.0 (3H, m), 5.85 (1H, br s), 4.73–4.4
(2H, m), 3.7 (3H, s), 2.1–2.4 (2H, m); 13C NMR (26)
(CDCl3) d 165, 153, 130, 125, 123, 121, 120.6, 108, 87,
1
71, 62, 51, 29; m/z 248 (AP+, M+1); H NMR (27)
(CDCl3) d: 8.2 (1H, m), 7.95 (1H, m), 7.4–7.16 (2H, m),
4.66 (2H, t, J=6.7 Hz), 3.86 (3H, s), 3.0 (2H, t,
J=6.7 Hz); m/z 246 (AP+, M+1).
Preparation of (Æ)-methyl 3,4-dihydro-4-[dimethyl-(1,1-
dimethylethyl)silyl]oxy]-2H-[1,3]oxazino[3,2-a]indole-10-
carboxylate (28). To a solution of 26 (2 g, 8.1 mmol)
and 2,6-lutidine (4.2 mL, 36 mmol) in dichloromethane
(64 mL) at À70 ꢀC was added TBDMS triflate (4 mL,
174 mmol) drop-wise over 3 min. The solution was stir-
red for 1 h, then allowed to warm to room temperature.
The solution was re-cooled to À70 ꢀC, then methanol
(9 mL) was added drop-wise over 5 min. The solution
was diluted with dichloromethane (200 mL), washed
with water, brine and dried (Na2SO4) to give an oil.
This was chromatographed (SiO2, 10% ethyl acetate/
dichloromethane to give 28, 2 g(69%). 1H NMR
(CDCl3) d: 8.1 (1H, m), 7.31–7.2 (3H+CHCl3, m), 6.0
(1H, m), 4.86–4.7 (2H, m), 3.95 (3H, s), 2.44–2.37 (1H,
m), 2.22–2.14 (1H, m), 0.91 (9H, s), 0.29 (3H, s), 0.22
(3H, s); m/z 362 (AP+, M+1).
Preparation of N-[(1-butyl-4-piperidinyl)methyl]-3,4-di-
hydro-7-hydroxy-2H-[1,3]oxazino[3,2-a] indole-10-carbox-
amide (4). A solution of 23 (9.1 g, 19 mmol) in methanol
(190 mL) was hydrogenated over 10% Pd/C (0.99 g) at
atmospheric pressure for 28 h. The reaction mixture was
filtered through Celite and the filtrate evaporated under
reduced pressure to give a solid which was recrystallised
from ethyl acetate obtaining 4, 6.2 g(84%) which mel-
ted at 125–126 C. H NMR (CD3OD) d 7.85 (1H, d,
8.4 Hz), 6.66 (1H, dd, J=2.3 Hz, 8.4 Hz), 6.58 (1H, d,
J=2.2 Hz), 4.48 (2H, t, J=5.0 Hz), 3.92 (2H, t,
J=6.2 Hz), 3.25 (2H, d, J=6.7 Hz), 2.95 (2H, m), 2.29
(4H, m), 1.95 (2H, m), 1.73 (2H, m), 1.47 (1H, m), 1.34
(2H, m), 1.30 (4H, m), 0.93 (3H, t, J=7.3 Hz); 13C
NMR (CD3OD) d 168, 154, 151, 134, 122, 120, 112,
96, 89, 68, 60, 55, 45, 40, 38, 31, 30, 22.3, 22, 14;
nOe experiments confirmed 7-OH substitution; m/z
385.2353 (calcd 385.2365); HPLC retention time 12.1
min, purity 99.1% (PAR), column: Hichrome RPB
150Â4.6 id mm, Mobile Phase: (A) 0.15 M NH4OAc
adjusted to pH 4.0 with TFA, (B) Methanol, Gradient:
25 to 60% B over 40 min, then 5 min at 60% B, detec-
tion: UV at 265 nm.
ꢀ
1
Preparation of (Æ)-N-[(1-butyl-4-piperidinyl)methyl]-
3,4-dihydro-4-[dimethyl-(1,1-dimethylethyl)silyl]oxy]-2H-
[1,3]oxazino[3,2-a]indole-10-carboxamide (29). To
a
solution of 1-n-butyl-4-piperidinylmethylamine (22)
(0.95 g, 47 mmol) in toluene (2.5 mL) was added tri-
methylaluminium (2.3 mL, 2 M in toluene, 46 mmol)
followed by 28 (1.62 g, 45 mmol) in toluene (5 mL). The
solution was heated under reflux for 4 h, then allowed to
cool to room temperature and stirred with 10% aqueous
NaOH solution (2 mL) for 30 min. The organic layer
was washed with 10% aqueous NaOH solution, water,
brine, dried over sodium sulphate, filtered and the fil-
trate evaporated to leave an oil. This was filtered
through silica, washing with ethyl acetate, then metha-
nol obtaining 29, 2.0 g(89%). 1H NMR (CD3OD) d: 8.1
(1H, m), 7.3 (1H, m), 7.1 (2H, m), 6.17 (1H, m), 4.7 (2H,
m), 3.0 (2H, m), 2.6–0.9 (9H, m), 0.3 (3H, s), 0.2 (3H, s);
m/z 500 (AP+, M+1).
Preparation of methyl 2-(3-hydroxypropoxy)-1H-indole-
3-carboxylate (24). To a suspension of methyl indole-3-
carboxylate 10 (25.3 g, 144 mmol) and DABCO (8.7 g,
78 mmol) in chloroform (250 mL) at 5 ꢀC was added N-
chlorosuccinimide (21 g, 157 mmol). After 30 min, 1,3-
propanediol (110 g, 1.45 mol) was added, followed by
methanesulphonic acid (3 g). The solution was allowed
to warm to room temperature and stirred at room tem-
perature for 1 h, then washed with 10% aqueous
Na2CO3 solution, water, brine and dried over sodium
sulphate. The mixture was filtered and evaporated
under reduced pressure to leave an oil. Chromato-
graphy (5–50% ethyl acetate/dichloromethane) gave 24,
16.2 g(45%), and 25, 3.7 g(12%). 1H NMR (24)
(CD3OD) d 7.85 (1H, m), 7.25 (1H, m), 7.1 (2H, m),
4.45 (2H, t, J=5.9 Hz), 3.85 (5H, m), 2.1 (2H, m); m/z
Preparation of (Æ)-N-[(1-butyl-4-piperidinyl)methyl]-3,4
-dihydro-4-hydroxy-2H-[1,3]oxazino[3,2-a]indole-10-car-
boxamide (5). A solution of 29 (0.15 g, 0.3 mmol) in
water (5 mL) and glacial acetic acid (10 mL) was heated
at 75 ꢀC for 6.5 h. The solution was evaporated under
reduced pressure and the residue partitioned between
ethyl acetate (25 mL) and saturated aqueous NaHCO3
1
250 (EI, M+1); H NMR (25) (DMSO-d6) d: 7.8 (2H,
m), 7.26 (2H, m), 7.06 (4H, m), 4.55 (4H, t, J=6.0 Hz),
3.68 (6H, s), 2.3 (2H, m); m/z 421 (APÀ, MÀ1).

2128
M. Fedouloff et al. / Bioorg. Med. Chem. 9 (2001) 2119–2128
solution (3 mL). The ethyl acetate layer was washed
with further saturated aqueous NaHCO3 solution, then
with brine and dried over sodium sulphate. Evaporation
under reduced pressure gave 5, 0.096 g(83%) as a foam.
1H NMR (CD3OD) d 8.16 (1H, m), 7.5 (1H, m), 7.2
(2H, m), 6.05 (1H, m), 4.8 (2H, m), 3.4 (2H, m), 3.1 (2H,
m), 2.6–1.35 (15H, m), 1.03 (3H, t, J=7.2 Hz); 13C
NMR (CD3OD) d 168, 151, 132, 127, 123, 122, 121, 110,
90, 72, 65, 62, 60, 45, 38, 31.3, 31, 30, 22, 21, 15, 14.5; m/
z 385.2353 (calcd 385. 2365); HPLC retention time 14.5
min, purity 97.4% (PAR), column: Hichrome RPB
150Â4.6 mm id, mobile phase: (A) 0.15 M NH4OAc
adjusted to pH 4.0 with TFA, (B) methanol, gradient:
25 to 60% B over 40 min, then 5 min at 60% B, detec-
tion: UV at 265 nm.
29697 (SmithKline Beecham plc). (c) WO 00/17207 (Smith-
Kline Beecham plc).
3. Dimitriadis, E.; Massy-Westropp, R. A. Aust. J. Chem.
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4. Ohkubo, M.; Nishimura, T.; Jona, H.; Honma, T.; Mor-
ishima, H. Tetrahedron 1996, 52, 8099.
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A.D J. Chem. Soc., Chem. Commun., 1987, 1625 (b) Griffith,
W. P.; Ley, S. V. Aldrichimica Acta 1990, 23, 13.
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References and Notes
10. Kennett, G. A.; Wood, M. D.; Bright, F.; Trail, B.; Riley,
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