Novel synthesis of physovenine and physostigmine analogs

Article abstract of DOI:10.1016/j.tetlet.2016.06.005

This Letter describes a versatile synthetic approach to prepare physovenine and physostigmine analogs. A series of analogs were synthesized and evaluated for cholinesterase inhibition activities, including human acetylcholinesterase (AChE) and butyrylchol

Full text of DOI:10.1016/j.tetlet.2016.06.005

Tetrahedron Letters 57 (2016) 3046–3049
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Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Novel synthesis of physovenine and physostigmine analogs
C. H. Wang a, S. Alluri a, G. Nikogosyan a, C. DeCarlo a, C. Monteiro a, G. Mabagos a, H. H. Feng a,
A. R. White a, M. Bartolini b, V. Andrisano c, L. K. Zhang d, A. K. Ganguly a,
⇑
a Department of Biomedical Engineering, Chemistry and Biological Sciences Stevens Institute of Technology, Hoboken, NJ 07030, USA
b Department of Pharmacy and Biotechnology, University of Bologna, via Belmeloro 6, 40126 Bologna, Italy
c Department for Life Quality Studies, University of Bologna, Corso d0Augusto, 237, 47921 Rimini, Italy
d Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, NJ 07033-1300, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
This Letter describes a versatile synthetic approach to prepare physovenine and physostigmine analogs. A
series of analogs were synthesized and evaluated for cholinesterase inhibition activities, including human
acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) from human serum.
Ó 2016 Elsevier Ltd. All rights reserved.
Received 28 April 2016
Revised 29 May 2016
Accepted 1 June 2016
Available online 3 June 2016
Keywords:
Radical reactions
AChE inhibitors
Physovenine and physostigmine analogs
Introduction
Present study
(ꢀ)-Physovenine and (ꢀ)-physostigmine (Fig. 1) are known to
be acetylcholinesterase (AChE) inhibitors.1a–c These alkaloids
have been used in the past for the treatment of myasthenia gravis
and glaucoma and more recently AChE inhibitors such as
Donepezil2 and Rivastigmine3 have found use for the treatment
of Alzheimer’s patients.
The main challenge for the synthesis of physovenine and
physostigmine analogs is the construction of quaternary carbon
center. There are reported syntheses in the literature4a–e toward
achieving this goal. Our approach for the synthesis of physovenine
and physostigmine analogs is different and novel. The present syn-
thesis allows us to make structurally diverse compounds for estab-
lishing structure activity relationship. The key step in our synthesis
is the formation of the spiro-oxindiole ring using radical cycliza-
tion process followed by further elaboration yielding the crucial
alcohol intermediate (A).
Substituted
physostigmine and physovenine analogs were prepared as follows.
Treatment6 of
-butyrolactone 1a with sodium methoxide and aro-
matic aldehydes gave substituted butyrolactones 2a–d (Scheme 1).
However, the reaction of 1a with acetaldehyde yielded an unex-
pected product 2e involving the consumption of two equivalents
of acetaldehyde. For the synthesis7 of 2f and 2g we treated the cor-
responding ketone and aldehyde with NaH and diethyl(2-oxote-
trahydrofuran-3-yl)phosphonate 1c, which in turn was derived
c-butyrolactones required for the synthesis of
c
from
a-bromo-c-butyrolactone 1b. Compound 1a when treated
with benzyl bromide yielded 2i. It should be noted that the reac-
tion of 1a with 2-bromopropane yielded 2h, as a result of self-con-
densation of c-butyrolactone 1a.
Treatment of compounds 2a–h with 2-bromo-4-methoxyani-
line and trimethylaluminum in toluene gave amides 3a–3h
(Scheme 2), respectively. These derivatives were acetylated using
acetyl chloride and pyridine in DCM to protect the hydroxyl group
and then N-alkylated using Cs2CO3 and alkyl halides in DMF yield-
ing the desired precursors 5a–5h for reductive radical cyclization.
Following the above procedure radical precursors 8a–c used for
oxidative cyclization were prepared from 2i–j. Compounds 5a–5g
upon treatment with AIBN and TBTH in toluene solution yielded
both the exo and endo cyclization derived products through the
intermediacy of 9 yielding compounds 10a–g and 11a–e, respec-
tively (Scheme 3). Surprisingly, two atropisomers (10h and 10i)
were isolated from the radical reaction of 5h. High resolution mass
In a previous publication5 we have reported the synthesis of a
novel class of progesterone receptor antagonists using the interme-
diate (A) and in this publication we have used the same intermedi-
ate (A) to synthesize novel physostigmine and physovenine
analogs represented by structure (D, wherein X = N and X = O
respectively). These novel analogs were synthesized through the
intermediacy of aryl radicals (B) and (C).
⇑
Corresponding author. Tel.: +1 617 4843806.
E-mail address: akganguly1@aol.com (A.K. Ganguly).
http://dx.doi.org/10.1016/j.tetlet.2016.06.005
0040-4039/Ó 2016 Elsevier Ltd. All rights reserved.

C. H. Wang et al. / Tetrahedron Letters 57 (2016) 3046–3049
3047
O
H
H
O
N
N
O
O
H
O
N
N
N
N
H
O
N
H
O
O
N
O
O
Donepezil
(-)Physostigmine
Rivastigmine
R2
(-)Physovenine
R2
R2
N
R2
N
H
N
N
O
O
R
O
R
X
R3
R1
R1
O
R3
H3CO
R4
R4
H3CO
R1
(D)
R1
OH
(B)
(C)
R
(A)
Figure 1. Examples of AChE inhibitors.
and H11 (Fig. 2). Treatment of 8a–c with TBTH and AIBN produced
the desired spirooxindoles (10a, 10j–k), involving the rearrange-
ment of 12 to the more stable radical 13.
O
(1). Diisopropylamine,
O
O
n-BuLi, THF, -10oC
R1
R1
R
O
O
O
O
(2). 2-Bromopropane
THF, -78oC
Hydrolysis of 10a–b with 2 N NaOH in methanol yielded the
corresponding alcohols 14a and 14b, which were then converted
to aldehydes 15a and 15b, respectively using the Dess–Martin oxi-
dation (Scheme 4). Condensation of methyl amine with 15a fol-
lowed by reduction with LiAlH4 afforded 16a. O-demethylation of
16a using BBr3 yielded the phenol 17a. Similarly, 17b was obtained
from 14b. Treatment of the phenols 17a–b with NaH and substi-
tuted isocyanates yielded the desired physostigmine analogs
18a–c.
Treatment of compounds 10a–k and 14a–d with LiAlH4 in THF
under reflux yielded the compounds with physovenine core struc-
tures 19a–j. O-demethylation of compounds 19a–j followed by
reaction with substituted isocyanates furnished the desired physo-
venine analogs 21a–g. The biological activities of the physostigmi-
nes (18a–c) and physovenines (21a–g) analogs are summarized in
Table 1.
NaOMe,
R
MeOH, 0oC
O
2h (25.0%)
O
1a
2a: R=C6H5, R1=H (72.6%)
2b: R=4-Cl-C6H4, R1=H (89.6%)
(1). Diisopropylamine,
n-BuLi, THF, -10oC
O
2c: R=4-CH3-C6H4, R1=H (88.7%)
2d: R=4-CH3O-C6H4, R1=H (40.0%)
2e: R=CH=CHCH3, R1=H (9.4%)
(2). benzyl bromide
THF, -78oC
2i (13.0%)
O
O
R1
NaH
THF
R
O
O
O
P
O
R1
R
O
P(OEt)3
O
O
O
Br
reflux
140oC
1b
1c
2f: R=CH2CH(CH3)2, R1=H (27.0%)
2g: R=R1=CH3 (70.5%)
Scheme 1. Synthesis of substituted c-butyrolactones.
It is evident from the IC50 values reported in Table 1 that in the
physostigmine series the benzyl substituted analogs 18a, 18b, and
18c were inactive against hAChE but active against hBuChE. In the
physovenine series benzyl substituted compounds 21a, 21b, and
21c were selective hBuChE inhibitors with inhibitory potencies in
the submicromolar range. The most active against hBuChE being
21a with an IC50 value of 70 nM and selectivity around 58 folds.
In the alkyl substituted derivatives, 21d, 21e, 21f, and 21g were
slightly selective toward hAChE. Comparing compounds 21f and
spectroscopy established identical molecular compositions of both
the above compounds. NMR spectra of 10h and 10i indicated that
H10 in these compounds appear at d 4.14 and d 4.03 respectively.
The quaternary carbon C9 in 10h appeared at d 54.55, whereas C9
in 10i appeared at d 54.22. Both spiro-oxindoles show identical
correlations in COSY and HMBC. For example in HMBC, C9 shows
correlations with H7, H15, and H11, whereas C1 with H10, H14, and
H17. In addition, 10h showed long range NOE between H7 and
H10, whereas 10i showed the correlations of H7 with both H10
Br R2
Br
H
N
Br
H
N
N
O
R
Br
Cs2CO3
R2X
DMF
2a-2h
O
R
O
R
NH2
Me3Al
acetyl chloride
pyridine, DCM
R1
R1
R1
O
CH3
toluene
O
CH3
O
CH3
O
CH3
O
O
HO
H3C
O
H3C
O
3a: R=C6H5, R1=H (85.7%)
5a: R=C6H5, R1=H, R2=CH3 (97.2%)
4a: R=C6H5, R1=H (84.3%)
3b: R=4-Cl-C6H4, R1=H (88.6%)
3c: R=4-CH3-C6H4, R1=H (97.3%)
3d: R=4-CH3O-C6H4, R1=H (79.8%)
3e: R=CH=CHCH3, R1=H (41.4%)
3f: R=CH2CH(CH3)2, R1=H (57.1%)
3g: R=R1=CH3 (71.2%)
5b: R=4-Cl-C6H4, R1=H, R2=CH3 (97.7%)
5c: R=4-CH3-C6H4, R1=H, R2=CH3 (95.7%)
5d: R=4-CH3O-C6H4, R1=H, R2=CH3 (97.0%)
5e: R=CH=CHCH3, R1=H, R2=CH3 (90.5%)
5f: R=CH2CH(CH3)2, R1=H, R2=CH3 (95.4%)
5g: R=R1=CH3, R2=CH3 (97.4%)
4b: R=4-Cl-C6H4, R1=H (90.8%)
4c: R=4-CH3-C6H4, R1=H (98.7%)
4d: R=4-CH3O-C6H4, R1=H (90.9%)
4e: R=CH=CHCH3, R1=H (85.0%)
4f: R=CH2CH(CH3)2, R1=H (82.5%)
4g: R=R1=CH3 (88.5%)
3h: R, R1= 2-tetrahydrofuran (85.2%)
5h: R, R1= 2-tetrahydrofuran, R2=CH3 (94.2%)
4h: R, R1= 2-tetrahydrofuran (91.4%)
Br R2
Br
H
N
Br
H
N
Br
N
O
R
O
R
O
R
NH2
Cs2CO3
R2X
O
Me3Al
toluene
acetyl chloride
pyridine, DCM
R1
R1
R1
O
R1
O
CH3
O
CH3
O
CH3
O
CH3
DMF
R
O
O
HO
H3C
O
H3C
O
2i: R=C6H5, R1=H
2j: R=R1=H
8a: R=C6H5, R1=H, R2=CH3 (97.6%)
8b: R=R1=H, R2=CH2CH3 (95.5%)
8c: R=R1=H, R2=CH3 (92.5%)
7a: R=C6H5, R1=H (97.5%)
7b: R=R1=H (86.2%)
6a: R=C6H5, R1=H (70.2%)
6b: R=R1=H (75.4%)
Scheme 2. Preparation of radical precursors.

3048
C. H. Wang et al. / Tetrahedron Letters 57 (2016) 3046–3049
Br R2
N
R2
N
R2
N
R2
N
O
R
O
R
O
O
AIBN
CH3
O
O
CH3
R1
R1
TBTH
H3C
H3C
O
O
CH3
O
CH3
O
O
O
R1
R
R1
toluene
O
O
9
R
11a: R=C6H5, R1=H, R2=CH3 (11.2%)
10a: R=C6H5, R1=H, R2=CH3 (60.9%)
H3C
O
H3C
O
11b: R=4-Cl-C6H4, R1=H, R2=CH3 (12.9%)
11c: R=4-CH3-C6H4, R1=H, R2=CH3 (28.4%)
11d: R=4-CH3O-C6H4, R1=H, R2=CH3 (18.9%)
11e R=CH2CH(CH3)2, R1=H, R2=CH3 (24.8%)
10b: R=4-Cl-C6H4, R1=H, R2=CH3 (68.8%)
10c: R=4-CH3-C6H4, R1=H, R2=CH3 (60.2%)
10d: R=4-CH3O-C6H4, R1=H, R2=CH3 (61.9%)
10e1: R=CHCH2CH3, R1=H, R2=CH3 (24.7%)
10e2 :R=CH=CHCH3, R1=H, R2=CH3 (18.3%)
10f: R=CH2CH(CH3)2, R1=H, R2=CH3 (30.1%)
10g: R=R1=CH3, R2=CH3 (82.2%)
5a-h
10h: R, R1= 2-tetrahydrofuran, R2=CH3 (33.9%)
10i: R, R1= 2-tetrahydrofuran, R2=CH3 (31.1%)
Br R2
N
R2
N
R2
N
O
O
O
R
AIBN
R2
N
1, 5 transfer
TBTH
O
O
CH3
O
CH3
O
CH3
O
CH3
toluene
H3C
R
R
O
O
O
O
O
R1
R
H3C
O
H3C
O
H3C
O
10a: R=C6H5, R1=H, R2=CH3 (36.4%)
10j: R=R1=H, R2=CH2CH3 (43.5%)
10k: R=R1=H, R2=CH3 (46.2%)
8a: R=C6H5, R1=H, R2=CH3
8b: R=R1=H, R2=CH2CH3
8c: R=R1=H, R2=CH3
12
13
R2
N
O
R2
N
O
CH3
H3C
O
O
NiCl2 6H2O, NaBH4
CH3OH, THF (1:1)
O
H3C
OH
O
R1
R
R1
10e=CH2CH2CH3, R1=H, R2=CH3 (93.0%)
Scheme 3. Radical cyclizations.
R
10e1: R=CH=CHCH3, R1=H, R2=CH3
10e2: R=CHCH2CH3, R1=H, R2=CH3
active against hBuChE. In general physovenine and physostigmine
analogs, 18a–c and 21a–e with alkyl substituents were more active
against hAChE than the arylalkyl substituents at the quaternary
carbon center 3a. Indeed, it is evident that the size of the sub-
stituent at 3a has a strong effect on the inhibitory activity against
hAChE. For physovenine analogs, the inhibitory activity toward
hAChE decreased by about 2.8 times upon substitution of a methyl
group with a butyl group (compare 21d with 21e), and 13 times
upon introduction of a benzyl group (compare 21a with 21e), while
the introduction of a p-chloro-benzyl substituent led to an inactive
derivative (21b). This trend can be likely ascribed to the narrow
AChE’s gorge which might not allow to accommodate physovenine
derivatives with bulky substituents at the quaternary carbon cen-
ter 3a. Conversely, due to the larger dimensions of the BuChE cat-
alytic gorge (approximately 340 Å3 larger than the corresponding
portion of the hAChE gorge) (Zha et al.), BuChE could easily accom-
modate analogs 21a–c. The evident effect of the size of the sub-
stituent at 3a on the inhibitory potency toward hAChE also
determined a change of the selectivity profile. In fact while physo-
venine analogs 21e–g (R = R1 = H) and 21d (R = CH2CH2CH3; R1 = H)
turned out to be selective hAChE inhibitors, analogs 21a–c with
bulky aryl alkyl substituent at 3a were highly selective BuChE
inhibitors.
10h
10i
In conclusion, the most potent among the alkyl substituted
compounds was 21g with an IC50 value of 53 nM against hAChE
and selectivity of 18 fold in activities between the two enzymes.
We believe the present work helps to establish the trend in SAR
among physostigmine and physovenine analogs for activity against
hAChE and hBuChE.
Figure 2. NOE correlations of 10h and 10i.
21g (at their maximum inhibition, according to the inhibition
kinetics) it is apparent that 21g was five-fold more active than
the aromatic carbamate 21f against hAChE and two-fold more

C. H. Wang et al. / Tetrahedron Letters 57 (2016) 3046–3049
3049
R2
N
R2
N
R2
N
(1) CH3NH2 HCl
TEA, MgSO4, THF
H
2N NaOH
DMP
DCM
O
CH3
10a-k
O
H3C
N
OH
H3C
H3C
O
O
O
DCM:CH3OH
9:1
O
(2) LiAlH4
reflux
R1
R1
R1
R
R
R
14a: R=C6H5, R1=H, R2=CH3 (90.4%)
16a: R=C6H5, R1=H, R2=CH3 (50.2%)
16b: R=4-Cl-C6H4, R1=H, R2=CH3
(39.7%)
15a: R=C6H5, R1=H, R2=CH3 (94.4%)
15b: R=4-Cl-C6H4, R1=H, R2=CH3 (92.5%)
14b: R=4-Cl-C6H4, R1=H, R2=CH3 (85.2%)
14c: R=4-CH3-C6H4, R1=H, R2=CH3 (79.5%)
14d: R=4-CH3O-C6H4, R1=H, R2=CH3 (90.6%)
14e: R=R1=H, R2=CH3 (70.7%)
LiAlH4
reflux
THF
LiAlH4
THF
DCM
BBr3
14f: R=R1=H, R2=CH2CH3 (89.5%)
R2
R2
N
R2
N
19a: R=C6H5, R1=H, R2=CH3 (70.4%)
19b: R=4-Cl-C6H4, R1=H, R2=CH3 (83.9%)
19c: R=4-CH3-C6H4, R1=H, R2=CH3 (86.2%)
19d: R=4-CH3O-C6H4, R1=H, R2=CH3 (94.5%)
19e: R=CH2CH2CH3, R1=H, R2=CH3 (86.7%)
19f: R=R1=CH3, R2=CH3 (92.6%)
H
O
N
H
N
NaH
N C O
H
CH3
O
CH3
H3C
R3
N
O
HO
R3
N
H
O
R1
R1
THF
R1
R
R
R
19g: R, R1= 2-tetrahydrofuran, R2=CH3 (95.1%)
19h: R, R1= 2-tetrahydrofuran, R2=CH3 (96.9%)
19i: R=R1=H, R2=CH2CH3 (59.2%)
17a: R=C6H5, R1=H, R2=CH3 (80.6%)
17b: R=4-Cl-C6H4, R1=H, R2=CH3 (60.4%)
18a-c
20a: R=C6H5, R1=H, R2=CH3
(
78.9%)
BBr3
DCM
20b: R=4-Cl-C6H4, R1=H, R2=CH3 (90.8%)
20c: R=4-CH3-C6H4, R1=H, R2=CH3 (64.5%)
20d: R=CH2CH2CH3, R1=H, R2=CH3 (94.6%)
20e: R=R1=CH3, R2=CH3 (66.6%)
19j: R=R1=H, R2=CH3 (66.0%)
R2
NaH
N C O
R2
H
N
20f: R=R1=H, R2=CH3 (86.0%)
H
R3
N
O
20g: R=R1=H, R2=CH2CH3 (93.3%)
O
O
HO
R3
THF
N
H
O
R1
R1
R
21a-g
R
Scheme 4. Synthesis of physovenine and physostigmine analogs.
Table 1
hAChE and hBuChE of the analogs
R2
N
H
O
3a
a
a
R3
Yield (%)
X
R
R1
R2
R3
IC50 hAChEb
(lM) SEM
IC50 hBuChEb
(l
M) SEM
X
N
H
O
R1
R
18a
18b
18c
21a
21b
21c
21d
21e
21f
75.0
63.7
41.6
62.9
43.1
48.8
15.0
59.1
48.7
54.6
N-CH3
N-CH3
N-CH3
O
O
O
O
O
O
O
C6H5
C6H5
H
H
H
H
H
H
H
H
H
H
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
C2H5
C2H5
C(CH3)3
Not active
>30c
4.35 0.24
7.17 0.54
13.7 1.3
0.070 0.007
0.173 0.019
0.231 0.018
1.20 0.20
0.985 0.125
1.87 0.39
3-Cl-4-CH3-C6H3
3-Cl-4-CH3-C6H3
3-Cl-4-CH3-C6H3
3-Cl-4-CH3-C6H3
3-Cl-4-CH3-C6H3
3-Cl-4-CH3-C6H3
3-Cl-4-CH3-C6H3
3-Cl-4-CH3-C6H3
Cyclohexane
4-Cl-C6H4
C6H5
4-Cl-C6H4
4-CH3-C6H4
CH2CH2CH3
H
Not active
4.04 0.45
Not active
9.55 0.51
0.638 0.049
0.226 0.022
0.361 0.025
0.182 0.010
H
H
21g
2.74 0.11
0.053 0.005d
0.959 0.052e
a
IC50 values represent the concentration of inhibitor required to decrease enzyme activity by 50% and are the mean of two independent measurements, each performed in
duplicate. According to the inhibition kinetics, IC50 values were determined following a previously developed protocol (Zha8 et al) based on Ellman’s method (Ellman9 et al.)
and a standard incubation time of 20 min, if not otherwise specified.
b
Human recombinant AChE and BuChE from human serum were used.
% inhibition at 30 lM = 27.2 4.6.
IC50 value determined after a 120 min incubation period, according to the slower inhibition kinetics of 21g against hAChE.
IC50 value determined after a 60 min incubation period, according to the slower inhibition kinetics of 21g against hBuChE.
c
d
e
6949–6951; (c) Trost, B.; Zhang, Y. J. Am. Chem. Soc. 2006, 128, 4590–4591; (d)
Morales-Ríos, M.; Santos-Sánchez, N.; Joseph-Nathan, P. J. Nat. Prod. 2002, 65,
136–141; (e) Mukai, C.; Yoshida, T.; Sorimachi, M.; Odani, A. Org. Lett. 2006, 8,
83–86.
Acknowledgment
We would like to thank Stevens Institute of Technology for the
award of undergraduate fellowships to G.N., C.D., C.M., and G.M.
5. Alluri, S.; Feng, H.; Livings, M.; Samp, L.; Biswas, D.; Lam, T.; Lobkovsky, E.;
Ganguly, A. K. Tetrahedron Lett. 2011, 52, 3945–3948.
6. Zimmer, H.; Rothe, J.; Holbert, J. J. Org. Chem. 1960, 25, 1234–1235.
7. Murphy, J.; Rasheed, F.; Roome, S.; Scott, K.; Lewis, N. J. Chem. Soc., Perkin Trans. 1
1998, 2331–2340.
References and notes
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