Design and synthesis of potential new apoptosis agents: hybrid compounds containing perillyl alcohol and new constrained retinoids

Article abstract of DOI:10.1016/j.tetlet.2010.01.003

Novel retinoids 1-3 containing perillyl alcohol were synthesized through the addition of perillyl alcohol to the activated carboxylic acids (retinoids) promoted by DCC (N,N′-dicyclohexyl carbodiimide). A set of structurally and functionally diverse perillyl alcohol derivatives of retinoids were obtained in good yields (78-82%). Biological evaluation of these novel hybrid compounds (containing retinoids and perillyl alcohol) is currently underway in our laboratory.

Full text of DOI:10.1016/j.tetlet.2010.01.003

Tetrahedron Letters 51 (2010) 1462–1466
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Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Design and synthesis of potential new apoptosis agents: hybrid compounds
containing perillyl alcohol and new constrained retinoids
b
b
b
b
Bhaskar C. Das a,b, , Sakkarapalayam M. Mahalingam , Lipsa Panda , Bo Wang , Phillp D. Campbell ,
*
Todd Evans c
a Department of Nuclear Medicine, Albert Einstein College of Medicine, Bronx, NY 10461, USA
b Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, Bronx, NY 10461, USA
c Department of Surgery, Weill Cornell Medical College, Cornell University, New York, NY 10021, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Novel retinoids 1–3 containing perillyl alcohol were synthesized through the addition of perillyl alcohol
to the activated carboxylic acids (retinoids) promoted by DCC (N,N0-dicyclohexyl carbodiimide). A set of
structurally and functionally diverse perillyl alcohol derivatives of retinoids were obtained in good yields
(78–82%). Biological evaluation of these novel hybrid compounds (containing retinoids and perillyl alco-
hol) is currently underway in our laboratory.
Received 7 December 2009
Revised 29 December 2009
Accepted 5 January 2010
Available online 18 January 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Apoptosis is an essential cellular process for normal develop-
ment and homeostasis of multicellular organisms, to eliminate un-
wanted or damaged cells. Deregulated apoptosis plays a major role
in many human diseases, including cancer.1–4 Defects in the apop-
tosis machinery can confer apoptosis resistance of cancer cells to
therapeutic agents, which can render current anticancer therapies
less effective, and may lead ultimately to their failure in the
clinic.2–4 Accordingly, targeting critical regulators of apoptosis to
overcome apoptosis resistance is a promising cancer therapeutic
strategy. Perillyl alcohol and all-trans-retinoic acid (ATRA) have
antitumour activity toward liver, colon and hematopoietic tu-
mours, and are used as chemotherapeutic agents.5 The chemo-pre-
ventive activity of perillyl alcohol and ATRA is associated with an
increase in apoptosis of the tumour cells. Therefore, we hypothe-
sized that hybrid molecules containing both perillyl alcohol and
ATRA might further increase chemo-preventive activity through
enhancing apoptosis. The hybrid concept to synthesize biological
active molecules is a frontier area of research and this concept
has successfully employed to synthesize new therapeutic agents
for Alzheimer’s diseases.6
Toward this goal, we first sought to synthesize compounds
4a–b as new retinoic acid analogues by introducing a constrained
phenyl ring system in the place of conjugated alkene backbone
(spacers in ATRA) to inhibit the metabolism of ATRA into its iso-
mers, 9-CRA and 13-CRA (Fig. 1).
Furthermore, we considered the synthesis of novel perillyl alco-
hol containing retinoids (1–3) for studying tumour cell-targeted
apoptosis. To our surprise, a search of the literature failed to uncover
any examples of perillyl alcohol containing retinoic acid derivatives.
Here we report the first synthesis of perillyl alcohol containing ret-
inoids by simple addition of perillyl alcohol to the activated carbox-
ylic acids (retinoids), promoted by DCC (Scheme 1).
R
O
O
1
7
11
B
R
OH
HO
6
2
9
15 OH
13
14
10
12
A
8
3
18
B
5
ATRA
4
4
a: R = CH3
b: R = H
5: R = CH3
6: R= Ph
O
Figure 1. Structure of ATRA and our new retinoic acid analogues. Metabolic positions of all-trans-retinoic acid (ATRA) are shown by arrows.
* Corresponding author. Tel.: +1 718 430 2422; fax: +1 718 430 8853.
E-mail addresses: bdas@aecom.yu.edu (B.C. Das), tre2003@med.cornell.edu
(T. Evans).
0040-4039/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2010.01.003

B. C. Das et al. / Tetrahedron Letters 51 (2010) 1462–1466
1463
We next explored the synthesis of the constrained retinoids 4a–
b and the possible retro-synthetic analysis as shown in Scheme 2.
In the process of synthesizing 4a, based on disconnection route
A (Scheme 2) starting from the allylic alcohol 13, we encountered
the unusual Wittig salt 14 which led to the formation of the unu-
sual compound 5 when the ylide 14 was treated with aldehyde 9
(Scheme 3).7
O
DCC, DCM, 1.5 h
then
OH
ATRA
OH
12 h, r.t, 68%
To explore the mechanism for forming the unusual Wittig salt
14, we first tried to understand the substitution effect at the a-po-
O
O
sition of the allylic alcohol 13. For that purpose we synthesized the
allylic alcohol 15 from Grignard reaction of b-cyclocitral 11 with
phenyl magnesium bromide in THF. The allylic alcohol 15 was con-
verted into the corresponding ylide using PPh3HBr followed by the
Wittig reaction with aldehyde 9 furnishing the unusual product 6
with 83% yield (Scheme 4).
1
Scheme 1.
When the a-position of the allylic alcohol was substituted with
hydrogen 17 (1° alcohol) we found both unusual and usual Wittig
products 18 and 4b with the ratio of 1:0.6, respectively (Scheme 4).
O
O
R
OMe
R
OH
From these experiments we found that when the
substituted with methyl, phenyl gives exclusively the unusual Wit-
tig salt, whereas when the -position is substituted with hydrogen,
a-position is
O
+
B
a
4
PPh3
12
both usual and unusual products are obtained.
10: R = CH3
11: R = H
a: R = CH3
b: R = H
To further explore the formation of the unusual Wittig product,
we examined various solvents. The allylic alcohol 13 gave the unu-
sual Wittig product (5) in methanol, DMF and acetonitrile (Table 1,
entry 1). Interestingly, allylic alcohol 17 gave the normal Wittig
product (4b) in DMF (Table 1, entry 2) but produced mixture of
usual (4b) and unusual (18) Wittig products in both methanol
and acetonitrile (Table 1, entry 2). This observation indicates that
A
O
R
+
OMe
PPh3
O
the
a-methyl group plays an important role in the reaction. We
9
7: R = CH3
8: R = H
next examined the mechanistic insight to study the unusual Wittig
salt formation using the cyclic allylic alcoholic systems (the alcohol
group is inside the ring system).
Scheme 2. Retrosynthetic analysis.
tBuONa, DMF
H3CO2C CHO HO
.
PPh3 HBr,
OH
CH3CN
H
5; 85%
90 oC, 6 h,
92%
PPh3.Br
13
9
O
14
Scheme 3.
Ph
.
1. PPh3 HBr,
O
OH
Ph
CH3CN, 90 oC, 6 h
PhMgBr
H
2. tBuONa, DMF
THF, r.t, 3 h
11
15
H3CO2C
CHO
HO
6; 83%
NaBH4,
ethanol,
r.t, 3 h
9
O
O
.
1. PPh3 HBr,
OH
CH3CN, 90 oC, 6 h
OH
+
2. tBuONa, DMF
HO
O
H3CO2C
CHO
17
4b
(1:1)
18
9
Scheme 4. Results obtained from route A.

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B. C. Das et al. / Tetrahedron Letters 51 (2010) 1462–1466
Table 1
Studies of the solvent effect of the unusual Wittig reaction
Entry
Allylic alcohol
Product
Yielda (%)
(product ratio)b
MeOH
DMF
CH3CN
O
OH
OH
85
83
90
1
+
13
(5:4a = 1:0)
(5:4a = 1:0)
(5:4a = 1:0)
HO
O
4a
5
O
OH
OH
80
78
76
2
+
(18:4b = 1:0.6)
(18:4b = 0:1)
(18:4b = 1:0.6)
17
HO
O
4b
18
a
Isolated yield refers to aldehyde.
The ratios of the usual and unusual Wittig products were calculated from the 1H NMR.
b
To our surprise reactions utilizing allylic alcohols inside the ring
system (Table 2, entries 1–4) generated the more normal Wittig
Table 2 (continued)
Entry Allylic alcohol (19a–d)
Product
Yield (%)b
(E:Z)c
R1
R2
OH
Table 2
Studies of the Wittig reactions alcohol attached to the ring systema
4
69 (99:1)d
CO2H
20d: (R1 = H, R2 = 4-
HO2CC6H4)
H
.
1. PPh3 Br,
OH
a
All reactions were performed using 1 equiv of aldehyde and 3 equiv of tBuONa
CH3CN, 90 oC, 6 h
in DMF for 8–12 h.
n
n
b
2. tBuONa, DMF
Isolated yield (mixture of E and Z) refers to aldehyde.
R2
R2
The E:Z ratios were determined by 1H NMR.
c
R1
R1
R3
R3
Trace amount of Z isomer was observed in 1H NMR.
d
H3CO2C
CHO
19a-d
20a-d
9
n = 1, 2, 3; R1 = R2 = R3 =H/CH3
products (20a–d). We examined cyclic 5-, 6- and 7- member ring
systems and in all cases we isolated the regular Wittig products.
These new derivatives 20a–d also have the hydrophobic cyclo-
alkene ring and constrained phenyl ring system in the place of con-
jugated alkene backbone in ATRA, and the carboxylic acid group
remains intact. We compared the structural relation of our original
target-constrained retinoids 4a–b with the new unusual products
5–6 and these new compounds 20a–d. Having an extension of
the double bond conjugation via the 4th position of the cycloalkene
ring unit may block or limit retinoic acid metabolism caused by
cytochrome P450 enzymes (CYPs). This could be very advanta-
geous, since it is believed that the most prominent physiological
degradation pathway starts with the rate-limiting hydroxylation
at the C-4 position of the cyclohexenyl ring, leading to the forma-
tion of 4-hydroxy-ATRA, followed by a reductase enzyme into 4-
oxo-ATRA and further transformation by CYP(s) into more polar
metabolites.8
Entry
Allylic alcohol (19a–d)
Product
Yield (%)b
(E:Z)c
OH
R2
R1
1
76 (99:1)d
20a: (R1 = H, R2 = 4-
HO2CC6H4)
OH
R2
R1
2
3
83 (85:15)
20b: (R1 = H, R2 = 4-
HO2CC6H4)
Then we tried to synthesize our original target-constrained ret-
inoids 4a–b through the disconnection route B (Scheme 1). First we
tried with the ylide 4-[(triphenyl-phosphanyl)-methyl]-benzoic
acid methyl ester 12 with b-cyclocitral 11 with different reaction
conditions and the reactions failed to obtain the constrained reti-
noid 4a. We then prepared the reactive ylide 4-(diethoxy-phospho-
rylmethyl)-benzoic acid methyl ester 21, and treating with b-
OH
R2
R1
85 (72:28)
20c: (R1 = H, R2 = 4-
HO2CC6H4)
t
cyclocitral 11 using BuOK as a base yielded the final product 4b
with 79% yield (Table 3, entry 3).

B. C. Das et al. / Tetrahedron Letters 51 (2010) 1462–1466
1465
Table 3
Results obtained from route B
O
O
O
R
OH
O
t
R
BuOK
+
4
THF, r.t, 3 h
EtO P O
OEt
a: R1 = CH3
b: R1= H
10: R1 = CH3
11: R1= H
21
Entry
R
Reaction condition
Yielda (%)
1
2
3
4
5
6
7
H
H
H
tBuONa in DMF
tBuONa in THF
tBuOK in THF
tBuONa in DMF
tBuONa in THF
tBuOK in THF
TMG, THF
—
16
79
—
—
—
CH3
CH3
CH3
CH3
—
a
Isolated yield.
R
R
DCC, DCM, 1.5 h
then
OH
O
HO
12 h, r.t, 68%
O
O
5: R = CH
6: R = Ph
3
2: R = CH ; 78%
3: R = Ph; 75%
3
Scheme 5.
However, very surprisingly the Wittig reaction of ylide 4-(dieth-
oxy-phosphorylmethyl)-benzoic acid methyl ester 21 with ketone
10 clearly failed to give the olefination product 4a.
ported by the Albert Einstein College of Medicine and in part by
Grants from the NSF (DBI9601607 and DBI0331934), the NIH
(RR017998) and the HHMI Research Resources for Biomedical Sci-
ences. T.E is supported by the NIH (HL056182, HL064282).
Encouraged by these results and having established a new strat-
egy for synthesizing the new retinoid derivatives we synthesized
the perillyl alcohol containing constrained retinoids 2–3.9 These
were synthesized from the acids 5 and 6 using DCC as a carboxylic
acid activating agent in DCM. The activated carboxylic acid was
treated with perillyl alcohol in DCM at room temperature and gen-
erated the products 2 and 3 with 78% and 75% yields, respectively
(Scheme 5).
Supplementary data
Supplementary data (copies of 1H, 13C NMR and Mass spectra)
associated with this article can be found, in the online version, at
doi:10.1016/j.tetlet.2010.01.003.
In conclusion, we have synthesized novel constrained retinoids
(4b, 5, 6 and 20a–d) and found when alcohols are outside the ring
produce unusual Wittig salts, whereas when alcohols are inside the
ring produce usual Wittig salts. These acids were further esterified
using perillyl alcohol to synthesize new perillyl alcohol containing
retinoids through DCC coupling. This is based on the addition of
perillyl alcohol to the activated carboxylic acids, promoted by
DCC. A novel set of structurally and functionally diverse perillyl
alcohol containing retinoids were obtained in moderate to good
yields. Biological evaluation of these novel hybrid retinoids is cur-
rently underway in our laboratory.
References and notes
1. Kerr, J. F.; Wyllie, A. H.; Currie, A. R. Br. J. Cancer 1972, 26, 239–257.
2. Thompson, C. B. Science 1995, 267, 1456–1462.
3. Nicholson, D. W. Nature 2000, 407, 810–816.
4. Reed, J. C. Nat. Rev. Drug Discovery 2002, 1, 111–121.
5. Keith, R. S.; Jamie, H. M.; Yvette, D. B.; Yvonne, A. B.; Pamela, L. C. Carcinogenesis
1997, 18, 1655–1658.
6. Pelayo, C.; Xavier, F.; Carles, G.; Tánia, G.; Diego, M.; Michele, S.; Elisabet, V.;
Albert, B.; Victória Clos, M.; Antoni, C.; Mercé, P.; Manuela, B.; Francesca, M.;
Vincenza, A.; Joan, E.; Mónica, L.; Axel, B.; Javier Luque, F. J. Med. Chem. 2008, 51,
3588–3598.
7. (a) Das, B. C.; Kabalka, G. W. Tetrahedron Lett. 2008, 49, 4695; (b) Das, B. C.;
Mahalingam, S. M.; Evans, T.; Kabalka, G. W.; Anguiano, J.; Hema, K. Chem.
Commun. 2009, 2133.
8. Napoli, J. L. FASEB J. 1996, 10, 993–1001.
9. General procedure for synthesis of perillyl alcohol containing new retinoids (1–3): A
mixture of all trans-retinoic acid (100 mg, 0.35 mmol) in dry CH2Cl2 (5 mL) was
stirred under nitrogen atmosphere was added DCC (79 mg, 0.38 mmol) and cat.
DMAP at room temperature. The reaction mixture was stirred for 2 h at room
temperature under nitrogen atmosphere. To this stirred solution perillyl alcohol
Acknowledgments
The author B.C.D is thankful to AECOM for start up funding. The
instrumentation in the AECOM Structural NMR Resource is sup-

1466
B. C. Das et al. / Tetrahedron Letters 51 (2010) 1462–1466
(58.6 mg, 0.38 mmol) was added drop wise. The reaction mixture was stirred at
Compound 2: 1H NMR (300 MHz, CDCl3): d 1.07–1.15 (m, 9H), 1.23–1.38 (m, 2H),
1.50–1.54 (m, 3H), 1.77 (s, 3H), 1.94 (s, 4H), 2.18–2.30 (m, 6H), 2.60–2.68 (m,
2H), 4.70–4.79 (m, 4H), 5.59 (s, 1H), 6.48 (s, 1H), 7.33–7.36 (d, J = 9 Hz, 2H),
8.00–8.03 (d, J = 9 Hz, 2H). 13C NMR (75 MHz, CDCl3): d 15.3, 15.7, 21.0, 23.5,
24.2, 25.1, 26.0, 27.1, 27.8, 28.0, 31.1, 35.8, 36.4, 39.2, 41.7, 69.4, 109.2, 120.8,
125.9, 127.8, 128.0, 129.8, 130.0, 133.8, 142.6, 144.7, 149.2, 150.8, 166.0. ESI MS:
[M+H]+ 419.2946, calcd 419.2950 for C29H39O2.
room temperature until TLC analysis indicated none remaining (about 10–12 h).
The reaction was quenched with saturated NH4Cl and extracted with ethyl
acetate. The extracts were washed with H2O and brine, then dried overage
Na2SO4, and evaporated. The residue was purified by flash column
chromatography using hexane/ethyl acetate (8:1) as the eluent to give 1 as
colourless oil.
Compound 1: 1H NMR (300 MHz, CDCl3): d 1.05 (s, 6H), 1.45–1.52 (m, 2H), 1.58–
1.67 (m, 2H), 1.73 (s, 3H), 1.76 (s, 3H), 1.97–2.10 (m, 6H), 2.10–2.20 (m, 4H),
2.38 (s, 3H), 4.53 (s, 2H), 4.76 (m, 2H), 5.78–5.82 (m, 2H), 6.12–6.15 (m, 1H),
6.20 (s, 1H), 6.25–6.31 (2H, d, J = 18 Hz), 6.92–7.05 (m, 1H). 13C NMR (75 MHz,
CDCl3): d 13.3, 14.3, 19.6, 21.2, 22.1, 26.8, 27.7, 29.4, 30.9, 33.5, 34.6, 39.9, 41.3,
68.2, 109.1, 118.7, 125.8, 129.1, 129.8, 130.4, 131.4, 133.4, 135.5, 137.6, 138.1,
140.0, 150.1, 153.4, 167.5.
Compound 3: 1H NMR (300 MHz, CDCl3): d 1.05 (s, 6H), 1.58–1.65 (m, 5H), 1.77
(s, 4H), 1.87 (s, 4H), 2.15–2.30 (m, 5H), 2.70–2.76 (m, 2H), 3.71 (s, 2H), 4.73–4.77
(m, 4H), 5.88 (br s, 1H), 6.57 (s, 1H), 7.17–7.25 (m, 3H), 7.29–7.33 (m, 2H), 7.38–
7.41 (d, J = 9 Hz, 2H), 8.02–8.05 (d, J = 9 Hz, 2H). 13C NMR (75 MHz, CDCl3): d
16.8, 21.2, 24.4, 25.0, 25.8, 27.0, 27.7, 28.2, 30.9, 35.3, 35.8, 36.1, 39.0, 41.6, 56.0,
69.7, 110.0, 121.8, 125.4, 128.7, 129.6, 129.7, 130.8, 133.0, 140.3, 142.5, 144.2,
145.0, 150.1, 166.8. ESI MS: [M+H]+ 481.3160, calcd 481.3107 for C34H40O2.