Synthesis and biological activities of azalamellarins

Article abstract of DOI:10.1002/asia.201000237

The synthesis of azalamellar-ins, a new series of lactam analogues of biologically active lamellarins, was achieved using CuI-mediated and microwave-assisted C-N_amide bond formation. Seventeen azalamellarins, including N-allylazalamellarins and

Full text of DOI:10.1002/asia.201000237

DOI: 10.1002/asia.201000237
Synthesis and Biological Activities of Azalamellarins
Sasiwadee Boonya-udtayan,[a] Nattawut Yotapan,[b] Christina Woo,[b] Carson J. Bruns,[b]
Somsak Ruchirawat,[a, b] and Nopporn Thasana*[a, b]
Abstract: The synthesis of azalamellar-
ins, a new series of lactam analogues of
biologically active lamellarins, was ach-
ieved using CuI-mediated and micro-
were synthesized and evaluated for
their cytotoxicity against the cancer
cell lines HuCCA-1, A-549, HepG2,
and MOLT-3. The results showed that
certain azalamellarins exhibited good
activities in the micromolar IC50 value
range (IC50 =the drug concentration
that causes 50% of cell-growth inhibi-
tion after 72 h of continuous exposure
to the test molecule), comparable to
their parent lamellarin analogue.
À
wave-assisted C Namide bond formation.
Keywords: antitumor
agents
·
Seventeen azalamellarins, including N-
allylazalamellarins and N-propylazala-
mellarins c-D, L-N, and J-dehydro J,
azalamellarins · microwave-assisted
reactions · lactams · lactones
Introduction
alogues of lamellarin have been synthesized with the aim of
improving their inhibitory potencies as well as other desira-
ble physicochemical properties.[5] Among the modifications
that were investigated to achieve these goals were the re-
placement of the lactone moiety with the lactam moiety,
which produced the desired azalamellarin D (2) to poten-
tially increase the solubility and stability.
Lactams occur in a wide range of synthetic and natural
product derivatives, many of which have potential uses as
pharmacologically active compounds.[6] This useful pharma-
cophore has attracted much interest specifically for its use
as an antitumor and anticancer agent.[7] Oxynitidine (3), a
benzo[c]phenanthridine alkaloid from Xanthoxylum (Ruta-
ceae), has been investigated as a potential antitumor and an-
tiviral agent (Scheme 1).[8] Pancratistatin (4), a highly oxy-
genated phenanthridone, shows a high level of in vivo
cancer-cell-growth inhibitory activity and antiviral activity.[9]
Indenoisoquinoline (NSC 314622) (5) acts as a novel non-
camptothecin topoisomerase I (top1) inhibitor that is cyto-
toxic in cancer cell cultures and is under development as a
potential anticancer agent.[10]
Lamellarins are compounds of marine origin that display a
wide range of biological activities including topoisomerase I
(top1) and kinase inhibition as well as cytotoxic activities in
a range of cancer cell cultures. As such, they are currently
reported to be in development as potential anticancer
agents.[1] As a novel, potent inhibitor of DNA top1, lamel-
larin D (LAM-D) (1) also displays potent cytotoxic activi-
ties against multidrug-resistant tumor cell lines and is highly
cytotoxic to prostate-cancer cell lines.[2] Recently our group
reported the synthesis[3] and structure–activity relationship
(SAR) study of 25 lamellarin derivatives based on a 6H-
[1]benzopyrano[4’,3’:4,5]pyrroloACTHNUGTRNEUNG[2,1-a]isoquinolinone penta-
cyclic planar chromophore.[4] Although several lamellarin
derivatives show potent anticancer potency in vitro, they
suffer from several drawbacks that result from relatively low
aqueous solubility and the relatively labile nature of the lac-
tone ring under basic conditions. Therefore, a number of an-
[a] S. Boonya-udtayan, Prof. Dr. S. Ruchirawat, Dr. N. Thasana
Program on Chemical Biology
À
Previous methods of forming C N bonds have been re-
Center of Excellence on Environmental Health
Toxicology and Management of Chemicals (ETM)
Chulabhorn Graduate Institute, Laksi, Bangkok 10210 (Thailand)
ported extensively.[11] The Buchwald–Hartwig amination is a
popular method in modern organocopper chemistry to form
[12]
À
various C N bonds.
Indeed, copper-mediated cross-cou-
[b] N. Yotapan, C. Woo, C. J. Bruns, Prof. Dr. S. Ruchirawat,
Dr. N. Thasana
À
pling reactions to form aryl C N bonds have been studied
under microwave conditions between arylboronic acids and
pyrazinones.[13]
Laboratory of Medicinal Chemistry
Chulabhorn Research Institute, Laksi, Bangkok 10210 (Thailand)
Fax : (+662)574-2027-1503
The target phenanthridin-6-one,[14] which consists of a d-
lactam in the center of the structure, has attracted attention
owing to its presence in natural products and ability to un-
E-mail: nopporn@cri.or.th
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/asia.201000237.
Chem. Asian J. 2010, 5, 2113 – 2123
ꢀ 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
2113

FULL PAPERS
reaction induced by
a copper(I)thiophene-2-carboxylate
(CuTC),[15] and is facilitated by microwave irradiation for
milder conditions and faster reaction times.[16]
Results and Discussion
Model Study in the Synthesis of Azalamellarin
Aminolysis[17] of the 2-halobiarylcarboxylates 13[16] with
available amines 9a–d was studied to reduce the sequence
from the ester to the amides into one step. The amide for-
mation under microwave (MW) irradiation using trimethyla-
luminum in toluene as solvent at 1008C gave the 2-halobiar-
ylcarboxamides 14a–d in moderate to good yields. With var-
ious amides 14 in hand, the CuI-mediated and MW-assisted
Scheme 1. Lamellarin D (1), azalamellarin D (2), and biologically active
compounds 3–5 that contain aromatic lactam.
À
C Namide bond formation was studied. The phenanthridin-6-
one derivatives 15a–d were obtained in moderate to excel-
lent yields (27 to 96%) under mild conditions with a tem-
perature of 2008C in DMF (Table 1). Copper-mediated lac-
tamization reactions allow for milder conditions and a faster
reaction time, compared to classical heating during the syn-
thesis of the O-analogue lactone.[16]
Having successfully developed a method for the synthesis
of phenanthridin-6-one derivatives 15, and with the desire to
study the effect of substitution patterns on the SAR, we
then applied the above approach to the synthesis of azala-
mellarin model 17. The tandem Michael-addition/ring-clo-
sure (Mi–RC) Grob-type condensation of benzyldihydroiso-
quinoline 11a and a-nitrocinnamate 12a using NaHCO3 in
acetonitrile heated at reflux gave ethyl-1-aryl-2-(2-bromoar-
dergo further transformations (Scheme 1). Accordingly, we
À
made an effort to develop the method for forming Caryl
Namide bonds among phenanthridin-6-one derivatives and
successfully applied it to the synthesis of azalamellarins,
lactam analogues of lamellarins, as shown in Scheme 2. The
intramolecular cross-coupling consists of an Ullmann-type
yl)-5,6-dihydropyrroloACTHNUTRGNE[NUG 2,1-a]isoquinoline-3-carboxylate (10)
in moderate yield (54%; Scheme 3). The key step involves
the Mi–RC of the imines 11 and mixture of cis/trans nitro-
cinnamate derivatives 12.[3c] Ethyl-1-aryl-2-(2-bromoaryl)-
5,6-dihydropyrroloACTHNUTRGENUG[N 2,1-a]isoquinoline-3-carboxylates 18 to
20 were also synthesized using Mi–RC chemistry and were
used further in the synthesis of six pairs of N-allyl and N-
propylazalamellarin c-D, L-N, and J-dehydro J analogues,
which are synthetic analogues that mimic natural lamellar-
ins.
The aminolysis[17] of compound 10 with various amines
9a–d was studied using trimethylaluminum (2m AlMe3) in
toluene as solvent at 1008C to give 1,2-biarylpyrroloisoqui-
noline-3-carboxamide derivatives 16a–d in good yields (76
to 94%) (Table 2, entries 1 to 4). The CuI-mediated and
Scheme 2. Retrosynthetic plan of azalamellarin synthesis.
Abstract in Thai:
Scheme 3. Synthesis of compounds 10 and 18–20.
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Synthesis and Biological Activities of Azalamellarins
Table 1. Synthesis of phenanthridin-6-ones 15 using CuI-mediated and MW-assisted Caryl Namide coupling.
À
bined with an increase in
amine to 4–5 equiv and AlMe3
to 2.0–2.4 equiv, led to much
improved yields (Table 3, en-
tries 5 to 7).
From the above-mentioned
improved amidation condi-
tions, the reaction of carboxy-
lates 18–20 with 5.0 equiv
amine 9d in the presence of
2.0 equiv AlMe3 in THF gave
carboxamides 23–25 in moder-
ate to good yields. The CuI-
Entry
Products
X
Amidation yield [%] 14[a]
Lactamization yield [%] 15[b]
1
2
15a
Br
Cl
14a, 96
14b, 93
15a, 37
15a, 72
3
4
15b
15c
15d
Br
Cl
14c, 84
14d, 86
15b, 62
15b, 72
À
mediated and MW-assisted C
Namide bond formation of car-
boxamides 23–25 were per-
formed in DMF under mild
condition with a temperature
of 1508C in order to affect the
lactamization. These conditions
gave benzyl-protected azala-
mellarins 26–28 in good yields
(Scheme 4).
5
6
Br
Cl
14e, 82
14 f, 85
15c, 96
15c, 55
7
8
Br
Cl
14g, 38[c]
14h, 41[d]
15d, 31
15d, 27
Synthesis of Azalamellarins
c-D, L-N, and J-dehydro J
[a] Amidation conditions: ester (1 equiv), amine (1.2 equiv), AlMe3 (1 equiv), toluene, MW 100 W, 1008C,
100 psi, 5 min. [b] Lactamization: CuTC (0.5 equiv), DMF, MW 200–250 W, 2008C, 38–100 psi, 10–60 min.
[c] Recovered starting material 61% yield. [d] Recovered starting material 57% yield.
We next turned to the deben-
zylation step that leads to satu-
rated N-allylazalamellarins 29–
À
MW-assisted C Namide bond formation was then studied
under mild conditions with a temperature of 2008C in DMF
to yield azalamellarins 17a–d in good to excellent yields (74
to 98%).
31 and saturated N-propylazalamellarins 33–35, respectively,
depending on the methodology. A hydrogenolysis route was
used for both cleavage of benzyl group and reduction at the
C2’–C3’ position to yield N-propylazalamellarins 33–35 in
good yields, whereas the saturated N-allylazalamellarins 29–
31 were obtained from the application of solid-support de-
benzylation as shown in Scheme 5.[18] This reaction was per-
formed initially for the polydebenzylation of the complex
molecules by using pTsOH immobilized on silica (PTS–Si)
Synthesis of Benzyl-Protected Azalamellarins 26–28
We then turned our attention to the synthesis and cytotoxic
evaluation of azalamellarin analogues of c-D, L-N, and J-de-
hydro J, which are mimics of natural lamellarins. The effec-
tive methodology of the synthesis of azalamellarins 17a–d
was used as the key step, including the use of solid support
for the debenzylation step and the oxidation step of C5–C6-
saturated azalamellarins to unsaturated azalamellarins. The
reaction of dihydroisoquinolines 11b–d and a-nitrocinna-
mate 12b in the presence of NaHCO3 and acetonitrile
heated at reflux overnight gave the protected form of com-
pounds 18–20 in moderate yields (32–43%; Scheme 3). Ami-
dation of the carboxylates was conducted under microwave
irradiation at 2008C using AlMe3 in toluene, as previously
reported, to generate carboxamide derivatives. However,
substitution of the benzyl groups on the periphery signifi-
cantly lowered the yields. Increasing the equivalent amount
of amine 9a, AlMe3, temperature, and reaction time gave
disappointing yields (Table 3, entries 1 to 4). We then modi-
fied the amidation conditions to improve the yield. THF
was found to be the most effective solvent and, when com-
Scheme 4. Synthesis of saturated N-allylazalamellarins 26–28.
Chem. Asian J. 2010, 5, 2113 – 2123
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N. Thasana et al.
FULL PAPERS
Table 2. Synthesis of azalamellarins 17 using CuI-mediated and MW-assisted Caryl Namide coupling.
À
larins 29–31 were purified and
obtained in moderate yields.
The main side product was
characterized as monobenzyl-
substituted diarylmethane 32
of benzyl-cation product in a
tiny quantity that was support-
ed by NMR spectroscopic data
(see the Supporting Informa-
tion).
Entry
Products
Amidation yield [%]
Lactamization yield [%]
16[a]
17[b]
1
17a
17b
17c
17d
16a, 76[c]
16b, 89
16c, 94
16d, 80
17a, 90
17b, 98
17c, 60[d]
17d, 74
With the C5–C6-saturated
precursors in hand, we next in-
vestigated the preparation of
the unsaturated form. Both N-
allylazalamellarins and N-pro-
pylazalamellarins were investi-
gated in the same manner and
involved three steps as shown
in Schemes 6 and 7. Treatment
of saturated azalamellarins
with acetyl chloride in the
presence of 4-dimethylamino-
pyridine (DMAP) and triethyl-
amine in dichloromethane at
room temperature furnished
acetates 36–41. To elaborate
the C5–C6 moiety, the intro-
duction of the double bond
was achieved by using 2,3-di-
chloro-5,6-dicyanobenzoqui-
none (DDQ) in dichloroethane
heated at reflux overnight to
yield the desired products 42–
47 in high yields. Finally, the
acetyl groups were hydrolyzed
under basic conditions to give
the final products, unsaturated
azalamellarins 48–53.
2
3
4
[a] Amidation conditions: ester (1 equiv), amine (1.2 equiv), AlMe3 (1 equiv), toluene, MW 100 W, 1008C,
100 psi, 5 min. [b] Lactamization: CuTC (0.5 equiv), DMF, MW 200–250 W, 2008C, 38–100 psi, 10–60 min.
[c] AlMe3 (1 equiv), toluene, 300 W, 2508C, 100 psi, 20–40 min. [d] 300 W, 2508C, 100 psi, 60 min, recovered
starting material 24% yield.
Table 3. Conditions for the MW-assisted amidation of ethyl dihydropyrrolo
derivatives.
ACHTUNGTRENN[UNG 2,1-a]isoquinoline-3-carboxylate
In addition, the N-unsubsti-
tuted azalamellarin D (2) was
also studied to determine the
SAR associated with the side
chain. Azalamellarin
D (2)
was prepared by means of
deallylation of N-allylazala-
mellarin 26 by using [ClRh-
Entry
Ester[a]
Solvent[c]
9a [equiv]
AlMe3 [equiv]
T [8C]
t [min]
Product yield [%]
1
2
3
4
5
6
7
19
19
19
19
toluene
toluene
toluene
toluene
THF
1.1
1.5
1.8
3.0
4.0
5.0
4.0
2.0
2.4
2.8
3.0
2.4
2.4
2.0
250
300
250
250
100
100
100
60
60
90
60
20
20
40
22, 15
22, 2
22, 8
AHCTNUGTREN(GNNU PPh3)3] and OsO4 with NaIO4
for isomerization and oxida-
tion,[19] respectively, to yield
tribenzyl azalamellarin 54,
which was further debenzylat-
ed by hydrogenolysis to cleave
the benzyl groups (Scheme 8).
After acetylation with acetic
anhydride, followed by oxida-
22, 7
19
22, 60
21, 77
21, 75
18
THF
THF
18[b]
[a] Reaction scale is 0.1 mmol. [b] Reaction scale is 1.0 mmol. [c] MW power 300 W for toluene and 100 W for
THF.
in toluene and a minute amount of methanol under a con-
stant temperature of 808C. After filtration, N-allylazalamel-
tion using DDQ, which utilized the same conditions as N-
allyl and N-propylazalamellarins, the azalamellarin D triace-
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Chem. Asian J. 2010, 5, 2113 – 2123

Synthesis and Biological Activities of Azalamellarins
Scheme 6. Synthesis of unsaturated N-allylazalmellarins D, N, and dehy-
dro J (48–50)
Scheme 5. Synthesis of saturated N-allylazalamellarins c, L, and J (29–31)
and N-propylazalamellarins c, L, and J (33–35).
DMSO. Azalamellarins 17a–b and 17d as well as carboxa-
mide analogues 16a–d, except N-phenylazalamellarin 17c,
are soluble in DMSO. Interestingly, N-allylazalamellarin
17d showed cytotoxicity in a lower micromolar IC50 range
(3.435 mm) against the HepG2 hepatoblastoma. The open
lactam ring of this azalamellarin, N-allylcarboxamide 16d,
gave lower cytotoxicity by an order of magnitude
(38.51 mm). The other side chains of lactams and carboxa-
mides resulted in either a considerable decrease or no ob-
served potency. This study provided crucial information
about the importance of the lactam moiety to significantly
increase the solubility of lamellarin derivatives in the biolog-
tate 55 was obtained. Basic hydrolysis yielded azalamellarin
D (2) in 4% overall yield from azalamellarin 18.
Biological Activity of Azalamellarins 17a–d and Their
Corresponding Carboxamides 16a–d
To examine the effect of the N-aryl/alkyl side chain of syn-
thetic azalamellarins, a panel of four human tumor cell lines
was used to evaluate the cytotoxic potential of azalamell-
ACHTUNGTRENNUNGarins 17a–d and their corresponding carboxamides 16a–d;
the cell lines were HuCCA-1 cholangiocarcinoma, A549
lung carcinoma, HepG2 hepatoblastoma, MOLT-3 T-lym-
phoblast (acute lymphoblastic
leukemia). 3-(4,5-Dimethylth-
iazol-2-yl)-2,5-diphenyltetrazo-
lium bromide (MTT) and 2,3-
bis-(2-methoxy-4-nitro-5-sulfo-
phenyl)-2H-tetrazolium-5-car-
ical medium. The N-allylACTHNUTRGNEaNUG mine was used as a side chain for
boxanilide (XTT) assays were
used to estimate IC50 values,
that is, the drug concentration
that causes 50% of cell-growth
inhibition after 72 h of contin-
uous exposure to the test mol-
ecule. The results obtained are
shown in Table 4.
We found that azalamell-
A
with
hexamethoxyl
groups have significantly in-
creased solubility in DMSO
compared with lamellarin G
trimethyl ether, which is the
core lactone structure of this
class but is insoluble in
Scheme 7. Synthesis of unsaturated N-propylazalmellarins D, N, and dehydro J (51–53)
Chem. Asian J. 2010, 5, 2113 – 2123
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N. Thasana et al.
FULL PAPERS
ture of lamellarins. Most synthetic azalamellarins exhibited
better cytotoxic activities than the positive control etoposide
in HepG2 cell line at micromolar levels of IC50 as shown in
Table 5.
Without any side chains, azalamellarin D (2) was more cy-
totoxic than its N-allyl/propyl side-chain analogues among a
panel of four human tumor cell lines (Table 5, entry 6) and
gave results that were as good as its equivalent lactone core,
lamellarin D (Table 5, entry 7).
Conclusion
In conclusion, we have achieved the synthesis of azalamel-
larin D (2) and its derivatives by means of a copper(I)-medi-
À
ated and microwave-assisted C Namide bond formation pro-
cess. The method we have developed and reported here
should be applicable to the
Scheme 8. Synthesis of azalamellarin D (2).
synthesis of other lactam sys-
tems in natural products and
synthetic compounds. Interest-
ingly, this study reveals for the
first time a significant contri-
bution from the change of a
lactone to a lactam moiety,
which in turn significantly in-
creases the aqueous solubility
of the corresponding lamellar-
in lactam derivatives in the
biological medium. Azalamel-
larin D (2) exhibited good ac-
tivities in the micromolar IC50
value range, comparable to its
parent lamellarin D (1). The
presence of allyl and propyl
side chains on the lactam ni-
Table 4. Biological activities of azalamellarins 17a–d and their corresponding carboxamides 16a–d in MTT
and XTT assays.[a]
Entry
Compound
IC50 [mM]
HuCCA-1[c,g]
A549[d,g]
HepG2[e,g]
MOLT-3[f,h]
1
2
3
4
5
6
7
8
9
16a
16b
16c
16d
inactive
inactive
inactive
inactive
inactive
inactive
N/A
34.35
0.52
ND
inactive
inactive
inactive
75.51
inactive
inactive
N/A
42.94
0.43
ND
inactive
inactive
inactive
38.51
inactive
inactive
N/A
3.435
ND
27.18
11.98
6.40
5.11
12.55
0.21
1.37
N/A
2.85
ND
17a
17b
17c[b]
17d
doxorubicin
etoposide
10
0.017
[a] Soluble in DMSO (10 mgmLÀ1). [b] Insoluble in DMSO. [c] HuCCA-1: Cholangiocarcinoma. [d] A549:
Lung carcinoma. [e] HepG2: Hepatoblastoma. [f] MOLT-3: T-lymphoblast (acute lymphoblastic leukemia).
[g] MTT assay. [h] XTT assay. N/A=Not available owing to the insolubility of the compounds in DMSO.
ND=Not determined.
the synthesis of the subsequent azalamellarin analogues, as
discussed in the preceding sections.
trogen of the azalamellarins resulted in either a considerable
decrease or no observed potency. This is the first report of a
structure–activity relationship at this position, since previous
studies have focused on lamellarin D and its derivatives,
which possess a hydrogen atom at C7, a C5=C6 double
bond, C8 and C20 hydroxy groups, as well as a lactone
moiety. This SAR study has helped to delineate the impor-
tant structural elements in the lactone–lactam skeleton that
contribute to cytotoxicity. Information from our synthetic
work and SAR studies is critical to improve our understand-
ing of the structural requirements for good anticancer activi-
ty and to help us more efficiently design new molecules to
probe SAR further.
Biological Activity of Saturated and Unsaturated N-Allyl
and N-Propylazalamellarins c-D, L-N, and J-dehydro J
The importance of the lactam in the cytotoxic activities of
azalamellarins was studied by comparing the IC50 of six
pairs of azalamellarins and azalamellarin D (2), each of
which contains exactly the same substituents and differs
only in the side chain of N-lactam. Thirteen azalamellarins,
both saturated and unsaturated N-allyl and N-propylazala-
mellarins c-D, L-N, J-dehydro J, and azalamellarin D were
evaluated for their cytotoxicity using MTT and XTT assays
depending on the cell-line types. Although the cytotoxicities
of azalamellarins were only comparable to the native lac-
tone structure of lamellarins, we found the tendency of the
selective activity of azalamellarins that was effective against
HepG2 cell line. The presence of allyl and propyl side
chains of azalamellarins showed a significant decrease in the
biological activities compared to their native lactone struc-
Experimental Section
General Methods
Microwave reactions were performed with a CEM Discover instrument.
Melting points were determined with an electrochemical melting-point
apparatus and reported without correction. 1H NMR spectra were re-
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Chem. Asian J. 2010, 5, 2113 – 2123

Synthesis and Biological Activities of Azalamellarins
Table 5. Biological activities of synthetic azalamellarins and their natural lamellarins in MTT and XTT assays.[a]
Entry
Structure
Compound
IC50 [mM]
HuCCA-1[b]
A549[b]
HepG2[b]
MOLT-3[c]
1
2
3
azalam 29; X=N-allyl
azalam 33; X=N-nPr
lam c X=O
31.4
19.35
4.0
20.3
19.35
8.3
8.64
8.85
0.95
3.09
2.30
0.48
4
5
6
7
azalam 48; X=N-allyl
azalam 51; X=N-nPr
azalam D 2; X=NH
lam D 1; X=O
19.50
inactive
0.12
7.06
inactive
0.74
4.70
inactive
0.13
1.28
0.425
0.03
0.078
0.085
0.28
0.0049
8
9
10
azalam 30; X=N-allyl
azalam 34; X=N-nPr
lam L; X=O
7.68
56.2
inactive
7.46
49.76
2.99
4.07
10.93
1.75
3.53
3.58
0.66
11
12
13
azalam 49; X=N-allyl
azalam 52; X=N-nPr
lam N; X=O
7.43
18.5
inactive
6.50
8.75
inactive
6.50
5.55
0.27
2.60
1.55
0.011
14
15
16
azalam 31; X=N-allyl
azalam 35; X=N-nPr
lam J; X=O
66.7
23.4
inactive
17.1
11.68
inactive
10.5
9.0
inactive
2.24
1.49
0.388
17
18
19
azalam 50; X=N-allyl
azalam 53; X=N-nPr
dehydrolam J; X=O
inactive
10.82
0.22
inactive
5.86
0.3
inactive
5.7
0.34
2.00
0.577
0.0064
20
21
doxorubicin
etoposide
0.52
ND
0.75
ND
0.60
28.9
ND
0.027
[a] Soluble in DMSO (10 mgmLÀ1). [b] MTT assay. [c] XTT assay. HuCCA-1=Cholangiocarcinoma. A549=Lung carcinoma. HepG2=Hepatoblastoma.
MOLT-3=T-lymphoblast (acute lymphoblastic leukemia). ND=Not determined.
corded with 200 MHz and 400 MHz instruments at 200 and at 400 MHz,
respectively; 13C NMR spectra were recorded with 200 and 400 MHz in-
struments at 50 and at 100 MHz, respectively. FTIR spectra were record
with a universal ATR (UATR) accessory. Low-resolution mass spectra
were obtained with an LC–MS instrument using electron-impact ioniza-
tion (EI). High-resolution mass spectra were obtained with a MicroTOF
instrument using atmospheric pressure chemical ionization (APCI) in
positive or negative mode. Column chromatography was carried out
using aluminum oxide (100–125 mesh) or Merck silica gel (70–230 mesh).
Thin-layer chromatography (TLC) and preparative thin-layer chromatog-
raphy (PTLC) were carried out on silica gel. All reagents were purified
and dried according to the standard procedures.
General Procedure for the Preparation of DihydropyrroloACTHNGUTERN[UNG 2,1-
a]isoquinoline Carboxylates (18–20)
A mixture of benzyldihydroisoquinoline 11 (1.5 equiv), nitrocinnamate
12 (1 equiv), and NaHCO3 (1.5 equiv) in acetonitrile (15 mL) was heated
to reflux for 16 h. After being allowed to cool to RT, water and EtOAc
were added. The two layers were separated, and the aqueous phase was
extracted with EtOAc (3ꢁ). The combined organic layers were washed
once with brine, dried over anhydrous Na2SO4, filtered, and concentrated
under reduced pressure to give a crude product, which was further puri-
fied by column chromatography on silica (30% EtOAc/hexanes) to fur-
nish the desired products in moderate yield (32–43%):
Compound 18
Employing the general procedure for the synthesis of pyrrolo
ACHTUNGTRENNUNG[2,1-a]iso-
quinoline carboxylate with benzyldihydroisoquinoline 11b (4.85 g,
Chem. Asian J. 2010, 5, 2113 – 2123
ꢀ 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemasianj.org
2119

N. Thasana et al.
FULL PAPERS
9.83 mmol) and nitrocinnamate 12b (2.86 g, 6.55 mmol) in acetonitrile
(50 mL) gave 18 as
Compound 23
a
yellow sticky gum (1.68 g, 33%). 1H NMR
Employing the general procedure with ester 18 (87.6 mg, 0.1 mmol), allyl
amine 9d (28.5 mg, 0.5 mmol), and AlMe3 (0.12 mL) in THF (1 mL)
(60 min, 1508C, 300 W, 100 psi) gave product 23 as a yellow sticky gum
(80.1 mg, 90%). 1H NMR (400 MHz, CDCl3, TMS): d=2.89–3.07 (m,
(400 MHz, CDCl3, TMS): d=0.86 (t, 3JH,H =7.1 Hz, 3H; CH3), 2.88–3.10
(m, 2H; CH2), 3.23 (s, 3H; CH3), 3.63 (s, 3H; CH3), 3.65 (s, 3H; CH3),
3.92–4.1 (m, 2H; CH2), 4.33 (m, 1H; CHH), 4.88 (m, 1H; CHH), 5.10 (d,
2JH,H =2.4 Hz, 2H; CH2), 5.13 (s, 4H; 2CH2), 6.55 (s, 1H; CH), 6.71–6.75
(m, 5H; 5CH), 7.04 (s, 1H; CH), 7.28–7.44 ppm (m, 15H; 15CH);
13C NMR (100 MHz, CDCl3, TMS): d=13.6, 28.9, 42.7, 55.2, 55.8, 56.0,
59.7, 70.6, 70.9, 71.0, 108.9, 113.2, 113.5, 114.3, 115.0, 115.6, 116.9, 118.6,
121.3, 121.4, 122.8, 125.5, 127.0, 127.2, 127.3, 127.8, 127.9, 128.0, 128.2,
128.5, 128.6, 130.4, 130.8, 131.6, 136.5, 136.9, 137.2, 146.5, 147.1, 147.2,
2H; CH2), 3.23 (s, 3H; CH3), 3.66 (s, 6H; 2CH3), 3.78–3.82 (m, 2H;
2
CH2), 4.39 (m, 1H; CHH), 4.84–4.93 (m, 3H; 2CHH), 5.10 (d, JH,H
=
2.0 Hz, 2H; CH2), 5.12(s, 4H; 2CH2), 5.55–5.65 (m, 2H; NH, CH), 6.66
(s, 1H; CH), 6.68 (s, 1H; CH), 6.72–6.74 (m, 4H; 4CH), 7.09 (s, 1H;
CH) 7.28–7.43 ppm (m, 15H; 15CH); 13C NMR (100 MHz, CDCl3, TMS):
d=29.0, 41.5, 43.0, 55.2, 55.8, 56.1, 70.7, 71.0, 71.2, 108.7, 113.3, 113.6,
114.2, 115.5, 115.7, 115.8, 117.4, 120.7, 121.5, 121.7, 122.8, 125.5, 126.4,
127.0, 127.2, 127.4, 127.8, 127.8, 128.2, 128.4, 128.5 (2C), 128.6, 129.3,
À
147.8, 148.1, 149.2, 161.5 ppm; IR (UATR): n˜max =1739 (C=O), 1251 (C
+
+
O), 1211 cmÀ1 (C N); EIMS: m/z (%): 881 (31) [M+2H ], 879 (32) [M
À
], 789 (43), 712 (15), 698 (17), 91 (100); HRMS (FAB): m/z: calcd for
133.8, 136.1, 137.0, 137.1, 146.5, 146.9, 147.8, 148.3, 149.0, 149.2,
C51H47BrNO8: 880.2480; found: 880.2458.
À1
À
161.6 ppm; IR (UATR): n˜max =3423 (N H), 1640 cm (C=O); EIMS: m/
z (%): 892 (0) [M+2H+], 890 (0) [M+], 802 (25), 711 (29), 631 (28), 541
(24), 91 (100); HRMS (FAB): m/z: calcd for C52H48BrN2O7: 891.2639;
found: 891.2665.
Compound 19
Employing the general procedure for the synthesis of pyrroloACTHNUGRTNEUNG[2,1-a]iso-
quinoline carboxylate with benzyldihydroisoquinoline 11c (2.49 g,
5.05 mmol) and nitrocinnamate 12b (1.47 g, 3.37 mmol) in acetonitrile
(25 mL) gave 19 as
Compound 24
a
yellow sticky gum (0.84 g, 32%). 1H NMR
(400 MHz, CDCl3, TMS): d=0.86 (t, 3JH,H =7.1 Hz, 3H; CH3), 2.98 (m,
2H; CH2), 3.32 (s, 3H; CH3), 3.62 (s, 3H; CH3), 3.82 (s, 3H; CH3), 4.02
(m, 2H; CH2), 4.37 (m, 1H; CHH), 4.87(m, 1H; CHH), 4.90 (s, 2H;
CH2), 5.08 (d, 2JH,H =3.7 Hz, 2H; CH2), 5.14 (s, 2H; CH2), 6.55 (s, 1H;
CH), 6.65 (s, 1H; CH), 6.74 (s, 1H; CH), 6.76–6.81 (m, 3H; 3CH), 7.03
(s, 1H; CH), 7.24–7.44 ppm (m, 15H; 15CH); 13C NMR (100 MHz,
CDCl3, TMS): d=13.6, 28.8, 42.7, 55.2, 55.9 (2C), 59.6, 70.8, 70.9, 71.0,
108.9, 111.4, 113.1, 115.0, 115.5, 116.3, 116.9, 118.5, 121.2, 121.3, 123.5,
125.5, 127.2, (4C), 127.3 (2C), 127.7 (2C), 127.8 (2C), 127.9 (2C), 128.4
(2C), 128.5 (4C), 130.3, 130.9, 131.5, 136.4, 136.9, 147.1, 147.2, 147.8,
148.0, 148.3, 161.5 ppm; IR (UATR): n˜max =1739 cmÀ1 (C=O); EIMS: m/z
(%): 881 (0) [M+2H+], 879 (0) [M+], 698 (13), 91 (100); HRMS (FAB):
m/z: calcd for C51H47BrNO8: 880.2480; found: 880.2456.
Employing the general procedure with ester 19 (93.6 mg, 0.1 mmol), allyl
amine 9b (30.2 mg, 0.53 mmol), and AlMe3 (0.12 mL) in THF (1 mL)
(60 min, 1508C, 300 W, 100 psi) gave product 24 as yellow sticky gum
(55.0 mg, 62%). 1H NMR (400 MHz, CDCl3, TMS): d=2.90–3.07 (m,
2H; CH2), 3.32 (s, 3H; CH3), 3.65 (s, 3H; CH3), 3.79 (m, 2H; CH2), 3.83
(s, 3H; CH3), 4.40 (m, 1H; CHH), 4.84–4.93 (m, 1H; CHH), 4.91 (d,
2JH,H =1.6 Hz, 2H; CH2), 4.91 (s, 2H; CH2), 5.08 (d, 2JH,H =3.8 Hz, 2H;
CH2), 5.14 (s, 2H; CH2), 5.53–5.65 (m, 2H; CH, NH), 6.62 (s, 1H; CH),
6.65 (s, 1H; CH), 6.73 (s, 1H; CH), 6.78 (m, 3H; 3CH), 7.08 (s, 1H;
CH), 7.23–7.46 ppm (m, 15H; 15CH); 13C NMR (100 MHz, CDCl3,
TMS): d=29.0, 41.5, 43.0, 55.2, 55.9, 56.1, 70.8, 70.9, 71.1, 108.7, 111.5,
113.2, 115.4, 115.7, 115.8, 116.1, 117.3, 120.7, 121.5, 121.6, 123.5, 125.5
(2C), 126.4, 127.2 (3C), 127.4 (2C), 127.8 (2C), 127.9 (2C), 128.2 (2C),
128.4 (2C), 128.6 (4C), 129.4, 133.9, 136.1, 136.9, 137.0, 146.9, 147.8,
À
148.3, 148.9, 161.6 ppm, 1C not observed; IR (UATR): n˜max =3420 (N
Compound 20
H), 1695 cmÀ1 (C=O); EIMS: m/z (%): 892 (0) [M+2H+], 890 (0) [M+],
564 (23), 179 (31), 91 (100); HRMS (FAB): m/z: calcd for C52H48BrN2O7:
891.26; found: 891.2632.
Employing the general procedure for the synthesis of pyrroloACTHNUGRTNEUNG[2,1-a]iso-
quinoline carboxylate with benzyldihydroisoquinoline 11d (2.54 g,
6.1 mmol) and nitrostyrene 12b (1.78 g, 4.07 mmol) in acetonitrile
(25 mL) gave 20 as
a
yellow sticky gum (1.40 g, 43%). 1H NMR
Compound 25
(400 MHz, CDCl3, TMS): d=0.87 (t, 3JH,H =7.1 Hz, 3H; CH3), 2.92–3.10
(m, 2H; CH2), 3.37 (s, 3H; CH3), 3.65 (s, 3H; CH3), 3.65 (s, 3H; CH3),
3.84 (s, 3H; CH3), 3.93–4.11 (m, 2H; CH2), 4.39 (m, 1H; CHH), 4.89 (m,
1H; CHH), 5.10 (d, 2JH,H =2.5 Hz, 2H; CH2), 5.15 (s, 2H; CH2), 6.57 (s,
1H; CH), 6.74 (d, 4JH,H =2.2 Hz, 1H; CH), 6.75 (s, 2H; 2CH), 6.75 (d,
3JH,H =8.6 Hz, 1H; CH), 6.81 (dd, 3JH,H =8.0 Hz, 4JH,H =1.6 Hz, 1H; CH),
7.03 (s, 1H; CH), 7.29–7.45 ppm (m, 10H; 10CH); 13C NMR (100 MHz,
CDCl3, TMS): d=13.6, 28.9, 42.7, 55.3, 55.7, 55.8, 56.0, 59.7, 71.0 (2C),
109.0, 110.7, 113.2, 113.9, 115.0, 115.6, 116.9, 118.6, 121.3, 121.4, 122.9
(2C), 125.6, 127.2 (2C), 127.3 (2C), 127.7, 127.9, 128.0, 128.6 (3C), 130.4,
Employing the general procedure with ester 20 (195.0 mg, 0.24 mmol),
allyl amine 9b (68.9 mg, 1.21 mmol), and AlMe3 (0.29 mL) in THF
(1 mL) (60 min, 1508C, 300 W, 100 psi) gave product 25 as a yellow sticky
gum (168.0 mg, 86%). 1H NMR (400 MHz, CDCl3, TMS): d=2.91–3.08
(m, 2H; CH2), 3.37 (s, 3H; CH3), 3.66 (s, 3H; CH3), 3.67 (s, 3H; CH3),
3.80 (m, 2H; CH2), 3.83 (s, 3H; CH3), 4.41 (m, 1H; CHH), 4.85–4.93 (m,
3H; CH2, CHH), 5.10 (d, 2JH,H =1.9 Hz, 2H; CH2), 5.14 (s, 2H; CH2),
5.55–5.66 (m, 2H; CH, NH), 6.68 (s, 1H; CH), 6.73–6.81 (m, 5H; 5CH),
7.09 (s, 1H; CH), 7.29–7.47 ppm (m, 10H; 10CH); 13C NMR (100 MHz,
CDCl3, TMS): d=29.0, 41.5, 43.0 (2C), 55.3, 55.7, 55.8, 56.1, 71.0 (2C),
108.8, 110.8, 113.3, 113.7, 115.4, 115.7, 115.8, 117.4, 120.8, 121.5, 121.7,
122.9, 125.6, 126.4, 127.2 (2C), 127.4 (2C), 127.9 (2C), 128.2 (2C), 128.5
130.9, 131.6, 136.5, 136.9, 147.1, 147.2, 147.6, 147.9, 148.1, 148.5,
À1
À
À
161.5 ppm; IR (UATR): n˜max =1687 (C=O), 1251 (C O), 1212 cm (C
N); EIMS: m/z (%): 805 (58) [M+2H+], 803 (61) [M+], 725 (32), 713
(84), 622 (4), 91 (36); HRMS (FAB): m/z: calcd for C45H43BrNO8:
804.2167; found: 804.2158
(2C), 128.6 (2C), 129.3, 133.8, 136.0, 137.0, 147.0, 147.6, 147.8, 148.2,
À1
À
148.5, 148.9 ppm; IR (UATR): n˜max =3416 (N H), 1695 cm (C=O);
EIMS: m/z (%): 816 (0) [M+2H+], 814 (3) [M+], 576 (4), 91 (100);
HRMS (FAB): m/z: calcd for C46H44BrN2O7: 815.2326; found: 815.2343.
General Procedure for Preparation of DihydropyrroloACHTNUTRGNEUNG[2,1-a]isoquinoline
Carboxamides 23–25
General Lactamization Procedure (26–28)
In a 10 mL microwave vessel, a mixture of ester 18–20 (1 equiv), amine
9d (5 equiv), and trimethylaluminum (2m AlMe3 in toluene, 2.4 equiv) in
THF (1 mL) was sealed and heated in a microwave reactor to 1508C for
60 min (100 W, 120 psi). The reaction was quenched with water and ex-
tracted with CH2Cl2. The combined organic layer was washed with water,
brine, and dried over anhydrous Na2SO4, filtered, and concentrated
under reduced pressure to give a crude product, which was further puri-
fied by column chromatography on silica (30% EtOAc/hexanes) to fur-
nish the desired products as a yellow sticky gum (62–90%):
In a 10 mL microwave vessel, a mixture of dihydropyrroloACHTUNTRGNEU[GN 2,1-a]isoquino-
line carboxamides 23–25 (1 equiv) and CuTC (2.8 equiv) in DMF (1–
2 mL) was sealed and heated in a microwave reactor to 1508C for 30 min
(200 W, 100 psi). The reaction was monitored by means of TLC until
completion. The reaction was filtered on silica gel and concentrated
under reduced pressure to yield a yellow-brown oil. The crude product
was then purified by column chromatography on silica (30% EtOAc/hex-
anes) to furnish the desired products (71–84%):
2120
www.chemasianj.org
ꢀ 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Chem. Asian J. 2010, 5, 2113 – 2123

Synthesis and Biological Activities of Azalamellarins
Compound 26
reaction was cooled to room temperature, diluted with H2O, and extract-
ed with CH2Cl2. The combined organic layers were washed with brine,
dried over anhydrous Na2SO4, filtered, and concentrated under reduced
pressure to give a crude product, which was further purified by prepara-
tive thin-layer chromatography (PTLC) to give the desired product 54 as
Employing the general procedure with amide 23 (348.2 mg, 0.4 mmol)
and CuTC (209.5 mg, 1.1 mmol) in DMF (2 mL) (30 min, 1508C, 200 W,
100 psi) gave product 26 as
a yellow sticky gum (225.0 mg, 71%).
1H NMR (400 MHz, CDCl3, TMS): d=3.01 (t, 3JH,H =7.2 Hz, 2H; CH2),
1
a brown oil (162.7 mg, 42%). H NMR (400 MHz, CDCl3, TMS): d=3.03
3.28 (s, 3H; CH3), 3.34 (s, 3H; CH3), 3.87 (s, 3H; CH3), 4.86–5.04 (m,
(brt, 2H; CH2), 3.29 (s, 3H; CH3), 3.35 (s, 3H; CH3), 3.87 (s, 3H; CH3),
4.85–5.00 (m, 2H; CH2), 5.15 (d, 2JH,H =8.0 Hz, 2H; CH2), 5.27 (s, 2H;
CH2), 5.30 (s, 2H; CH2), 6.72 (s, 1H; CH), 6.75 (s, 1H; CH), 6.76 (s, 1H;
CH), 6.76 (s, 1H; CH), 7.04 (d, 3JH,H =8.07 Hz, 1H; CH), 7.07 (s, 1H;
4H; 2CH2), 4.96 (dd, 3JH,H =17.5 Hz, JH,H =1.0 Hz, 1H; CHH), 5.08 (dd,
2
3JH,H =10.5 Hz, 2JH,H =1.0 Hz, 1H; CHH), 5.14 (s, 2H; CH2), 5.19 (s, 2H;
CH2), 5.27 (s, 2H; CH2), 5.80–5.90 (m, 1H; CH), 6.70 (s, 1H; CH), 6.74
(s, 1H; CH), 6.79 (s, 1H; CH), 6.89 (s, 1H; CH), 7.02–7.09 (m, 3H;
3CH), 7.27–7.49 ppm (m, 15H; 15CH); 13C NMR (100 MHz, CDCl3,
TMS): d=28.9, 29.7, 42.3, 44.3, 55.1, 55.2, 56.2, 70.9 (2C), 71.4, 102.7,
105.7, 108.9, 112.4, 113.3, 114.1, 114.6, 114.7, 116.4, 118.8, 121.2, 123.6,
125.5, 126.5, 127.0, 127.1, 127.2, 127.9, 128.0, 128.6 (2C), 128.7, 129.9,
130.3, 132.4, 133.6, 136.8, 137.0, 144.9, 146.8, 147.4, 147.5, 147.9, 150.4,
3
CH), 7.08 (d, JH,H =7.8 Hz, 1H; CH), 7.39 (m, 15H; 15CH), 9.68 ppm (s,
1H; NH); 13C NMR (100 MHz, CDCl3, TMS): d=28.9, 42.4, 55.2, 55.3,
56.2, 70.9 (2C), 71.0, 101.1, 105.3, 108.9, 111.3, 113.4, 114.6, 114.7, 116.9,
118.7, 119.0, 121.1, 123.6, 126.3, 127.0 (2C), 127.2 (2C), 127.3 (2C), 128.0
(2C), 128.6 (3C), 128.7 (2C), 129.2, 129.6, 136.8, 137.0, 147.4, 148.0,
À
150.4, 156.1 ppm, 5C not observed; IR (UATR): n˜max =3355 (N H), 1651
155.5 ppm; IR (UATR): n˜max =1636 (C=O), 1212, 1180 cmÀ1 (C N);
À
+
(C=O), 1214 cmÀ1 (C N); EIMS: m/z (%): 770 (2) [M ], 680 (6), 91
À
EIMS: m/z (%): 810 (8) [M+], 630 (35), 629 (9); HRMS (FAB): m/z:
(100); HRMS (FAB): m/z: calcd for C49H43N2O7:771.3065; found:
calcd for C52H47N2O7: 811.3378; found: 811.3406.
771.3088.
Compound 27
Compound 55
Employing the general procedure with amide 24 (50.1 mg, 0.06 mmol)
and CuTC (29.9 mg, 0.16 mmol) in DMF (2 mL) (30 min, 1508C, 200 W,
100 psi) gave product 27 as a yellow sticky gum (38.1 mg, 84%). H NMR
A solution of lactam 54 (1 equiv) in EtOAc was placed in a high-pressure
Parr apparatus at room temperature. Palladium on activated charcoal
(ca. 100 mg) was added to this solution. The resulting mixture was hydro-
genated (75 psi) until all starting material was consumed (normally 15 h)
as indicated by TLC. The mixture was then filtered through a plug of
Celite and concentrated under reduced pressure to give a gray solid. The
obtained gray solid (1 equiv) was then treated with Et3N (4.5 equiv),
DMAP (1 equiv), and acetic anhydride (4.5 equiv) in CH2Cl2
(25 mLmmolÀ1 of starting material) at 08C. The resulting mixture was
stirred at room temperature and monitored with TLC until completion.
The reaction was quenched with water and extracted with CH2Cl2. The
combined organic layer was washed with water, brine, and dried over an-
hydrous Na2SO4, filtered, and concentrated under reduced pressure to
give a crude product, which was further oxidized using DDQ (2.5 equiv)
in dichloroethane (100 mLmmolÀ1 of starting material) while heating to
reflux for 16 h. After being allowed to cool to RT, water and CH2Cl2
were added. The combined organic layers were washed with brine, dried
over anhydrous Na2SO4, filtered, and concentrated under reduced pres-
sure to give a crude product, which was further purified by column chro-
matography on silica (1% MeOH/CH2Cl2) to furnish the desired product
1
(400 MHz, CDCl3, TMS): d=3.01 (t, 3JH,H =6.7 Hz, 2H; CH2), 3.34 (s,
3H; CH3), 3.40 (s, 3H; CH3), 3.95 (s, 3H; CH3), 4.86–5.01 (m, 4H;
2CH2), 4.92 (d, 3JH,H =17.2 Hz, 1H; CHH), 5.09 (d, 3JH,H =10.5 Hz, 1H;
CHH), 5.12 (d, 2JH,H =2.6 Hz, 2H; CH2), 5.15 (s, 2H; CH2), 5.20 (s, 2H;
CH2), 5.81–5.90 (m, 1H; CH), 6.64 (s, 1H; CH), 6.74 (s, 1H; CH), 6.79
(s, 1H; CH), 6.89 (s, 1H; CH), 7.11 (s, 1H; CH), 7.12 (d, 3JH,H =6.1 Hz,
1H; CH), 7.13 (d, 3JH,H =6.4 Hz, 1H; CH), 7.21–7.44 ppm (m, 15H;
15CH); 13C NMR (100 MHz, CDCl3, TMS): d=29.0, 42.3, 44.3, 55.1,
55.2, 56.4, 70.9 (2C), 71.4, 102.7, 105.8, 108.9, 112.5, 112.6, 113.2, 114.0,
116.5, 116.8, 118.8, 121.2, 124.2, 125.5, 126.4, 127.1 (2C), 127.2 (2C),
127.4 (2C), 127.9 (2C), 128.0 (2C), 128.5 (2C), 128.6 (4C), 129.3, 130.3,
132.4, 133.7, 136.4, 136.9, 144.9, 146.8, 147.5, 147.8, 148.9, 149.3,
155.5 ppm; IR (UATR): n˜max =1635 cmÀ1 (C=O); EIMS: m/z (%): 810
(0.87) [M+], 720 (10), 629 (12), 91 (100); HRMS (FAB): m/z: calcd for
C52H47N2O7: 811.3378; found: 811.3398.
Compound 28
1
Employing the general procedure with amide 25 (609.8 mg, 0.75 mmol)
and CuTC (199.4 mg, 1.05 mmol), in DMF (2 mL) (30 min, 1508C, 200 W,
55 as pale yellow solid (45.1 mg, 60%). M.p. 2818C; H NMR (400 MHz,
CDCl3, TMS): d=2.34 (s, 3H; CH3), 2.35 (s, 3H; CH3), 2.38 (s, 3H;
100 psi) gave product 28 as
a brown sticky gum (451.4 mg, 82%).
CH3), 3.45 (s, 3H; CH3), 3.45 (s, 3H; CH3), 3.84 (s, 3H; CH3), 6.94 (d,
1H NMR (400 MHz, CDCl3, TMS): d=3.03 (t, 3JH,H =6.6 Hz, 2H; CH2),
3.37 (s, 3H; CH3), 3.44 (s, 3H; CH3), 3.86 (s, 3H; CH3), 3.96 (s, 3H;
CH3), 4.87 (d, 3JH,H =2.37 Hz, 2H; CH2), 4.95 (m, 3H; CH2, CHH), 5.09
(dd, 3JH,H =10.5 Hz, 2JH,H =1.0 Hz, 1H; CHH), 5.14 (s, 2H; CH2), 5.19 (s,
2H; CH2), 5.81–5.90 (m, 1H; CH), 6.73 (s, 1H; CH), 6.76 (s, 1H; CH),
3JH,H =7.5 Hz, 1H; CH), 6.96 (s, 1H; CH), 7.01 (s, 1H; CH), 7.21 (s, 1H;
3
CH), 7.25 (d, 3JH,H =6.4 Hz, 1H; CH), 7.25 (s, 1H; CH), 7.29 (d, JH,H
=
3
5.9 Hz, 1H; CH), 7.36 (s, 1H; CH), 9.49 ppm (d, JH,H =7.4 Hz, 1H; CH);
13C NMR (100 MHz, CDCl3, TMS): d=20.6 (3C), 55.5 (2C), 56.2, 195.0,
106.4 (2C), 110.4, 111.2, 111.8, 114.0, 115.3, 115.6, 120.5,123.4, 123.5,
123.8, 123.9, 127.6, 129.5, 131.6, 135.4, 139.6, 140.0, 140.1, 146.6, 150.5,
4
6.80 (s, 1H; CH), 6.93 (s, 1H; CH), 7.08 (d, JH,H =1.5 Hz, 1H; CH), 7.10
(s, 1H; CH), 7.13 (dd, 3JH,H =8.1 Hz, 4JH,H =1.8 Hz, 1H; CH), 7.28–
7.43 ppm (m, 10H; 10CH); 13C NMR (100 MHz, CDCl3, TMS): d=29.0,
42.3, 44.3, 55.2 (2C), 56.1, 56.2, 70.9, 71.4, 102.7, 105.8, 108.9, 112.0 (2C),
113.3, 114.1, 114.2, 116.5, 118.8, 121.2, 123.8, 125.5, 126.5, 127.1 (2C),
127.2 (2C), 127.9 (3C), 128.6 (4C), 129.4, 130.4, 132.4 (2C), 133.7, 136.8,
145.0, 146.8, 147.5, 147.9, 148.6, 149.8 ppm; IR (UATR): n˜max =1636 cmÀ1
(C=O); EIMS: m/z (%): 735 (10) [M+H+], 644 (46), 553 (39), 91 (51);
HRMS (FAB): m/z: calcd for C46H43N2O7: 735.3065; found: 735.3037.
À
152.3, 156.1, 168.9, 169.2 ppm (2C); IR (UATR): n˜max =3442 (N H), 1765
(C=O), 1648 cmÀ1 (C=O); EIMS: m/z (%): 624 (35) [M+], 582 (69), 540
(100), 498 (56), 43 (29); HRMS (FAB): m/z: calcd for
C39H29N2O10:625.1817; found: 625.1838.
Compound 2
Compound 55 (42 mg, 0.067 mmol) was dissolved by 5% KOH/EtOH so-
lution (KOH 1 equiv). The resulting mixture was stirred at room temper-
ature and monitored with TLC until completion. The reaction was acidify
with 2n HCl until reaching pH 7 and extracted with EtOAc. The com-
bined organic layer was washed with water, brine, and dried over anhy-
drous Na2SO4, filtered, and concentrated under reduced pressure to give
Compound 54
ACHTUNGTRENNUNG[ClRhACHTUNGTRENNUNG(PPh3)3] (10 mol%) was added to a solution of lactam 26 (1 equiv)
in dry toluene (40 mLmmolÀ1 of starting material). The resulting mixture
was heated at reflux for 14 h, cooled to room temperature, concentrated,
and passed through a short plug of Celite (3% MeOH/CH2Cl2) to gener-
ate a crude product as a brown viscous oil that was used for the next re-
action without further purification. The crude product in THF
(10 mLmmolÀ1 of starting material) was added to OsO4 followed by a so-
lution of NaIO4 in H2O (10 mLmmolÀ1 of starting material). The reaction
mixture was heated to 808C and monitored with TLC. At that time, the
desired product
2 as a
pale green solid (23.7 mg, 71%). 1H NMR
(400 MHz, (CD3)2SO): d=3.33 (s, 3H; CH3), 3.36 (s, 3H; CH3), 3.75 (s,
3H; CH3), 6.79 (s, 1H; CH), 6.88 (s, 1H; CH), 6.99 (dd, 3JH,H =7.5,
1.9 Hz, 1H; CH), 7.00 (d, 3JH,H =7.1 Hz, 1H; CH), 7.08 (d, 3JH,H =8.0 Hz,
1H; CH), 7.11 (s, 1H; CH), 7.12 (s, 1H; CH), 7.12 (s, 1H; CH), 9.36 (d,
3JH,H =7.2 Hz, 1H; CH), 9.27 (s, 1H; OH), 9.49 (s, 1H; OH), 9.71 (s, 1H;
OH), 11.3 ppm (s, 1H; NH); 13C NMR (100 MHz, (CD3)2SO): d=54.9,
Chem. Asian J. 2010, 5, 2113 – 2123
ꢀ 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemasianj.org
2121

N. Thasana et al.
FULL PAPERS
55.2, 56.4, 102.7, 105.9, 106.3, 109.0, 110.5, 110.8, 112.0, 112.4, 115.6,
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À
147.5, 147.8, 148.4, 149.1, 155.8 ppm; IR (UATR): n˜max =3356 (N H),
1605 cmÀ1 (C=O); EIMS: m/z (%): 499 (29) [M+H+], 498 (100) [M+],
95 (12); HRMS (FAB): m/z: calcd for C28H23N2O7: 499.1500; found:
499.1506.
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Cytotoxicity Test[4]
All azalamellarins were dissolved in DMSO and tested for their cytotoxic
activities against HuCCA-1, A-549, HepG2, and MOLT-3 cancer cell
lines. The cells suspended in the corresponding culture medium were ino-
culated in 96-well microtiter plates (Corning Inc., NY, USA) at a density
of 10000–20000 cells per well, and incubated at 378C in a humidified at-
mosphere of 95% air and 5% CO2. After 24 h, an equal volume of addi-
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pounds, positive control (etoposide), or negative control (DMSO) was
added to the desired final concentrations, and the microtiter plates were
further incubated for an additional 48 h. The number of surviving cells in
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XTT assay (for suspended cells) to determine the IC50, which is defined
as the concentration that inhibits cell growth by 50% (relative to nega-
tive control) after 48 h of continuous exposure to each test compound.
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Acknowledgements
Financial support from the Thailand Research Fund (TRF; RMU4980048
for N.T), the award of the Royal Golden Jubilee Scholarship (PHD/0028/
2552 for S.B.), and from NSF-REU research fellowship (CHE-0453126
for C.J.B. and C.W.) are gratefully acknowledged. The Center of Excel-
lence on Environmental Health, Toxicology and Management of Chemi-
cals (ETM) is gratefully acknowledged. The authors are thankful to Dr.
Poonsakdi Ploypradith and Ms. Supattra Karnkra for providing lamellar-
in derivatives. Special thanks go to Ms. Pagamas Intachote (Laboratory
of Immunology) and Ms. Busakorn Saimanee (Integrated Research Unit)
Chulabhorn Research Institute for conducting the cytotoxicity assays.
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Received: March 30, 2010
Published online: August 2, 2010
Chem. Asian J. 2010, 5, 2113 – 2123
ꢀ 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemasianj.org
2123