Total synthesis of seco (+)- and ent-(-)-oxaduocarmycin SA: Construction of the (chloromethyl)indoline alkylating subunit by a novel intramolecular aryl radical cyclization onto a vinyl chloride

Article abstract of DOI:10.1021/jo971880b

A practical, total synthesis of seco-(+)-oxaduocarmycin 3a, an analogue of the highly cytotoxic natural product, duocarmycin SA (1), is described. the 13-step synthesis feature a novel and efficient intramolecular aryl radical cyclization onto a vinyl chloride as a direct entry to the (chloromethyl)indoline alkylating subunit 14. Subsequent resolution, utilizing a preparative Chiralpak AD column, provided enantiomerically pure alkylating subunits 14a and 14b which were elaborated to seco-(+) and ent-(- )-oxduocarmycins, 3a and 3b. respectively. The natural enantiomer 3a was active at pM concentrations and exhibited 7-50-fold higher potentcy than its enantiomer 3b in in vitro cytotoxicity assays.

Full text of DOI:10.1021/jo971880b

8868
J . Org. Chem. 1997, 62, 8868-8874
Tota l Syn th esis of Seco (+)- a n d en t-(-)-Oxa d u oca r m ycin SA:
Con str u ction of th e (Ch lor om eth yl)in d olin e Alk yla tin g Su bu n it by
a Novel In tr a m olecu la r Ar yl Ra d ica l Cycliza tion on to a Vin yl
Ch lor id e
Vinod F. Patel,*,† Sherri L. Andis,‡ J ulia K. Enkema,‡ David A. J ohnson,‡
J oseph H. Kennedy,† Fariborz Mohamadi,1,† Richard M. Schultz,‡ Daniel J . Soose,† and
Michael M. Spees†
Lilly Research Laboratories, Cancer Research Division, Eli Lilly and Company,
Indianapolis, Indiana 46285
Received October 14, 1997X
A practical, total synthesis of seco-(+)-oxaduocarmycin 3a , an analogue of the highly cytotoxic
natural product, duocarmycin SA (1), is described. The 13-step synthesis features a novel and
efficient intramolecular aryl radical cyclization onto a vinyl chloride as a direct entry to the
(chloromethyl)indoline alkylating subunit 14. Subsequent resolution, utilizing a preparative
Chiralpak AD column, provided enantiomerically pure alkylating subunits 14a and 14b which were
elaborated to seco-(+) and ent-(-)-oxaduocarmycins, 3a and 3b, respectively. The natural
enantiomer 3a was active at pM concentrations and exhibited 7-50-fold higher potentcy than its
enantiomer 3b in in vitro cytotoxicity assays.
In tr od u ction
represent the most exciting agent in the series.7,8 The
first feature relates to the extent of the noncovalent DNA
binding8 which stabilizes the reversible9 alkylation of
DNA while providing (+)-duocarmycin SA (1) with the
full biological properties of the irreversible alkylating
agent, (+)-CC-1065 (2).10 This, in combination with the
implications that the extent of the noncovalent stabiliza-
tion may be related to the delayed, fatal hepatotoxicity
associated with (+)-CC-1065 (2),9a,10 suggests that (+)-
duocarmycin SA (1) may prove to be a better therapeutic
agent. Second, as the most stable natural product in this
class of agents,11 (+)-duocarmycin SA12 (1) is also the
most potent,6a a finding that is consistent with a correla-
tion between solvolytic stability and cytotoxic potency
that was established within a series of agents possessing
sufficient reactivity to alkylate DNA.13 Together, these
characteristics provide rationale for the development of
(+)-duocarmycin SA (1) and related agents as potent
oncolytics for the treatment of human cancers.14
(+)-Duocarmycin SA (1),2 isolated from Streptomyces
sp. DO113, is the newest member of a class of potent
antitumor antibiotics3,4 that are structurally and biologi-
cally related to (+)-CC-1065 (2).5 In common, these
natural products possess a cyclopropanoindolinone linked
to a DNA binding subunit as the pharmacophore respon-
sible for the sequence-selective minor groove alkylation
of duplex DNA5 and the exceptionally potent cytotoxic
activity.5,6 Detailed studies, however, reveal two critical
features that distinguish (+)-duocarmycin SA (1) in its
interaction with duplex DNA and suggest that it may
* To whom correspondence should be addressed.
† Cancer Chemistry, Eli Lilly & Co.
‡ Cancer Pharmacology, Eli Lilly & Co.
X Abstract published in Advance ACS Abstracts, December 1, 1997.
(1) Present address: Abbott Laboratories, 1 Abbott Park Rd., IL
60064-3500.
(2) Isolation: (a) Ichimura, M.; Ogawa, T.; Takahashi, K.; Koba-
yashi, E.; Kawamoto, I.; Yasuzawa, T.; Takahashi, I.; Nakano, H. J .
Antibiot. 1990, 43, 1037. (b) Ichimura, M.; Ogawa, T.; Katsumata, S.;
Takahashi, K.; Takahashi, I; Nakano, H. J . Antibiot. 1991, 44, 1045.
(c) Yasuzawa, T.; Muroi, K.; Ichumura, M.; Takahashi, I.; Ogawa, T.;
Takahashi, K.; Sano, H.; Saitoh, Y. Chem. Pharm. Bull. 1995, 43, 378.
Total syntheses: (d) See ref 30. (e) Boger, D. L.; Machiya, K.; Hertzog,
D. L.; Kitos, P. A.; Holmes, D. J . Am. Chem. Soc. 1993, 115, 9025. (f)
Muratake, H.; Abe, I.; Natsume M. Chem. Pharm. Bull. 1996, 44, 67.
(g) Murake, H.; Tonegawa, M.; Natsume, M. Chem. Pharm. Bull. 1996,
44, 1631.
(3) Duocarmycin A, B1, B2, C1, and C2: (a) Ichimura, M; Muroi, K.;
Asano, K.; Kawamoto, I.; Tomita, F.; Morimoto, M.; Nakano, H. J .
Antibiot. 1988, 41, 1285. (b) Takehashi, I.; Takehashi, K.; Ichimura,
M.; Morimoto, M.; Asano, K.; Kawamoto, I.; Tomita, F.; Nakano, H. J .
Antibiot. 1988, 41, 1915. (c) Yasuzawa, T.; Iida, T.; Muroi, K.; Ichimura,
M.; Takahashi, K.; Sano, H. Chem. Pharm. Bull. 1988, 36, 3728. (d)
Ogawa, T.; Ichimura, M.; Katsumata, S.; Morimoto, M.; Takahashi,
K. J . J . Antibiot. 1989, 42, 1299.
(6) (a) Boger, D. L.; Ishizaki, T.; Zarrinmayeh, H.; Munk, S. A.; Kitos,
P. A.; Suntornwat, O. J . Am. Chem. Soc. 1990, 112, 8961. (b) Sugiyama,
H.; Hosoda, M., Saito, I. Tetrahedron Lett. 1990, 31, 7197. (c) Warpe-
hoski, M. A. In Advances in DNA Sequence Specific Agents; Hurley, L.
H., Ed.; J AI: Greenwich, CT, 1992; Vol. 1, p 217. (d) Hurley, L. H.;
Draves, P. H. In Molecular Aspects of Anticancer Drug-DNA Interac-
tions; Neidle, S., Waring, M., Eds; CRC: Ann Arbor, MI, 1993; Vol. 1,
p 89.
(7) Boger, D. L. In Advances in Nitrogen Heterocycles; Moody, C. J .,
Ed.; J AI Press: Greenwich, CT, 1995; Vol. 1, pp 229-247.
(8) (a) Boger, D. L.; J ohnson, D. S.; Yun, W. J . Am. Chem. Soc. 1994,
116, 1635. (b) Boger, D. L.; J ohnson, D. S. Proc. Natl. Acad. Sci. U.S.A.
1995, 92, 3642. (c) Boger, D. L.; J ohnson, D. S. J . Am. Chem. Soc. 1995,
117, 1443.
(9) (a) Boger, D. L.; Yun, W. J . Am. Chem. Soc. 1993 115, 9872. (b)
Asai, A.; Nagamura, S.; Saito, H.; Takahashi, I; Nakano, H. Nucleic
Acids Res. 1994, 22, 88.
(10) Warpehoski, M. A.; Harper, D. E.; Mitchell, M. A.; Monroe, T.
J . Biochemistry 1992, 31, 2502.
(4) Pyrindamycin A and B: (a) Ohba, K.; Watabe, H.; Sasaki, T.;
Takeuchi, Y.; Kodama, S. J . Antibiot. 1988, 41, 1515. (b) Ishii, S.;
Nagasawa, M.; Kariya, Y.; Yamamoto, H.; Inouye, S.; Kondo, S. J .
Antibiot. 1989, 42, 1713.
(5) (a) Hurley, L. H.; Needham-VanDevanter, D. R. Acc. Chem. Res.
1986, 19, 230. (b) Warpehoski, M. A.; Hurley, L. H. Chem. Res. Toxicol.
1988, 1, 315. (c) Coleman, R. S.; Boger, D. L. In Studies in Natural
Products Chemistry; Atta-ur-Rahman, Ed.; Elsevier: Amsterdam, 1989;
Vol. 3, p 301. (d) Boger, D. L. In Advances in Heterocyclic Natural
Products Synthesis; Pearson, W. H., Ed.; J AI Press: Greenwich, CT,
1992; Vol. 2, pp 1-188. (e) Boger, D. L. Acc. Chem. Res. 1995, 28, 20.
(f) Boger, D. L.; J ohnson, D. S. Angew. Chem., Int. Ed. Engl. 1996, 35,
1439.
(11) (a) Boger, D. L.; Coleman, R. S.; Invergo, B. J .; Sakya, S. M.;
Ishizaki, T.; Munk, S. A.; Zarrinmayeh, H.; Kitos, P. A.; Thompson, S.
C. J . Am. Chem. Soc. 1990, 112, 4623. (b) Boger, D. L. In Heterocycles
in Bioorganic Chemistry; Bergman, J ., van der Plas, H. C. Simonyi,
M., Eds.; Royal Society of Chemistry: Cambridge, 1991. (c) Boger, D.
L. In Proc. Robert A. Welch Found. Conf. Chem. Res. 35th Chem. Front.
Med. 1991, 35, 137.
(12) The name duocarmycin SA originates from duocarmycin stable
A.
S0022-3263(97)01880-X CCC: $14.00 © 1997 American Chemical Society

Synthesis of Seco (+)- and ent-(-)-Oxaduocarmycin SA
J . Org. Chem., Vol. 62, No. 25, 1997 8869
Ch a r t 1
Sch em e 1
Seco-(+)-oxaduocarmycin SA 3a (LY307918) is an
analogue in which the fused pyrrole ring in the alkylating
subunit of 1 is replaced with a potentially metabolically
more inert furan ring,15 and the cyclopropanoindolinone
pharmacophore is masked as a (chloromethyl)hydroxy-
indoline, a prodrug form which is known to demonstrate
indistinguishable biological properties from its active
form.16 The unexpected observation that both (+)-1 and
its enantiomer ent-(-)-1 alkylate DNA with comparable
efficiency,17 prompted us to evaluate both (+)-3a and ent-
(-)-3b as potentially useful oncolytics. A practical
synthetic route was, therefore, required to provide suf-
ficient quantities of both the enantiomers for a full
evaluation of their biological properties.
Herein, we report an efficient and practical total
synthesis of (+)-3a and ent-(-)-3b in which the key
transformation involves a C-C bond formation via a
novel 5-exo-trig cyclization of an aryl radical onto a vinyl
chloride18 to provide a direct entry into the (chlorometh-
yl)indoline ring system (Scheme 1).19 Subsequent resolu-
tion, by chromatographic separation on a chiral column,
leads to enantiomerically pure alkylating subunits, which
are then elaborated to furnish both enantiomers of seco-
oxaduocarmycin SA (3).20
Resu lts a n d Discu ssion
Alkylation of the potassium salt of commercially avail-
able hydroxybenzaldehyde 4 with ethyl bromoacetate
cleanly provided 521 (Scheme 2). Intramolecular aldol
cyclization of 5 followed by dehydration was most ef-
fectively achieved in one step with DBU in refluxing
ethanol to directly give benzofuran 621 in a modest 44%
yield. O-Demethylation, with concomitant ester hydroly-
sis, occurred on heating 6 with excess, neat pyridine
hydrochloride22 at 170 °C to give hydroxy acid 7, which
was directly reesterified with acidic methanol to give the
desired methyl ester 8 in an 95% overall yield. Reduction
(18) To our knowledge, the intramolecular addition of carbon
radicals to vinyl chlorides has not been reported; however, vinyl
fluorides are known to be suitable radical acceptors: (a) Morikawa,
T.; Uchida, J .; Hasegawa, Y.; Taguchi, T. Chem. Pharm. Bull. 1991,
39, 2464. (b) Morikawa, T.; Uchida, J .; Imoto, K.; Taguchi, T. J .
Fluorine Chem. 1992, 58, 119. (c) Dolbier, W. R., J r.; Rong, X. X.
Tetrahedron Lett. 1996, 37, 5321. The corresponding intermolecular
reactions with vinyl chlorides and vinyl fluorides are well-docu-
mented: (d) Low, H. C.; Tedder, J . M.; Walton, J . C. J . Chem. Soc.,
Faraday Trans. 1976, 72, 1707. (e) Henning, R.; Urbach, H. Tetrahe-
dron Lett. 1983, 24, 5343. (f) Giese, B.; Gonzalez-Gomez, J . A.; Witzel,
T. Angew. Chem., Int. Ed. Engl. 1984, 23, 69.
(13) (a) Boger, D. L.; Ishizaki, T. Tetrahedron Lett. 1990, 31, 793.
(b) Boger, D. L.; Munk, S. A.; Ishizaki, T. J . Am. Chem. Soc. 1991,
113, 2779. (c) Ichimura, M.; Ogawa, T.; Takasahi, K.; Mihara, A.;
Takahashi, I.; Nakano, H. Oncol. Res. 1993, 5, 165. (d) Boger, D. L.;
Goldberg, J .; McKie, J . A. Bioorg. Med. Chem. Lett. 1996, 6, 1955.
(14) KW-2189, a novel derivative of duocarmycin B2, is presently
in clinical trials: (a) Asai, A.; Nagamura, S.; Saito, H. J . Am. Chem.
Soc. 1994, 116, 4171. (b) Kobayashi, E.; Okamato, A.; Asada, M.;
Okabe, M.; Nagamura, S.; Asai, A.; Saito, H.; Gomi, K.; Hirata, T.
Cancer Res. 1994, 54, 2404. (c) Nagamura, S.; Asai, A.; Kanda, Y.;
Kobayashi, E.; Gomi, K.; Saito, H. Chem. Pharm. Bull. 1996, 44, 1723.
(15) Mohammadi, F.; Spees, M. M.; Staten, G. S.; Marder, P.; Kipka,
J . K.; J ohnson, D. A.; Boger, D. L.; Zarrinmayeh, H. J . Med. Chem.
1994, 37, 232.
(19) Review: Boger, D. L.; Boyce, C. W.; Garbaccio, R. M.; Goldberg,
J . A. Chem. Rev. (Washington, D.C.) 1997, 97, 787.
(20) This approach provides a useful improvement to established
synthetic approaches to such agents: (a) 5-exo-dig aryl radical cycliza-
tion: Boger, D. L.; Ishizaki, T.; Wysocki, R. J ., J r.; Munk, S. A.; Kitos,
P. A.; Suntornwat, O. J . Am. Chem. Soc. 1989, 111, 6461. Boger, D.
L.; Ishizaki, T.; Kitos, P. A.; Suntornwat, O. J . Org. Chem. Soc. 1990,
55, 5823. (b) 5-exo-trig aryl radical-alkene cyclization (functionalized
alkene): Boger, D. L.; Yun, W.; Teegarden, B. R. J . Org. Chem. Soc.
1992, 57, 2873. (c) Boger, D. L.; Mckie, J . A. J . Org. Chem. 1995, 60,
1271.
(16) Seco-(+)-duocarmycin SA exhibits cytotoxic activity as well as
a DNA alkylating selectivity and efficiency identical with those of (+)-
duocarmycin SA: see ref 8a.
(17) ent-(-)-Duocarmycin SA alkylates DNA but only at concentra-
tions approximately 10× that required for (+)-duocarmycin SA.
Consequently, the natural enantiomer is 10× more effective than the
unnatural enantiomer: see ref 5e.
(21) (a) Aneja, R.; Mukerjee, S. K.; Seshadri, T. R. Tetrahedron 1958,
2, 203. (b) Sahm, W.; Schinzel, E.; J urges, P. J ustus Liebigs Ann. Chem.
1974, 523. (c) Gammill, R. B.; Nash, S. A. J . Org. Chem. 1986, 51,
3116.

8870 J . Org. Chem., Vol. 62, No. 25, 1997
Patel et al.
Sch em e 2
Sch em e 3
of the nitro group in 8, using H2 over 5% Pd-Al2O3 in a
solvent mixture of THF:MeOH, followed by protection of
the resulting amine 9, led to Boc aniline 10. O-Benzyla-
tion of phenol 10, effected under standard Mitsunobu23
conditions, then gave fully protected benzofuran 11. A
low-temperature, electrophilic aromatic bromination of
11, using NBS in the presence of catalytic concentrated
H2SO4,24 led to regiospecific introduction of the C-4
bromide to provide 12 as the sole product in an excellent
yield. Deprotonation of amide 12, using NaH, followed
by alkylation24 of the resulting anion with commercially
available (E:Z)-1,3-dichloropropene gave an inconsequen-
tial mixture of E:Z isomers of vinyl chloride 13, the
desired precursor for the key aryl radical cyclization
(Scheme 3). A deoxygenated solution of aryl bromide 13
in 0.015 M benzene was heated at reflux for 3.5 h in the
presence of 1.1 equiv of tri-n-butyltin hydride and cat.
AIBN to give the desired, fully functionalized alkylating
subunit 14 in 72% yield.25 Particularly noteworthy was
the simple workup employed that involved trituation in
hexanes followed by filtration to provide pure 14 as a
white solid with no detectable levels of tin residues. The
cyclized product 14 is formed in a highly chemoselective
manner believed to proceed via an initial, preferential
homolysis of the weaker aryl C-Br bond26 in 13 to
generate an aryl radical which undergoes a preferred
5-exo-trig intramolecular cyclization27 onto the proximal
vinyl chloride acceptor. A hydrogen radical quench of the
resulting (chloromethyl)indoline radical terminates the
chain reaction. The absence of a methylindoline product
that could originate from reduction of 14 indicates that
the chloromethyl functionality was, indeed, stable to
homolysis under these reaction conditions.
Resolution,28 on a preparatively useful scale was
readily accomplished by direct chromatographic separa-
tion of racemate 14 on a preparative Chiralpak AD
(Amylose)29 column (8 cm × 24 cm, n-PrOH:hexane (51:
49), column volume (CV) ) 964 mL, 225 mL/min flow
rate), to provide multigram quantities (360 mg in 300
mL eluent/injection) of enantiomers (+)-14a (tR ) 5.6 min,
97.6 % ee, 49 % recovery) and ent-(-)-14b (tR ) 7.4 min,
95.55 % ee, 48 % recovery). Acid-mediated deprotection
(27) (a) Baldwin, J . E. J . Chem. Soc., Chem. Commun. 1976, 734.
(b) Beckwith, A. J . L.; Schiesser, C. H. Tetrahedron 1985, 41, 3945.
(28) The amine, derived from deprotection of 14, proved unstable
and was unamenable to resolution through classical fractional recrys-
tallization of diastereomeric salts. Covalent diastereomers, prepared
by attachment of chiral agents to either the amine or phenol derivatives
of 14, were not readily separable by chromatograhic means.
(29) Effective separations on related agents have been previously
achieved by direct chromatographic resolution on a ChiraCel OD
column: (a) Boger, D. L.; Yu, W. J . Am. Chem. Soc. 1994, 116, 7996.
(b) Boger, D. L.; Bollinger, B.; J ohnson, D. S. Bioorg. Med. Chem. Lett.
1996, 6, 2207. (c) Boger, D. L. et al, J . Am. Chem. Soc. 1997, 119, 4977.
See also ref 8a.
(22) Rene L.; Buisson J -P.; Roger, R. Bull. Soc. Chim. Fr. 1975,
11-12, 2763.
(23) Mitsunobu, O. Synthesis 1981, 1.
(24) Boger, D. L.; Coleman, R. S. J . Am. Chem. Soc. 1988, 110, 1321.
(25) A minor uncyclized product (<2%), resulting from reduction of
the aryl bromide, was detected by 1H NMR but not isolated.
(26) An example of regiospecific homolysis of a C-Br bond in the
presence of a C-Cl bond has been reported: Stork, G.; Mook, R. J .
Am. Chem. Soc. 1983, 105, 3721.

Synthesis of Seco (+)- and ent-(-)-Oxaduocarmycin SA
J . Org. Chem., Vol. 62, No. 25, 1997 8871
Ta ble 1. In Vitr o Cytotoxicity, IC50 (n M)
J ) 2.57 Hz, 1H), 5.06 (s, 2H), 4.14 (q, J ) 6.99 Hz, 2H), 4.00
(s, 3H), 1.19 (t, J ) 6.99 Hz, 3H); 13C NMR (CDCl3, 63 MHz)
δ 188.4 (CH), 168.6 (C), 154.4 (C), 152.0 (C), 129.3 (C), 125.9
(C), 115.4 (CH), 111.6 (CH), 69.2 (CH2), 61.6 (CH2), 56.7 (CH3),
14.1 (CH3); IR (KBr) υmax 3100, 3080, 3000, 2950, 2910, 1755,
1698, 1585, 1526, 1479, 1352, 1203, 1053, 744 cm-1; UV
(EtOH) λmax 209 (ꢀ ) 10739), 245 (ꢀ ) 12353), 294.5 (ꢀ ) 5178)
nm; FDMS m/z 284 (M+, 100). Anal. Calcd for C12H13NO7:
C, 50.89; H, 4.63; N, 4.95. Found: C, 50.82; H, 4.50; N, 4.76.
tumor cell line
T222
CCRF-CEM
GC3/C1
(+)-3a
0.39
19.4
0.23
1.59
0.10
0.70
ent-(-)-3b
of (+)-14a and ent-(-)-14b selectively removed the Boc
group to give unstable amine salts (+)-15a and ent-(-)-
15b, which were directly coupled with 5,6,7-trimethoxy-
indole-2-carboxylic acid6a 16, under standard EDCI-
promoted coupling conditions,30 to furnish enantiomerically
pure penultimate intermediates (+)-17a (96 % ee) and
ent-(-)-17b (94 % ee), respectively, in overall 50% yield.
Final, biphasic, transfer catalytic hydrogenolysis31 of 17a
and 17b served to quantitatively remove the benzyl ether
Meth yl 7-Meth oxy-5-n itr oben zofu r an -2-car boxylate (6).
1,8-Diazabicyclo[5.4.0]undec-7-ene (80 mL, 0.535 mol) was
added to a solution of benzaldehyde 5 (136.4 g, 0.482 mol) in
ethanol (2.5 L) and the solution heated at reflux for 4 h. The
reaction mixture was cooled to room temperature and the
precipitated solid filtered, washed with cold ethanol (600 mL),
and dried (in vacuo at 40 °C) to provide benzofuran 6 (56.1 g,
1
44%) as a yellow solid: mp 161-162 °C; H NMR (DMSO-d6,
250 MHz) δ 8.38 (d, J ) 1.84 Hz, 1H), 7.93 (s, 1H), 7.90 (d, J
) 2.21 Hz, 1H), 4.40 (q, J ) 6.99 Hz, 2H), 4.10 (s, 3H), 1.36 (t,
J ) 6.99 Hz, 3H); 13C NMR (CDCl3, 63 MHz) δ 158.3 (C), 148.4
(C), 147.8 (C), 145.8 (C), 145.3 (C), 127.7 (C), 114.2 (CH), 111.3
(CH), 103.8 (CH), 61.9 (CH2), 56.5 (CH3), 14.2 (CH3); IR (KBr)
υmax 3500, 3200, 3000, 2950, 1718, 1529, 1345, 1322, 1297,
1188, 1096, 979, 944, 882, 739 cm-1; UV (EtOH) λmax 213 (ꢀ )
14538), 264.5 (ꢀ ) 28797) nm; FDMS m/z 265 (M+, 100). Anal.
Calcd for C12H11NO6: C, 54.34; H, 4.18; N, 5.28. Found: C,
54.08; H, 4.10; N, 5.29.
and provided (+)-3a ([R]23 -1.73° (c 0.11, DMF))32 and
D
its enantiomer (-)-3b ([R]23 -2.1° (c 0.12, DMF)),
D
respectively.
The biological activity, determined in an in vitro
cytotoxicity assay,33 showed that duocarmycin SA ana-
logue, (+)-3a (GC3/C1; IC50 ) 100 pM) was highly potent
and in the same range as that of the natural product
(+)-1 (L1210; IC50 ) 10 pM). Furthermore, the natural
enantiomer (+)-3a displayed 7-50-fold higher potentcy
than the unnatural isomer, ent-(-)-3b (Table 1), consis-
tent with the biological findings of duocarmycin SA ((+)-
1) and its enantiomer.8a These preliminary results are
sufficiently encouraging to warrant further studies to
fully characterize the biological properties of these agents
and provided the incentive to develop the refined syn-
thetic approach detailed herein.
7-Hyd r oxy-5-n itr oben zofu r a n -2-ca r boxylic Acid (7).
Methoxybenzofuran 6 (55.9 g, 0.211 mol) was combined with
pyridine hydrochloride (243 g, 2.11 mol) in a flask equipped
with a paddle stirrer. The two solids were heated neat at 170
°C with vigorous stirring for 18 h. The mixture was allowed
to cool to ∼50 °C and then poured into ice-water (2 L). The
precipitated solid was filtered and washed with water (1 L).
The crude solid was dissolved in 1 N sodium hydroxide (1.5
L) and washed with ethyl acetate (2 × 500 mL). Concentrated
hydrochloric acid was added to the aqueous solution until pH
3 was reached. The precipitated product was filtered, washed
with water, and dried (in vacuo at 50 °C) to give phenol 7 (44
g, 88%) as a pale yellow solid: mp 189-191 °C; 1H NMR
(DMSO-d6, 300 MHz) δ 13.93 (s, 1H), 11.41 (s, 1H), 8.21 (d, J
) 2.21 Hz, 1H), 7.81 (s, 1H), 7.71 (d, J ) 2.21 Hz, 1H); IR
(KBr) υmax 3274 (br), 3093, 2542, 1694, 1530, 1347, 1287, 1225,
1193, 791 cm-1; UV (EtOH) λmax 208 (ꢀ ) 15442), 263.5 (ꢀ )
18538) nm; FDMS m/z 223 (M+, 100). Anal. Calcd for
C9H5NO6: C, 48.44; H, 2.26; N, 6.28. Found: C, 48.74; H, 2.41;
N, 6.28.
Con clu sion
A concise, total synthesis of seco-(+)- and ent-(-)-
oxaduocarmycins has been achieved in 13 synthetic steps.
Central to this strategy was an aryl radical cyclization
onto a vinyl chloride, followed by a preparatively useful
resolution procedure. The tactical improvement defined
in this work shortens approaches to related agents20 by
2-4 synthetic steps. This methodology should find
application as a general synthetic approach to other
members of this class of agents and related analogues.
Meth yl 7-Hydr oxy-5-n itr oben zofu r an -2-car boxylate (8).
Acetyl chloride (13 mL, 186 mmol) was added dropwise to dry
methanol (1.5 L) at 0 °C and the solution stirred at room
temperature for 0.5 h. Carboxylic acid 7 (39.6 g, 177 mmol)
was added to the acidic methanol and the mixture heated at
reflux, under a nitrogen atmosphere, for 18 h. The reaction
was cooled to room temperature and the resulting precipitate
filtered, washed with cold methanol (500 mL), and dried to
give methyl ester 8 (41.2 g, 98%) as a light tan solid: mp 151-
Exp er im en ta l Section
Eth yl 2-(6-F or m yl-2-m eth oxy-4-n itr op h en oxy)a ceta te
(5). To a rapidly stirring solution of 2-hydroxy-3-methoxy-5-
nitrobenzaldehyde 4 (100 g, 0.507 mol) in dry methanol (1.5
L) was added powdered potassium hydroxide (85%, 37.1 g,
0.562 mol) and the mixture heated at reflux for 45 min.
Methanol solvent was removed in vacuo and the resulting solid
residue suspended in dimethylformamide (1.5 L). The mixture
was cooled to 0 °C and ethyl bromoacetate (103 mL, 0.925 mol)
added. The solution was warmed to room temperature and
stirred for 36 h. The solvent was removed in vacuo and the
resulting solid residue was diluted with water (3 L) and
extracted with ethyl acetate (8 × 1 L). The combined organic
extracts were washed with water (1 L) and brine (1 L), dried
(Na2SO4), and concentrated to give the desired product 5 (136.5
1
152 °C; H NMR (DMSO-d6, 300 MHz) δ 11.48 (s, 1H), 8.23
(d, J ) 2.21 Hz, 1H), 7.91 (s, 1H), 7.73 (s, 1H), 3.93 (s, 3H); IR
(KBr) υmax 3281, 1688, 1582, 1523, 1442, 1356, 1328, 1256,
1102, 969, 901, 810, 744 cm-1; UV (EtOH) λmax 213 (ꢀ ) 16556),
267.5 (ꢀ ) 26855) nm; FDMS m/z 238 (M+, 100). Anal. Calcd
for C10H7NO6: C, 50.64; H, 2.97; N, 5.91. Found: C, 50.85;
H, 3.01; N, 5.88.
Meth yl 5-Am in o-7-h yd r oxyben zofu r a n -2-ca r boxyla te
(9). To a solution of nitrobenzofuran 8 (42 g, 177 mmol) in
tetrahydrofuran:methanol (1:1, 1.5 L) was added 5% pal-
ladium-Al2O3 (5 g) and the mixture stirred at room temper-
ature under 60 psi hydrogen atmosphere for 4 h. The reaction
mixture was filtered through Celite and the filtrate concen-
trated in vacuo to give a solid, which was purified by flash
chromatography (gradient: 0-5% methanol:dichloromethane)
to provide aminobenzofuran 9 (35.6 g, 97%) as a tan solid: mp
201-202 °C; 1H NMR (DMSO-d6, 250 MHz) δ 9.93 (s, 1H), 7.44
(s, 1H), 6.29 (d, J ) 2 Hz, 1H), 6.24 (d, J ) 1.75 Hz, 1H), 4.92
(s, 2H), 3.85 (s, 3H); IR (KBr) υmax 3416, 3335, 2947, 1713,
1
g, 95%) as a tan solid: mp 109-110 °C; H NMR (DMSO-d6,
300 MHz) δ 10.48 (s, 1H), 8.09 (d, J ) 2.94 Hz, 1H), 8.06 (d,
(30) Boger, D. L.; Machiya, K. J . Am. Chem. Soc. 1992, 114, 10056.
(31) (a) Ram, S.; Ehrenkaufer, R. E. Synthesis 1988, 91. Bieg, T.;
Szeja, W. Synthesis 1985, 76.
(32) The absolute configuration at C1 in analogue (+)-3a was
assigned based on analogy of the biological activity with (+)-DSA (1).
(33) Procedures are described in the experimental section. For T222
tumor cell line also see: ref 15; for CCRF-CEM and GC3/C1 tumor
cell lines also see: Mosmann, T. J . Immunol. Meth. 1983, 65, 55-62.

8872 J . Org. Chem., Vol. 62, No. 25, 1997
Patel et al.
1610, 1568, 1454, 1434, 1356, 1204, 1155 cm-1; UV (EtOH)
λmax 235.5 (ꢀ ) 20635), 290.5 (ꢀ ) 15655) nm; FDMS m/z 207
(M+, 100). Anal. Calcd for C10H9NO4 requires: C, 57.97; H,
4.38; N, 6.76%. Found: C, 57.72; H, 4.28; N, 6.63%.
yellow mixture was stirred at room temperature for 40 min
and cooled to 0 °C and then neat (E:Z)-1,3-dichloropropene (846
µL, 9 mmol) added. The reaction was allowed to warm to room
temperature, stirred for a further 15 h, quenched with brine
(4 mL), and extracted with ethyl acetate (3 × 150 mL).
Combined, dried (MgSO4) organics were concentrated in vacuo,
and the resulting crude brown oil was purified by flash
chromatography (gradient: 0-20% ethyl acetate/hexanes) to
give vinyl chloride 13 (1.36 g, 82%) as a low melting yellow
glass: (E:Z vinyl chlorides and NBoc rotomers): 1H NMR
(CDCl3, 300 MHz) δ 7.56 (s, 1H), 7.48-7.20 (m, 5H), 6.85-
6.71 (m, 1H), 5.99-5.85 (m, 2H), 5.32 (br s, 2H), 4.46 (dd, J )
15.5 and 6.2 Hz, 1H), 4.35-4.20 (m, 1H), 3.98 (s, 3H), 1.5 (br
s, 3H), 1.28 (br s, 6H);13C NMR (CDCl3, 63 MHz) δ 159.1 (C),
153.8 (C), 146.4 (C), 146.3 (C), 135.8 (C), 135.7 (C), 128.6 (CH),
128.5 (CH), 128.3 (CH), 128.2 (CH), 127.4 (CH), 127.0 (CH),
120.7 (CH), 114.8 (CH), 114.7 (C), 113.3 (CH), 80.6 (C), 71.4
(CH2), 71.3 (CH2), 52.4 (CH3), 28.0 (CH3); IR (CHCl3) υmax 2981,
1729,1 698, 1368, 1165 cm-1; UV (EtOH) λmax 203 (ꢀ ) 32586),
245.5 (ꢀ ) 30885), 286.5 (ꢀ ) 17684) nm; FDMS m/z 551 (M+,
100). Anal. Calcd for C25H25NO6Cl Br: C, 54.51; H, 4.57; N,
2.58; Cl, 6.44; Br, 14.51; Found: C, 54.31; H, 4.60; N, 2.68;
Cl, 6.72; Br, 14.42.
Meth yl 5-(N-(ter t-Bu tyloxycar bon yl)am in o)-7-h ydr oxy-
ben zofu r a n -2-ca r boxyla te (10). To a solution of amine 9
(35.6 g, 172 mmol) in tetrahydrofuran (750 mL) was added
di-tert-butyl dicarbonate (48 g, 222 mmol) and the mixture
stirred at room temperature for 24 h. The solvent was
removed in-vacuo and the residue dissolved in ethyl acetate
(1.5 L), washed with 1 N aqueous hydrochloric acid (2 × 200
mL), water (2 × 600 mL), and brine (1 × 600 mL), and then
dried (Na2SO4). The organic solution was passed through a
pad of silica gel and concentrated in vacuo to furnish Boc
amine 10 (49.4 g, 93%) as a white powder: mp 190-192 °C;
1H NMR (DMSO-d6, 300 MHz) δ 10.34 (s, 1H), 9.34 (s, 1H),
7.65 (s, 1H), 7.37 (d, J ) 1.47 Hz, 1H), 7.10 (d, J ) 1.84 Hz,
1H), 3.88 (s, 3H), 1.48 (s, 9H); IR (KBr) υmax 3291, 1698, 1606,
1584, 1534, 1453, 1340, 1249, 1164, 1109, 1062, 976, 852, 763,
739 cm-1; UV (EtOH) λmax 211 (ꢀ ) 25983), 244.5 (ꢀ ) 31990),
287.5 (ꢀ ) 19236) nm; FDMS m/z 308 (M+, 100). Anal. Calcd
for C15H17NO6: C, 58.63; H, 5.58; N, 4.56. Found: C, 58.88;
H, 5.63; N, 4.60.
Meth yl 5-(N-(ter t-Bu tyloxyca r bon yl)a m in o)-7-(ben zyl-
oxy)ben zofu r a n -2-ca r boxyla te (11). To a solution of phenol
10 (48.9 g, 159 mmol) in dichloromethane (1.5 L) were added
benzyl alcohol (30 mL, 291 mmol) and triphenylphosphine
(75.1 g, 286 mmol) at room temperture and under a dry
nitrogen atmosphere. The solution was cooled to 0 °C and
diethyl azodicarboxylate (DEAD) (50 g, 287 mmol) added
dropwise. The reaction mixture was stirred at room temper-
ature for 18 h and concentrated in vacuo and the solid residue
purified by flash chromatoghaphy (gradient: 5-20% ethyl
acetate/hexanes) to provide benzyl ether 11 (57 g, 90%) as a
Meth yl (1R/S)-5-(Ben zyloxy)-3-(ter t-bu tyloxycar bon yl)-
1-(ch lor om et h yl)-1,2-d ih yd r o-3H -fu r a n o[3,2-e]in d ole-7-
ca r boxyla te (14). To a solution of aryl bromide 13 (12.30 g,
22.34 mmol) in dry benzene (1.49 L) were added tri-n-butyltin
hydride (6.52 mL, 24.57 mmol) and catalytic 2,2′-azobis(isobu-
tyronitrile) (AIBN) (183 mg, 1.12 mmol). The solution was
deoxygenated by bubbling dry nitrogen through the solution
at room temperature for ∼1 h. The reaction mixture was then
heated at reflux for 3.5 h and concentrated in vacuo to give
an oily residue. Trituation of the crude oil with hexanes
provided a solid which was filtered and washed with hexanes
(3 × 500 mL) to give indoline 14 (7.79 g, 72%) as an off-white
solid: mp 137-139 °C; 1H NMR (CDCl3, 300 MHz) δ 7.85 (br
s, 1H), 7.54-7.30 (m, 6H), 5.28 (s, 2H), 4.17 (apparent t, J )
10.4 Hz, 1H), 4.08-3.98 (m, 1H), 3.95 (s, 3H), 3.91-3.71 (m,
2H), 3.56 (apparent t, J ) 10.3 Hz,1H), 1.56 (s, 9H); 13C NMR
(CDCl3, 63 MHz) δ 159.4 (C), 152.2 (C), 146.5 (C), 144.9 (C),
142.3 (C), 136 1 (C), 128.4 (CH), 128.0 (CH), 127.7 (CH), 111.6
(CH), 100.1 (C), 70.9 (CH2), 52.9 (CH2), 52.2 (CH3), 46.6 (CH3),
41.5 (C), 28.4 (3 × CH3); IR (CHCl3) υmax 2956, 1725, 1695,
1497, 1158 cm-1; UV (EtOH) λmax 219 (ꢀ ) 27621), 254.5 (ꢀ )
38111), 285 (ꢀ ) 14659), 346 (ꢀ ) 2729) nm; FDMS m/z 471
(M+, 100). Anal. Calcd for C25H26NO6Cl; C, 63.63; H, 5.55;
N, 2.97; Cl, 7.51; Found: C, 63.35; H, 5.51; N, 2.74; Cl, 7.30.
1
white solid: mp 141-143 °C; H NMR (DMSO-d6, 250 MHz)
δ 9.43 (s, 1H), 7.71 (s, 1H), 7.55 (s, 2H), 7.53 (d, J ) 1.75 Hz,
1H), 7.51-7.39 (m, 3H), 7.29 (d, J ) 1.75 Hz, 1H), 5.22 (s,
2H), 3.87 (s, 3H), 1.48 (s, 9H); 13C NMR (CDCl3, 63 MHz) δ
159.6 (C), 152.9 (C), 146.0 (C), 144.6 (C), 142.1 (C), 136.1 (C),
135.2 (C), 128.6 (C), 128.5 (CH), 128.0 (CH), 127.6 (CH), 114.2
(CH), 104.3 (CH), 103.7 (CH), 80.5 (C), 71.0 (CH2), 52.1 (CH3),
28.3 (3 x CH3); IR (KBr) υmax 3440, 2982, 2954, 2934, 1725,
1606, 1582, 1528, 1320, 1158 cm-1; UV (EtOH) λmax 242 (ꢀ )
29172), 280 (ꢀ ) 16141) nm; FDMS m/z 397 (M+, 100). Anal.
Calcd for C22H23NO6: C, 66.48; H, 5.83; N, 3.52. Found: C,
66.25; H, 5.86; N, 3.69.
Meth yl 5-(N-(ter t-Bu tyloxyca r bon yl)a m in o)-7-(ben zyl-
oxy)-4-br om oben zofu r a n -2-ca r boxyla te (12). A solution
of benzofuran 11 (30 g, 75.5 mmol) in dry tetrahydrofuran (250
mL) was cooled to -60 °C and two drops of concentrated
sulfuric acid, followed by N-bromosuccinamide (14.8 g, 83
mmol), added under a stream of dry nitrogen. The mixture
was warmed to 0 °C over a period of 3 h and was then
quenched with 0.1 N aqueous sodium bisulfite (5 mL). The
organic solvent was removed in vacuo and the residue diluted
with ethyl acetate (300 mL), washed with water (2 × 150 mL),
and brine (2 × 150 mL), dried (Na2SO4), and concentrated in
vacuo to afford aryl bromide 12 (34.3 g, 95%) as a light yellow
Resolu tion of 14. Twenty micron Chiralpak AD (Amylose)
(600 g) was slurried in 1-propanol (1200 mL) and transferred
to a 8 cm Prochrom column and the bed compressed at a
pressure of 85 Bar resulting in a 8 cm × 24 cm configuration
with a column volume (CV) of 964 mL. Preparative separa-
tions were preformed at a flow rate of 225 mL/min with a
column pressure of 11 Bar. Racemate 14 (360 mg) was
dissolved in eluent 1-propanol/hexanes (51:49, 300 mL) and
charged on the column using a sample pump. Chromatograms
from a run on the Prochrom column is illustrated in Figure 1.
Racemate 14 (10.57 g) was resolved, in 35 runs on the 8 cm
column, with an optimized time of 13 min per run and effluent
monitored at 280 nm, to give:
1
solid: mp 140-142 °C; H NMR (DMSO-d6, 300 MHz) δ 8.07
(s, 1H), 7.54-7.49 (m, 3H), 7.42-7.34 (m, 3H), 6.94 (s, 1H),
5.31 (s, 2H), 3.97 (s, 3H), 1.55 (s, 9H); 13C NMR (CDCl3, 63
MHz) δ 159.2 (C), 152.5 (C), 146.0 (C), 144.2 (C), 141.2
(C),135.8 (C),133.2 (C), 129.1 (C), 128.4 (CH), 128.2 (CH), 128.0
(CH), 114.0 (CH), 104.6 (CH), 94.2 (C), 81.1 (C), 71.2 (CH2),
52.3 (CH3), 28.2 (3 × CH3); IR (KBr) υmax 3700, 3440, 3000,
2983, 17231453, 1233, 1157 cm-1; UV (EtOH) λmax 212.5 (ꢀ )
27223), 246 (ꢀ ) 28581), 287 (ꢀ ) 14281) nm; FDMS m/z 475
(M+, 100). Anal. Calcd for C22H22NO6Br: C, 55.48; H, 4.65;
N, 2.94; Br, 16.78%. Found: C, 55.20; H, 4.70; N, 3.20; Br,
16.50.
Meth yl (1S)-5-(Ben zyloxy)-3-(ter t-bu tyloxyca r bon yl)-
1-(ch lor om et h yl)-1,2-d ih yd r o-3H -fu r a n o[3,2-e]in d ole-7-
ca r boxyla te (14a ). First peak (tR ) 5.6 min, 97.6 % ee, 5.16
g, 49 % recovery), as a white solid: [R]23D -10.8 ° (c 0.5, CHCl3).
1
Mp 130-132 °C dec; H NMR (CDCl3, 300 MHz) δ 7.90 (br s,
1H), 7.55-7.33 (m, 6H), 5.29 (s, 2H), 4.18 (apparent dd, J )
11.3 and 10 Hz, 1H), 4.08-3.93 (m, 1H), 3.96 (s, 3H), 3.87-
3.80 (m, 2Hl), 3.60 (apparent t, J ) 10.4 Hz, 1H), 1.57 (s, 9H);
13C NMR (CDCl3, 63 MHz) δ 159.4 (C), 152.2 (C), 146.5 (C),
144.9 (C), 142.3 (C), 136 1 (C), 128.4 (CH), 128.0 (CH), 127.7
(CH), 111.6 (CH), 100.1 (C), 70.9 (CH2), 52.9 (CH2), 52.2 (CH3),
46.6 (CH3), 41.5 (C), 28.4 (3 × CH3); IR (KBr) υmax 2990, 1737,
1696, 1152, 1137, 767, 696 cm-1; UV (EtOH) λmax 219 (ꢀ )
27870), 254.5 (ꢀ ) 39132), 284.5 (ꢀ ) 14997), 346 (ꢀ ) 2901)
nm; FDMS m/z 470.9 (M+, 100). Anal. Calcd for C25H26NO6-
Met h yl 5-[N-(ter t-Bu t yloxyca r b on yl)-N-(3-ch lor o-2-
p r op en -1-yl)a m in o]-7-(ben zyloxy)-4-br om oben zofu r a n -2-
ca r boxyla te (13). Boc amine 12 (1.43 g, 3 mmol) was added
portionwise to a suspension of sodium hydride (60% in mineral
oil; 156 mg, 3.90 mmol) in dry dimethylformamide (8.0 mL)
at room temperature and under nitrogen. The resulting dark

Synthesis of Seco (+)- and ent-(-)-Oxaduocarmycin SA
J . Org. Chem., Vol. 62, No. 25, 1997 8873
16 (128 mg, 0.509 mmol) and finally 1-(3-(dimethylamino)pro-
pyl)-3-ethylcarbodiimide hydrochloride (EDCI) (244 mg, 1.272
mmol) and the mixture stirred at room temperature under
nitrogen for 18 h. The solvent was reduced to ∼1/2 the original
volume and ethyl acetate (20 mL) added. The organics were
washed with 0.1 N aqueous hydrochloric acid (2 × 10 mL),
water (10 mL), saturated sodium bicarbonate (10 mL), and
brine (10 mL) and then dried (MgSO4). Concentration of the
organics followed by purification of the resulting crude solid
by flash chromatography (gradient: 10-40% ethyl acetate/
hexanes) provided amide 17a (127 mg, 50%, HPLC (analytical
ChiraCel OD column; flow rate ) 1.0 mL/min; UV detection
at λ ) 254 nm; eluent, CH3CN:H2O 90:10): tR ) 11.08 min; 96
% ee) as a white powder: [R]23 +13.6° (c 0.5, CHCl3). mp
D
182-183.5 °C (EtOAc/hexanes, white needles); 1H NMR
(CDCl3, 300 MHz) δ 9.38 (brs, 1H), 8.31 (s, 1H), 7.56-7.52 (m,
3H), 7.43-7.28 (m, 3H), 6.96 (d, J ) 2.1 Hz, 1H), 6.87 (s, 1H),
5.35 (s, 2H), 4.71 (apparent t, J ) 10 Hz, 1H), 4.59 (dd, J )
10.9 and 4.0 Hz, 1H), 4.08 (s, 3H), 4.07-3.88 (m, 2H), 3.98 (s,
3H), 3.95 (s, 3H), 3.92 (s, 3H), 3.61 (dd, J ) 11 and 9.4 Hz,
1H); 13C NMR (CDCl3, 63 MHz) δ 159.9 (C), 159.3 (C), 150.1
(C), 146.7 (C), 144.7 (C), 143.1 (C), 141.0 (C), 140.5 (C), 138.8
(C), 136.0 (C), 129.6 (C), 128.5 (CH), 128.1 (CH), 127.7 (CH),
125.4 (C), 123.9 (C), 123.5 (C), 114.7 (C), 111.5 (CH), 106.4
(CH), 102.4 (CH), 97.6 (CH), 70.9 (CH2), 61.4 (CH3), 61.0 (CH3),
56.2 (CH3), 55.0 (CH2), 52.3 (CH3), 46.3 (CH2), 43.0 (CH); IR
(KBr) υmax 3459, 2936, 1732, 1622, 1494, 1308, 1203, 744, 697
cm-1; UV (EtOH) λmax 208.5 (ꢀ ) 35611), 297 (ꢀ ) 22216), 326
(ꢀ ) 24475) nm; FDMS m/z 604 (M+, 100). Anal. Calcd for
C32H29N2O8Cl: C, 63.52; H, 4.83; N, 4.63; Cl, 5.86. Found: C,
63.58; H, 4.87; N, 4.39; Cl, 6.02.
F igu r e 1. Typical chromatograms of the separation of race-
mate 14 on a Chiralpak AD (Amylose) column.
Cl: C, 63.63; H, 5.55; N, 2.97; Cl, 7.51; Found: C, 63.56; H,
5.53; N, 2.67; Cl, 7.72.
Meth yl (1R)-5-(Ben zyloxy)-3-(ter t-bu tyloxyca r bon yl)-
1-(ch lor om et h yl)-1,2-d ih yd r o-3H -fu r a n o[3,2-e]in d ole-7-
ca r boxyla te (14b). Second peak (tR ) 7.4 min, 95.5 % ee,
5.04 g, 48 % recovery) as a white solid: [R]23 +11.2° (c 0.5,
D
CHCl3). mp 131-133 °C dec; 1H NMR (CDCl3, 300 MHz) δ 7.90
(br s, 1H), 7.55-7.28 (m, 6H), 5.29 (s, 2H), 4.18 (dd, J ) 11.6
and 10 Hz,1H), 4.08-3.98 (m, 1H), 3.96 (s, 3H), 3.88-3.80 (m,
2Hl), 3.60 (apparent t, J ) 10.4 Hz, 1H), 1.57 (s, 9H); 13C NMR
(CDCl3, 63 MHz) δ 159.4 (C), 152.2 (C), 146.5 (C), 144.9 (C),
142.3 (C), 136 1 (C), 128.4 (CH), 128.0 (CH), 127.7 (CH), 111.6
(CH), 100.1 (C), 70.9 (CH2), 52.9 (CH2), 52.2 (CH3), 46.6 (CH3),
41.5 (C), 28.4 (3 × CH3); IR (KBr) υmax 2990, 1736, 1697, 1499,
1424, 1349, 1208, 1152, 1137, 762, 697 cm-1; UV (EtOH) λmax
219.5 (ꢀ ) 26377), 254.5 (ꢀ ) 37628), 284.5 (ꢀ ) 14440), 346.5
(ꢀ ) 2823) nm; FDMS m/z 470.9 (M+, 100). Anal. Calcd for
C25H26NO6Cl: C, 63.63; H, 5.55; N, 2.97; Cl, 7.51; Found: C,
63.91; H, 5.58; N, 3.13; Cl, 7.59.
(-)-(1R)-Meth yl 5-(Ben zyloxy)-1-(ch lor om eth yl)-1,2-d i-
h yd r o-3-[(5,6,7-t r im et h oxy-1H -in d ol-2-yl)ca r b on yl]-3H -
fu r a n o[3,2-e]in d ole-7-ca r boxyla te (17b). Amine 15b (216
mg, 0.530 mmol) was coupled to 5,6,7-trimethoxyindolecar-
boxylic acid 16, in the same manner described above, to furnish
amide 17b (161 mg, 50%, HPLC (analytical ChiraCel OD
column; flow rate ) 1.0 mL/min; UV detection at λ ) 254 nm;
eluent, CH3CN:H2O 90:10): 17b:: tR ) 8.42 min; 94 % ee) as
a white solid: [R]23 -13.2° (c 0.5, CHCl3); mp 138-140 °C;
Meth yl (1S)-5-(Ben zyloxy)-1-(ch lor om eth yl)-1,2-d ih y-
d r o-3H-fu r a n o[3,2-e]in d ole-7-ca r boxyla te (15a ). Boc in-
doline 14a (800 mg, 1.697 mmol) was added to a dry solution
of 4 N HCl in dioxane (25 mL) at 0 °C and under nitrogen.
The solution was stirred at 0 °C for 8 h and the solvent
removed at 0 °C under high vacuum to provide amine
hydrochloride salt 15a (690mg, 100%) as an unstable brown
solid: 1H NMR (DMSO-d6, 300 MHz) δ 8.07 (s, 1H), 7.55-
7.52 (m, 2H), 7.47-7.39 (m, 3H), 7.13 (s, 1H), 5.32 (s, 2H),
4.14-3.91 (m, 2H), 3.91 (s, 3H), 3.73-3.60 (m, 2H), 3.51-3.47
(m, 1H); IR (KBr) υmax 3400 (br), 1717, 1584, 1333, 1317, 1207,
1141, 1104, 915, 727 cm-1; UV (EtOH) λmax 243 (ꢀ ) 18536),
285 (ꢀ ) 12985), 359 (ꢀ ) 2272), 389 (ꢀ ) 1674) nm; FDMS
m/z 371 (Free amine, M+, 100). Anal. Calcd for C20H19NO4Cl2:
C, 58.84; H, 4.69; N, 3.43; Cl, 17.37. Found: C, 58.71; H, 4.85;
N, 3.13; Cl, 16.82.
D
1H NMR (CDCl3, 300 MHz) δ 9.41 (brs, 1H), 8.31 (s, 1H), 7.55-
7.51 (m, 3H), 7.43-7.33 (m, 3H), 6.95 (d, J ) 2.1 Hz, 1H), 6.86
(s, 1H), 5.35 (s, 2H), 4.70 (apparent t, J ) 9.9 Hz, 1H), 4.58
(dd, J ) 10.9 and 3.9 Hz, 1H), 4.08 (s, 3H), 4.04-3.88 (m, 2H),
3.98 (s, 3H), 3.94 (s, 3H), 3.91 (s, 3H), 3.60 (dd, J ) 10.8 and
9.6 Hz, 1H); 13C NMR (CDCl3, 63 MHz) δ 159.9 (C), 159.3 (C),
150.1 (C), 146.7 (C), 144.7 (C), 143.1 (C), 141.0 (C), 140.5 (C),
138.8 (C), 136.0 (C), 129.6 (C), 128.5 (CH), 128.1 (CH), 127.7
(CH), 125.4 (C), 123.9 (C), 123.5 (C), 114.7 (C), 111.5 (CH),
106.4 (CH), 102.4 (CH), 97.6 (CH), 70.9 (CH2), 61.4 (CH3), 61.0
(CH3), 56.2 (CH3), 55.0 (CH2), 52.3 (CH3), 46.3 (CH2), 43.0 (CH);
IR (KBr) υmax 2937, 1731, 1622, 1494, 1416, 1309, 1204, 1109,
744, 697 cm-1; UV (EtOH) λmax 209 (ꢀ ) 44076), 297.5 (ꢀ )
26058), 326 (ꢀ ) 30391) nm; FDMS m/z 603.7 (M+, 100). Anal.
Calcd for C32H29N2O8Cl: C, 63.53; H, 4.83; N, 4.63; Cl, 5.86.
Found: C, 63.74; H, 4.81; N, 4.40; Cl, 6.01.
Meth yl (1R)-5-(Ben zyloxy)-1-(ch lor om eth yl)-1,2-d ih y-
d r o-3H-fu r a n o[3,2-e]in d ole-7-ca r boxyla te (15b). Boc in-
doline 14b (1.00g, 2.12 mmol) was added to a dry solution of
4 N HCl in dioxane (28 mL) at 0 °C and under nitrogen. The
solution was stirred at 0 °C for 8 h and the solvent removed
at 0 °C under high vacuum to provide amine hydrochloride
salt 15a (865mg, 100%) as an unstable brown solid: 1H NMR
(DMSO-d6, 300 MHz) δ 8.08 (s, 1H), 7.55-7.52 (m, 2H), 7.47-
7.39 (m, 3H), 7.15 (s, 1H), 5.32 (s, 2H), 4.14-3.91 (m, 2H), 3.91
(s, 3H), 3.72-3.61 (m, 2H), 3.51-3.48 (m, 1H); IR (KBr) υmax
3400 (br), 1717, 1583, 1333, 1317, 1275, 1207, 1141, 1104, 728
cm-1; UV (EtOH) λmax 211 (ꢀ ) 26721), 243 (ꢀ ) 18796), 285 (ꢀ
) 13166), 359 (ꢀ ) 2189) nm; FDMS m/z 371 (Free amine, M+,
100). Anal. Calcd for C20H19NO4Cl2: C, 58.84; H, 4.69; N,
3.43; Cl, 17.37. Found: C, 58.78; H, 4.72; N, 3.18; (Cl, 16.82).
Meth yl (1R)-5-(Ben zyloxy)-1-(ch lor om eth yl)-1,2-d ih y-
dr o-3-[(5,6,7-tr im eth oxy-1H-in dol-2-yl)car bon yl]-3H-fu r a-
n o[3,2-e]in d ole-7-ca r boxyla te (17a ). To a dry solution of
amine hydrochloride salt 15a (173 mg, 0.424 mmol) in dimeth-
ylformamide (4.0 mL) was added sodium bicarbonate (178 mg,
2.12 mmol) followed by 5,6,7-trimethoxyindolecarboxylic acid
Meth yl (1S)-5-h yd r oxy-1-(ch lor om eth yl)-1,2-d ih yd r o-
3-[(5,6,7-tr im eth oxy-1H-in d ol-2-yl)ca r bon yl]-3H-fu r a n o-
[3,2-e]in d ole-7-ca r boxyla te (3a ). To a solution of benzyl
ether 17a (149 mg, 0.247 mmol) in tetrahydrofuran (5.80 mL)
was added 10% aqueous ammonium formate (560 µL). The
solution was cooled to 0 °C and 10% Pd-C (56 mg) added. The
reaction mixture was stirred for 5 h, filtered through a pad of
Celite, and concentrated to provide (+)-oxaduocarmycin SA
(3a ) (122 mg, 96%) as a yellow powder: [R]23 +1.73° (c 0.11,
D
1
DMF); mp 265-268 °C dec; H NMR (DMSO-d6, 300 MHz) δ
11.39 (d, J ) 1.14 Hz, 1H), 10.52 (s, 1H), 7.96 (s, 1H), 7.92 (s,
1H), 6.97 (d, J ) 1.7 Hz, 1H), 6.92 (s, 1H), 4.66 (apparent t, J
) 10 Hz, 1H), 4.34 (dd, J ) 10.6 and 3.54 Hz, 1H), 4.08-4.02
(m,1H), 3.98-3.94 (m, 1H), 3.88 (s, 3H), 3.87 (s, 3H), 3.77 (s,
3H), 3.75 (s, 3H), 3.34-3.26 (buried m, 1H); 13C NMR (DMSO-
d6, 63 MHz) δ 160.0 (C), 159.1 (C), 149.2 (C), 145.6 (C), 142.6
(C), 141.9 (C), 140.9 (C), 139.9 (C), 139.1 (C), 131.0 (C), 125.3
(C), 124.2 (C), 123.2 (C), 114.3 (C), 113.0 (CH), 106.0 (CH),
104.0 (CH), 98.1 (CH), 61.1 (CH3), 61.0 (CH3), 56.0 (CH3), 54.8
(CH2) 52.3 (CH3), 47.4 (CH2), 41.4 (CH); IR (KBr) υmax 3449,

8874 J . Org. Chem., Vol. 62, No. 25, 1997
Patel et al.
1726, 1589, 1492, 1453, 1310, 1198, 1106 cm-1; UV (EtOH)
λmax 304 (ꢀ ) 8970), 328 (ꢀ ) 9326) nm; FDMS m/z 514 (M+,
100). Anal. Calcd for C25H23N2O8Cl: C, 58.31; H, 4.50; N,
5.44; Cl, 6.89. Found: C, 58.25; H, 4.66; N, 5.29; Cl, 6.61.
Meth yl (1R)-5-Hyd r oxy-1-(ch lor om eth yl)-1,2-d ih yd r o-
3-[(5,6,7-tr im eth oxy-1H-in d ol-2-yl)ca r bon yl]-3H-fu r a n o-
[3,2-e]in d ole-7-ca r boxyla te (3b). Benzyl ether 17b (130 mg,
0.215 mmol) was deprotected, in the same manner described
previously, to give (-)-oxaduocarmycin SA (3b) (109 mg, 99%)
macromolecules was detemined using an automated cell
harvester and liquid scintillation techniques. Data were
evaluated as % reduction in incorporation of radioactivity
relative to controls incubated in medium without compound
to yield a 50% cytotoxic concentration (IC50).
A modification of the original MTT (3-(4,5-dimethylthiazol-
2-yl)-2,5-diphenyltetrazolium bromide) colorimeteric assay,
described by Mosmann33 that measures the reduction of MTT
to a violet-colored formazan by living cells, was used for the
CCRF-CEM and GC3/C1 cell assays. The tumor cells (1 ×
104) were seeded in assay medium (100 µL)/well in 96-well flat
bottom tissue culture plates (Costar, Cambridge, MA). Assay
medium consisted of RPMI-1640 medium supplemented with
10% dialyzed fetal bovine serum. Well 1A was left blank (100
µL of growth medium without cells). Stock solutions of test
compounds were prepared in DMSO at 1 mg/mL, and a series
of 2-fold dilutions were made in Dulbecco’s phosphate-buffered
saline (PBS). Aliquots (10 µL) of each concentration were
added to triplicate wells. The compounds were tested at
concentrations ranging from 0.01 to 0.00008 µg/mL. Plates
were incubated for 72 h at 37 °C in a humidified atmosphere
of 5% CO2-in-air. MTT was dissolved in PBS at 5 mg/mL, and
following incubation of plates, a stock solution of MTT (10 µL)
was added to the assay wells and the plates were incubated
at 37 °C for an additional 2 h. DMSO (100 µL) was added to
each well to thoroughly solubilize the formazan, and the plates
were then read on a Dynatech (Alexandria, VA) MR600 reader
using a test wavelength of 570 nm and a reference wavelength
of 630 nm. The IC50 was determined as the concentration of
drug required to inhibit cell growth by 50%.
as a yellow solid: [R]23 -2.1° (c 0.12, DMF); mp 264-267 °C;
D
1H NMR (DMSO-d6) δ 11.40 (s, 1H), 10.51 (s, 1H), 7.96 (s, 1H),
7.94 (s, 1H), 6.98 (d, J ) 1.7 Hz, 1H), 6.92 (s, 1H), 4.67
(apparent t, J ) 10 Hz, 1H), 4.32 (dd, J ) 11.4 and 4.2 Hz,
1H), 4.08-4.05 (m, 1H), 3.98-3.92 (m, 1H), 3.89 (s, 3H), 3.87
(s, 3H), 3.77 (s, 3H), 3.75 (s, 3H), 3.36-3.27 (buried m, 1H);
13C NMR (DMSO-d6, 63 MHz) δ 160.0 (C), 159.1 (C), 149.2 (C),
145.6 (C), 142.6 (C),141.9 (C), 140.9 (C), 139.9 (C), 139.1 (C),
131.0 (C), 125.3 (C), 124.2 (C), 123.2 (C), 114.3 (C), 113.0 (CH),
106.0 (CH), 104.0 (CH), 98.1 (CH), 61.1 (CH3), 61.0 (CH3), 56.0
(CH3), 54.8 (CH2) 52.3 (CH3), 47.4 (CH2), 41.4 (CH); IR (KBr)
υmax 3450, 1725, 1586, 1492, 1429, 1310, 1107, 746 cm-1; UV
(EtOH-sparingly soluble) λmax 208, 303, 328 nm; FDMS m/z
514 (M+, 100). Anal. Calcd for C25H23N2O8Cl: C, 58.31; H,
4.50; N, 5.44; Cl, 6.89. Found: C, 58.55; H, 4.67; N, 5.14; Cl,
6.92.
In Vitr o Cytotoxicity Assa ys. T222 (human lung epider-
moid carcinoma)14 cells (1 × 104) were distributed in each well
of 96 well tissue culture plates and incubated in leucine-
deficient media (leucine-free DMEM, 13 µg/mL L-leucine, 29.9
µg/mL L-glutamine, 50 µg/mL gentamicin, and 10% dialyzed
fetal bovine serum) for 16 h at 37 °C in 5% carbon dioxide/air
atmosphere. The medium was removed aseptically and com-
pound dilutions added in leucine-deficient medium (200 µL).
After 48 h the media was removed and 4 µCi (3H-Leucine-NEN,
Boston, MA) was added to each well. The plates were returned
to the incubator for 24 h. Radioactivity incorporated into
Ack n ow led gm en t. The authors would like to thank
the Physical Chemistry Department at Lilly Research
Laboratories for their generous support.
J O971880B