Small molecules that mimic the thiol-triggered alkylating properties seen in the natural product leinamycin

Article abstract of DOI:10.1021/ja029169y

Reaction of the antitumor agent leinamycin with cellular thiols results in conversion of the natural product to a DNA-alkylating episulfonium alkylating agent via an intriguing sequence of chemical reactions. To establish whether the chemistry first seen

Full text of DOI:10.1021/ja029169y

Published on Web 04/05/2003
Small Molecules That Mimic the Thiol-Triggered Alkylating Properties Seen in
the Natural Product Leinamycin
Tonika Chatterji, Murat Kizil,† Kripa Keerthi, Goutam Chowdhury, Toma´s Posp´ısil, and
Kent S. Gates*
Departments of Chemistry and Biochemistry, UniVersity of Missouri, Columbia, Missouri 65211
Received October 31, 2002; E-mail: gatesk@missouri.edu
Scheme 1
Identification and characterization of new chemical motifs that
can efficiently modify biological macromolecules under physi-
ological conditions is of general importance in medicinal chemistry,
toxicology, and biotechnology. Biologically active natural products
have long served as an inspirational source of chemical strategies
for the intracellular generation of intermediates that react with
proteins and nucleic acids.1-4 In a recent example, studies of the
antitumor natural product leinamycin have revealed a novel
mechanism for the thiol-triggered generation of a DNA-alkylating
episulfonium ion.5-7 Reaction of thiols with the 1,2-dithiolan-3-
one 1-oxide heterocycle found in leinamycin initiates a sequence
of reactions that leads to the 1,2-oxathiolan-5-one intermediate 2
(Scheme 1).8,9 This intermediate undergoes a profound rearrange-
ment in which reaction of the C6-C7 double bond with the
electrophilic sulfur of the 1,2-oxathiolan-5-one heterocycle yields
an episulfonium ion (3) that efficiently alkylates the N7-position
of guanosine residues in double-stranded DNA.5 Leinamycin can
Scheme 2
also be transformed to the episulfonium ion 3 in the absence of
thiol,10 although, under typical cellular conditions (1-5 mM
glutathione, pH ≈ 7.2), thiol-mediated activation of the antibiotic
is approximately 1000 times faster than the thiol-independent
process and also proceeds in higher yield.10
In leinamycin, nature has provided us with a novel assembly of
functional groups that generates a potent alkylating agent upon
Scheme 3
exposure to the thiol-rich environment found inside cells. To
establish whether the chemistry of leinamycin represents a general
motif that can function in various molecular frameworks, we
undertook the construction of greatly simplified leinamycin ana-
logues containing only what we perceived to be the “core”
functional groups required for thiol-accelerated production of an
alkylating agent (Scheme 2).
We targeted compound 4 as the first “stripped-down” analogue
of leinamycin (Scheme 3). Despite the obvious structural differences
between the benzo-fused sulfur heterocycle found in 4 and the
saturated heterocycle found in the natural product, previous work
indicated that the 3H-1,2-benzodithiol-3-one 1-oxide system (5)
would emulate key aspects of leinamycin’s reactivity (specifically,
thiol-mediated formation of an oxathiolanone intermediate).8,11-13
Synthesis of the desired leinamycin analogue 4 was achieved as
shown in Scheme 3. The diazonium salt of dibromoaniline (6) was
reacted with trimethylsilylethanethiolate14,15 in the presence of
powdered copper16 to provide the aromatic sulfide 7. Metal-
halogen exchange, followed by reaction of the resulting lithium
anion with prenyl bromide, allowed installation of the alkene side
trimethylsilylethanethiol, furnished the thioester 10. Removal of
both trimethylsilylethane protecting groups from 10 using tetra-
chain in compound 8. A second metal-halogen exchange reaction,
followed by quenching of the lithium anion with carbon dioxide,
provided the carboxylic acid derivative 9. Conversion of the
carboxylic acid to the acid chloride, followed by reaction with
butylammonium fluoride and trifluoroacetic anhydride,17 followed
by in situ oxidation with I2/KI, resulted in formation of the 3H-
1,2-benzodithiolan-3-one heterocycle 11 in modest yield. Finally,
oxidation of 11 with dimethyldioxirane18 in acetone gave the desired
leinamycin analogue 4. In addition, we prepared the allyl derivative
12 using the same general approach, with the exception that it was
† Visiting Scholar from University of Dicle, Department of Chemistry, 21280
Diyarbakir, Turkey.
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J. AM. CHEM. SOC. 2003, 125, 4996-4997
10.1021/ja029169y CCC: $25.00 © 2003 American Chemical Society

C O M M U N I C A T I O N S
Scheme 4
In summary, we find that the relatively simple molecule 4 is
able to effectively mimic key elements of leinamycin’s chemical
reactivity. This work helps define the “core” functional groups
required for thiol-accelerated generation of an alkylating intermedi-
ate from leinamycin. The results suggest that substantially altered
analogues of the natural product may retain DNA-alkylating
properties. From a broader perspective, our findings provide
evidence that the intriguing cascade of chemical reactions first seen
in the context of leinamycin represents a general motif that can
operate in a variety of molecular frameworks. This realization could
facilitate the discovery of new natural and synthetic agents that
effect thiol-triggered alkylation of macromolecular targets inside
cells.
Acknowledgment. We are grateful to the National Institutes
of Health (CA83925) for financial support of this research, and
we thank Professor Miquel Yus (Universidad de Alicante, Spain)
for providing us with spectral data that helped confirm the structures
13.
necessary to employ the metalation-transmetalation procedure
developed by Snieckus and co-workers19 for installation of the allyl
side chain into 7.
Treatment of 4 with 1 equiv of thiol (either 2-mercaptoethanol
or propanethiol) in a 1:1 mixture of acetonitrile and sodium
phosphate buffer (100 mM, pH 7) led to rapid formation of the
cyclized product 13a which was isolated in approximately 50%
yield as its methyl ester following treatment of the organic extract
with diazomethane. This product is envisioned to arise from
Markovnikov addition of water to the episulfonium ion intermediate
14 (Scheme 4).20 It is noteworthy that the leinamycin-derived
episulfonium ion 3 also yields Markovnikov products upon reaction
with DNA or water.5 Thiol-triggered conversion of 4 to an
alkylating species also proceeds effectively in a 99:1 mixture of
methanol/sodium phosphate buffer. In this case, the analogous
product (13b) resulting from nucleophilic attack of methanol on
the episulfonium ion 14 was obtained in 50% yield as the methyl
ester after workup of the organic extract with diazomethane.
Interestingly, we find that the allyl analogue 12 is also efficiently
transformed to the methanol adduct 13c under these reaction
conditions. This indicates that formation of the intermediate
episulfonium ion is not strongly dependent upon the presence of
electron-donating alkyl substituents on the alkene.21
The leinamycin model compound 4 also emulates the thiol-inde-
pendent activation chemistry reported recently for leinamycin.10
Incubation of 4 in a 1:1 mixture of acetonitrile and sodium
phosphate buffer (100 mM, pH 7) for 11 h in the absence of thiol,
followed by diazomethane workup, affords 13a in 35% yield.
Similar to the natural product, thiol-independent conversion of 4
to the episulfonium ion is slower and somewhat less efficient than
the thiol-triggered reaction. The reaction of 4 (100 µM) with thiol
(10 equiv) is complete within 100 s under these conditions, whereas
the half-life for the conversion of this compound to products in
the absence of thiol is approximately 2 h.
Supporting Information Available: Experimental procedures and
spectral data for all compounds, comments on the structure elucidation
of 13, and DNA-damage assays (PDF). This material is available free
of charge via the Internet at http://pubs.acs.org.
References
(1) Gates, K. S. In ComprehensiVe Natural Products Chemistry; Kool, E. T.,
Ed.; Pergamon: New York, 1999; Vol. 7, pp 491-552.
(2) Wolkenberg, S. E.; Boger, D. L. Chem. ReV. 2002, 102, 2477-2495.
(3) Verdine, G. L. Nature 1996, 384(supplement), 11-13.
(4) Newman, D. J.; Cragg, G. M.; Snader, K. M. Nat. Prod. Rep. 2000, 17,
215-234.
(5) Asai, A.; Hara, M.; Kakita, S.; Kanda, Y.; Yoshida, M.; Saito, H.; Saitoh,
Y. J. Am. Chem. Soc. 1996, 118, 6802-6803.
(6) Gates, K. S. Chem. Res. Toxicol. 2000, 13, 953-956.
(7) Reaction of leinamycin with thiols also results in the generation of reactive
oxygen species. See: (a) Mitra, K.; Kim, W.; Daniels, J. S.; Gates, K. S.
J. Am. Chem. Soc. 1997, 119, 11691-11692. (b) Behroozi, S. B.; Kim,
W.; Dannaldson, J.; Gates, K. S. Biochemistry 1996, 35, 1768-1774.
(8) Behroozi, S. B.; Kim, W.; Gates, K. S. J. Org. Chem. 1995, 60, 3964-
3966.
(9) Breydo, L.; Gates, K. S. J. Org. Chem. 2002, 67, 9054-9060.
(10) Breydo, L.; Zang, H.; Mitra, K.; Gates, K. S. J. Am. Chem. Soc. 2001,
123, 2060-2061.
(11) Mitra, K.; Gates, K. S. Tetrahedron Lett. 1995, 36, 1391-1394.
(12) Mitra, K.; Pohl, M. E.; MacGillivray, L. R.; Barnes, C. L.; Gates, K. S.
J. Org. Chem. 1998, 62, 99361-9364.
(13) Zang, H.; Breydo, L.; Mitra, K.; Dannaldson, J.; Gates, K. S. Bioorg.
Med. Chem. Lett. 2001, 11, 1511-1515.
(14) Anderson, M. B.; Ranasinghe, M. G.; Palmer, J. T.; Fuchs, P. L. J. Org.
Chem. 1988, 53, 3125-3127.
(15) Schwan, A. L.; Brillon, D.; Dufault, R. Can. J. Chem. 1994, 72, 325-
333.
(16) Meijs, G. F.; Beckwith, A. L. J. J. Am. Chem. Soc. 1986, 108, 5890-
5893.
(17) Wang, Y.; Koreeda, M.; Chatterji, T.; Gates, K. S. J. Org. Chem. 1998,
63, 8644-8645.
(18) Adam, W.; Bialas, J.; Hadjiarapoglou, L. Chem. Ber. 1991, 124, 2377.
(19) Sibi, M. P.; Jalil Miah, M. A.; Snieckus, V. J. Org. Chem. 1984, 49,
737-747.
(20) Markovnikov addition of nucleophiles to episulfonium ions is common.
See, for example: Smit, W. A.; Zefirov, N. S.; Bodrikov, I. V.; Krimer,
M. Z. Acc. Chem. Res. 1979, 12, 282-288. Toshimitsu, A.; Hirosawa,
C.; Tanimoto, S. Tetrahedron Lett. 1991, 32, 4317-4320.
(21) Alkyl substituents are known to increase the rate at which alkenes react
with electrophilic sulfur compounds. Schmid, G. H.; Garratt, D. G. Can.
J. Chem. 1973, 51, 2463-2468.
Finally, it is important to report that 4 does not efficiently alkylate
DNA. This is not surprising because existing data indicate that
efficient DNA alkylation by leinamycin is driven by noncovalent
association of the activated antibiotic (3) with the double helix.5,22
Clearly, small analogues intended to recapitulate the DNA-
alkylating properties of leinamycin will need to be equipped with
additional functional groups that furnish noncovalent DNA binding.
(22) Breydo, L.; Zang, H.; Dutta, S.; Gates, K. S., unpublished data.
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