Published on Web 06/13/2003
Solid-Phase Synthesis of Bleomycin Group Antibiotics.
Construction of a 108-Member Deglycobleomycin Library
Christopher J. Leitheiser, Kenneth L. Smith, Michael J. Rishel, Shigeki Hashimoto,
Kazuhide Konishi, Craig J. Thomas, Chunhong Li, Michael M. McCormick, and
Sidney M. Hecht*
Contribution from the Departments of Chemistry and Biology, UniVersity of Virginia,
CharlottesVille, Virginia 22904
Received November 27, 2002; E-mail: sidhecht@virginia.edu
Abstract: The bleomycins (BLMs) are structurally related glycopeptide antibiotics isolated from Streptomyces
verticillus that mediate the sequence-selective oxidative damage of DNA and RNA. Deglycobleomycin,
which lacks the carbohydrate moiety, cleaves DNA analogously to bleomycin itself, albeit less potently,
and has been used successfully for analyzing the functional domains of bleomycin. Although structural
modifications to bleomycin and deglycobleomycin have been reported, no bleomycin or deglycobleomycin
analogue having enhanced DNA cleavage activity has yet been described. The successful synthesis of a
deglycobleomycin on a solid support has permitted the facile solid-phase synthesis of 108 unique
deglycobleomycin analogues through parallel solid-phase synthesis. Each of the deglycobleomycin
analogues was synthesized efficiently; the purity of each crude product was greater than 60%, as determined
by HPLC integration. The solid-phase synthesis of the deglycobleomycin library provided near-milligram to
1
milligram quantities of each deglycobleomycin, thereby permitting characterization by H NMR and high-
resolution mass spectrometry. Each analogue demonstrated supercoiled plasmid DNA relaxation above
background cleavage; the library included two analogues that mediated plasmid relaxation to a greater
extent than the parent deglycobleomycin molecule.
Introduction
Despite a lesser cleavage efficiency and a decreased double- to
single-strand cleavage ratio for deglycobleomycin, the aglycon
The bleomycins (BLMs) are a family of glycopeptide-derived
antitumor antibiotics that were originally isolated from a culture
broth of Streptomyces Verticillus as copper chelates.1 A mixture
of bleomycins, consisting primarily of bleomycin A2 (∼60%)
and bleomycin B2 (∼30%) (Figure 1),2 are used for the treatment
of cancers.3 The therapeutic effects of bleomycin are believed
to derive in part from damage to chromosomal DNA in the
presence of a metal ion cofactor.4 Bleomycin can also degrade
some RNAs at micromolar concentrations,5 in both the presence5a,c
and absence of Fe2+ 5b
therapeutic locus.4e,5a,d
Deglycobleomycin (Figure 2) cleaves DNA in a sequence-
selective manner quite similar to that of bleomycin itself.6
is more readily accessible synthetically and has been used
successfully to define the essential structural elements necessary
to perform sequence-selective oxidative DNA degradation.7
Over the past two decades, numerous analogues of bleomycin
and deglycobleomycin have been synthesized to better under-
stand the mechanism of DNA cleavage by bleomycin.7 Most
of these have differed from the parent (deglyco)bleomycin only
within a single amino acid constituent.
Recently, the solid-phase synthesis of deglycobleomycin A5
has been reported.8 This has also permitted the solid-phase
syntheses of deglycobleomycins having varied C-termini9 and
,
and RNA may represent another
(6) (a) Oppenheimer, N. J.; Chang, C.; Ehrenfeld, G.; Rodriguez, L. O.; Hecht,
S. M. J. Biol. Chem. 1982, 257, 1606. (b) Sugiyama, H.; Ehrenfeld, G.
M.; Shipley, J. B.; Kilkuskie, R. E.; Chang, L.-H.; Hecht, S. M. J. Nat.
Prod. 1985, 48, 869. (c) Shipley, J. B.; Hecht, S. M. Chem. Res. Toxicol.
1988, 1, 25.
(1) Umezawa, H.; Suhara, Y.; Takita, T.; Maeda, K. J. Antibiot. 1966, 19A,
210.
(2) Anticancer Drugs: ReactiVe Metabolism and Drug Interactions; Powis,
G., Ed.; Pergamon Press: New York, 1994.
(7) (a) Takita, T.; Umezawa, Y.; Saito, S.; Morishima, H.; Naganawa, H.;
Umezawa, H.; Tsuchiya, T.; Miyake, T.; Kageyama, S.; Umezawa, S.;
Muraoka, Y.; Suzuki, M.; Otsuka, M.; Narita, M.; Kobayashi, S.; Ohno,
M. Tetrahedron Lett. 1982, 23, 521. (b) Aoyagi, Y.; Murugesan, N.;
Ehrenfeld, G. M.; Chang, L.-H.; Ohgi, T.; Shekhani, M. S.; Kirkup, M. P.;
Hecht, S. M. J. Am. Chem. Soc. 1982, 104, 5237. (c) Aoyagi, Y.; Katano,
K.; Suguna, H.; Primeau, J.; Chang, L.-H.; Hecht, S. M. J. Am. Chem.
Soc. 1982, 104, 5537. (d) Sugiura, Y.; Suzuki, T.; Otsuka, M.; Kobayashi,
S.; Ohno, M.; Takita, T.; Umezawa, H. J. Biol. Chem. 1983, 258, 1328.
(e) Umezawa, H.; Takita, T.; Sugiura, Y.; Otsuka, M.; Kobayashi, S.; Ohno,
M. Tetrahedron 1984, 40, 501. (f) Boger, D. L.; Honda, T.; Menezes, R.
F.; Colletti, S. L. J. Am. Chem. Soc. 1994, 116, 5631. (g) Boger, D. L.;
Honda, T. J. Am. Chem. Soc. 1994, 116, 5647. (h) Boger, D. L.; Cai, H.
Angew. Chem., Int. Ed. 1999, 38, 448. (i) Katano, K.; An, H.; Aoyagi, Y.;
Overhand, M.; Sucheck, S. J.; Stevens, W. C., Jr.; Hess, C. D.; Zhou, X.,
Hecht, S. M. J. Am. Chem. Soc. 1998, 120, 11285.
(3) Bleomycin Chemotherapy; Sikic, B. I., Rozencweig, M., Carter, S. K., Eds.;
Academic Press: Orlando, FL, 1985.
(4) (a) Hecht, S. M. Acc. Chem. Res. 1986, 19, 383. (b) Stubbe, J.; Kozarich,
J. W. Chem. ReV. 1987, 87, 1107. (c) Natrajan, A.; Hecht, S. M. In
Molecular Aspects of Anticancer Drug-DNA Interactions; Neidle, S.,
Waring, M. J., Eds.; Macmillan: London, 1994; pp 197-242. (c) Kane,
S. A.; Hecht, S. M. Prog. Nucleic Acid Res. Mol. Biol. 1994, 49, 313. (d)
Aso, M.; Kondo, M.; Suemune, H.; Hecht, S. M. J. Am. Chem. Soc. 1999,
121, 9023. (e) Hecht, S. M. J. Nat. Prod. 2000, 63, 158.
(5) (a) Hecht, S. M. Bioconjugate Chem. 1994, 5, 513. (b) Keck, M. V.; Hecht,
S. M. Biochemistry 1995, 34, 12029. (c) Holmes, C. E.; Duff, R. J.; van
der Marel, G. A.; van Boom, J.; Hecht, S. M. Bioorg. Med. Chem. 1997,
5, 1235. (d) Hecht, S. M. RNA as a Therapeutic Target for Bleomycin. In
The Many Faces of RNA; Eggleston, D. S., Prescott, C. D., Pearson, N.
D., Eds; Academic Press Ltd.: London, 1998; pp 3-17.
9
8218
J. AM. CHEM. SOC. 2003, 125, 8218-8227
10.1021/ja021388w CCC: $25.00 © 2003 American Chemical Society