Anthracycline Antitumor Antibiotics

    Daunomycin (Dau) and Adriamycin (Adm) are the prototypical members in the anthracycline antitumor antibiotic family [1].  Despite their severe cardiotoxicity and other side effects, these drugs have been widely used as dose-limited chemotherapeutic agents for the treatment of human cancers such as leukemia since their discovery in the early 1960's [1].  These antibiotics contain a quinone-containing chromophore and an aminoglycoside sugar (Figure 1) [2].  The antineoplastic activity of these drugs has been mainly attributed to their strong interactions with DNA in the target cells.  There are two major mechanisms for these drugs to deform DNA structure and terminate its biological function [3]:  (1) an intercalation of the drugs into the base pairs in the DNA minor grooves, where the major contributions of the intercalative binding arise from hydrogen bonding, electrostatic, van der Waals, and hydrophobic interactions; and (2) via a free radical damage of the ribose, where the free radicals are formed during the redox cycle of the anthraquinone.  These drugs can be reduced to their semiquinone form by biological reducing agents, such as NADH and NADPH.  Superoxide (O2) and hydrogen peroxide (H2O2) can be produced via dioxygen receiving electrons from the semiquinone.  Then, hydroxyl radical (OH·) can be generated, which can attack cell components, such as membrane and DNA, and stops cell growth.

    A number of papers reported that metal ions (Fe2+/3+, Cu+/2+, and Tb3+) played an important role in altering the biochemical properties of the anthracyclines, and indicated a new direction in the pursuit of chemotherapeutic efficacy and lowering toxicity of these antibiotics [4].  The binding of metal ions may cause a significant influence on the redox property of these drugs, thus affecting their activity.  The interactions of these metal-drug complexes with DNA and other cell components, and their subsequent damage by the metal complexes have also been previously studied by the use of various physical and biochemical methods [5].

    Iron is an important element in that it participates in the action of several drug functionings (such as bleomycin [6] and streptonigrin [7]) as a redox center, which can generate free radicals in the presence of dioxygen under reducing conditions and damage cell components.  It has been shown that Fe3+ ion can bind three anthracycline molecules in aqueous solution, with the metal chelated by the 11,12-a-ketophenolate group.4g  A 1:2 Fe3+-adriamycin complex forms a stable complex with calf-thymus DNA in solution, and this tertiary complex is distinct from both the free Fe3+-drug complex and DNA-intercalated drug on the basis of optical and chromatographic studies [5e].  However, Fe3+-anthracyclines have also been observed not being able to intercalate into DNA base pairs until releasing the Fe3+ ion, despite the strong binding of Fe3+ with the drugs [4g].  Although studies have shown significant interactions between metal-drug complexes and DNA that resulted in DNA damage, the structures of these metal complexes have not been clearly described in all the previous studies [5].

    Since a large variety of metal ions (such as Mg2+, Ca2+, and the transition metal ions) exist in living organisms and are presumably available for binding with these drugs, the study of metal-drug interactions is crucial for a better understanding of the drug action in vivo.  A better understanding of metal-drug interactions and the formation of the metal-drug complexes will enable us to gain further insight into their antibiotic mechanisms.  In addition to the above described, several other metal ions with preferable spectroscopic and magnetic properties have been utilized as probes for the study of metal-anthracycline interactions.  Particularly, the lanthanide(III) (Ln3+) ions have been used as substitutes for the alkaline earth metal ions, and have also served as non-redox active metal substitutes for transition metal ions in those studies [8].  However, some questions regarding the metal binding and the structure of the complexes remain unanswered, such as the identity of different species detected in the spectra, the stoichiometry of the metal-drug complexes, and the metal binding mode.

    The Ln3+ ions are considered useful probes for the study of this drug system because of their unique magnetic and chemical properties [9,10]:  (1) The paramagnetic nature (J = 1/2 to 15/2) of some Ln3+ ions allows us to look into the structures of their drug complexes by the use of NMR, where the 1H NMR signals of a coordinated drug are paramagnetically shifted by the Ln3+ ions which act as “shift reagents” [11].  (2) Like the alkaline earth metal ions, Ln3+ ions have a strong affinity to oxygen-rich ligands and can bind to the a-ketophenolate moiety of these antibiotics.  This ligand binding property makes Ln3+ ions the best substitutes for the alkaline earth metal ions in biomolecules with oxygen-rich binding environments.  (3) The radii, charges, Lewis acidity, and preferred oxygen-rich ligand environment of the Ln3+ ions are similar to those of Fe3+, despite their different preferred coordination geometries, suggesting that a better study of the physical and chemical properties of Ln3+-drug complexes may afford a better understanding of the iron-drug complexes.

    We have studied several paramagnetic Ln3+-anthracycline complexes by the use of optical, electrochemical, and 1D and 2D 1H NMR techniques [12,13].  The use of 2D 1H NMR techniques allows us to assign all the proton signals of these Ln3+-drug complexes and to determine their configurations in solution which was not accomplished in previous studies.  On the basis of the optical and NMR studies, we conclude that four different Ln3+-drug complexes with metal-to-drug ratios 1:1, 1:2 1:3, and 2:1 complexes can be prepared in solution (plus an amorphous polymeric form) are formed in solution under different proton and metal ion concentrations.  These complexes are under fast chemical exchange with each other in solution.  With a complete assignment of the 1H NMR spectrum of the 1:1 Yb3+-drug complex [12], the spectra of the other species are possible to assign by the use of exchange spectroscopy (EXSY).  The configurations of the lanthanide(III) complexes of the drug daunomycin have been determined to be the same as the crystal structure of the free drug (see structure).  We use these lanthanide(III)-anthracycline complexes as model systems for the understanding of the binding of alkaline earth metal and transition metal ions with these drugs [14], and the effect of metal ion binding on the action of other quinone-containing drugs such as streptonigrin.

    [1] (a) Arcamone, F. Topics Antibiot. Chem. 1979, 2, 102. (b) Arcamone, F. Doxorubicin Anticancer Antibiotics; Academic:  New York, 1981. (c) Weiss, R. B.; Sarosy, G.; Clagett-Carr, K.; Russo, M.; Leyland-Jones, B. Cancer Chemother. Pharmacol. 1986, 18, 185.  (d) Lown, J. W.; Ed.; Anthracycline and Anthracenedione-based Anticancer Agents, Elsevier:  Amsterdam, 1988.
    [2] Courseille, C.; Busetta, B.; Geoffre, S.; Hospital, M.  Acta Cryst., 1979, B35, 764-767.
    [3] (a) Lown, J.W. Mol. Cell. Biochem. 1983, 55, 17. (b) Pindur, U.; Haber, M.; Sattler, K. J. Chem. Edu. 1993, 70, 263. (c) Lown, J. W. Chem. Soc. Rev. 1993, 22, 165.
    [4] A general reference:  (a) Martin R. B. In Metal Ions in Biological Systems; Sigel, H.; Ed.; Dekker:  New York, 1985 Vol. 19.  About copper binding:  (b) Greenaway, F. T.; Dabrowiak, J. C. J. Inorg. Biochem. 1982, 16, 91-107.  (c) Tachibana, M. Iwaizumi, M.; Tero-Kubota, S. J. Inorg. Biochem. 1987, 30, 133-140.  About iron binding:  (d) Matzanke, B. F.; Bill, E.; Butzlaff, C.; Trautwein, A. X.; Winkler, H.; Hermes, C.; Nolting, H.-F.; Barbieri, R.; Russo, U. Eur. J. Biochem. 1992, 207, 747-755.  (e) Massoud, S. S.; Jordan, R. B. Inorg. Chem. 1991, 30, 4851-4856.  (f) Gelvan, D.; Berg, E.; Saltman, P.; Samuni, A. Biochem. Pharmacol. 1990, 39, 1289-1295. (g) Beralso, H. Garnier-Suillerot, A.;Tosi, L.; Lavelle, F. Biochemistry 1985, 24, 284-289.  About other metal binding:  (h) Moustatih, A.; Fiallo, M. M. L.; Garnier-Suillerot, A. J. Med. Chem. 1989, 32, 336-342.  (i) Fiallo, M. M. L.; Garnier-Suillerot, A. Biochemistry 1986, 25, 924-930.  (j) Pasini, A.; Pratesi, G.; Savi, G.; Zunino, F. Inorg. Chim. Acta 1987, 137, 123-124.  (k) Allman, T.; Lenkinski, R. E. J. Inorg. Biochem. 1987, 30, 35-43.
    [5] Copper complexes:  (a) Phillips, D. R.; Carlyle, G. A. Biochem. Pharmacol. 1981, 30, 2021-2024.  (b)Spinelle, M.; Dabrowiak, J. C. Biochemistry 1982, 21, 5862-5870.  (c) Mariam, Y. H.; Glover, G. P. Biochem. Biophys. Res. Commun. 1986, 136, 1-7.  Iron complex:  (d) Myers, C. E.; Gianni, L.; Simone, C. B.; Klecker, R.; Greene, R. Biochemistry 1982, 21, 1707-1713.  (e) Eliot, H.; Gianni, L.; Myers, C. Biochmistry 1984, 23, 928-936. (f) Beraldo, H.; Garnier-Suillerot, A.; Tosi, L.; Lavelle, F. Biochemistry 1985, 24, 284-289.  (g) Cullinane C.; Phillips, D. R. Biochmeistry 1990, 29, 5638-5646.  (h) Akman, S. A.; Doroshow, J. H.; Bruke, T. G.; Dizdaroglu, M. Biochemsitry 1992, 31, 3500-3506.
    [6] Hecht, S. M. Acc. Chem. Res. 1986, 19, 383-391.
    [7] Hajdu, J. In Metal Ions in Biological Systems, vol. 19; Siegel, H., ed.; Dekker:  New York, 1985.
    [8] Mclennan, I. J.; Lenkinski, R. E. J. Am. Chem. Soc. 1984, 106, 6905. (a) Lenkinski, R. E.; Sierke, S.: Vist, M. R. J. Less-comm Met. 1983, 94, 359-365. (b)Lenkinski, R. E.; Sierke, S. J. Inorg. Biochem. 1985, 24, 59-67. (c) Mariam, Y. H.; Wells, W. J. Sol. Chem. 1984, 13, 259, 269. (d) Canada, R., G.; Carpentier, R.G. Bioichim. Biophy. Acta 1991, 1073, 136.  (e) Haj-Tajeb, H. B.; Fiallo, M. M. L.; Garnier-Suillerot, A.; Kiss, T.; Kozlowski, H. J. Chem. Soc. Dalton Trans. 1994, 3689-3693.
    [9] (a) Ming, L.-J.; "Paramagnetic Lanthanide(III) Ions as NMR Probes for Biomolecular Structure and Function"  In La Mar, G. N.; Ed.  Nuclear Magnetic Resonance of Paramagnetic Macromolecules, NATO-ASI, Kluwer:  Dordrecht, Netherlands, 1995. (b) Ming, L.-J.; Magn. Reson. Chem. 1993 33, S104.
    [10] (a). Bunzli, J.-C. G.; Choppin, G. R.; Lanthanide Probes in Life, Chemical and Earth Science, Elsevier:  Amsterdam, 1989. (b) Evans, C. H.; Biochemistry of the Lanthanides, Plenum:  New York, 1990.
    [11] (a) Morrill, T. C., Ed.  Lanthanide Shift Reagents in Stereochemical Analysis; VCH:  NY, 1986.  (b) La Mar, G. N.; Horrocks, W. DeW., Jr.; Holm, R. H. NMR of Paramagnetic Molecules, Chapters 12 & 13; Academic: NY, 1973.
    [12] Ming, L.-J.; Wei, X. Inorg. Chem. 1994, 33, 4617-4618.
    [13] Wei, X.; Ming, L.-J. Inorg. Chem. 1998, 37, 2255-2262.
    [14] (a) We have studied the binding of the transition metal ions Co(II) and Fe(II) with these drugs in methanol, and found that their binding modes with these drugs are similar to that of Yb3+ ion.  (b) Wei, X. Ph.D. Dissertation, Chapter 2, University of South Florida, 1996