Kinetic Analysis of the Stepwise Formation of a Long-Range DNA Interstrand Cross-link by a Dinuclear Platinum Antitumor Complex: Evidence for Aquated Intermediates and Formation of Both Kinetically an
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- 作者：John W. Cox ; Susan J. Berners-Price ; Murray S. Davies ; Yun Qu ; and Nicholas Farrell
- 刊名：Journal of the American Chemical Society
- 出版年：2001
- 出版时间：February 21, 2001
- 年：2001
- 卷：123
- 期：7
- 页码：1316 - 1326
- 全文大小：367K
- 年卷期：v.123,no.7(February 21, 2001)
- ISSN：1520-5126
文摘
Reported here is a detailed study of the kinetics and mechanism of formation of a 1,4 GG interstrandcross-link by [{trans-PtCl(NH3)2}2(
f">-NH2(CH2)nNH2)]2+ (1,1/t,t (n = 6), 1), the prototype of a novel class ofplatinum antitumor complexes. The reaction of the self-complementary 12-mer duplex 5'-{d(ATATGTACATAT)2} with 15N-1 has been studied at 298 K, pH 5.4, by [1H,15N] HSQC 2D NMR spectroscopy. Initialelectrostatic interactions with the duplex are observed for 1 and the monoaqua monochloro species (2). Aquationof 1 to yield 2 occurs with a pseudo-first-order rate constant of (4.15 ± 0.04) × 10-5 s-1. 2 then undergoesmonofunctional binding to the guanine N7 of the duplex to form 3 (G/Cl) with a rate constant of 0.47 ± 0.06M-1 s-1. There is an electrostatic interaction between the unbound {PtN3Cl} group of 3 and the duplex, whichis consistent with H-bonding interactions observed in the molecular model of the monofunctional (G/Cl) adduct.Closure of 3 to form the 1,4 GG interstrand cross-link (5) most likely proceeds via the aquated (G/H2O)intermediate (4) (pseudo-first-order rate constant = (3.62 ± 0.04) × 10-5 s-1) followed by closure of 4 toform 5 (rate constant = (2.7 ± 1.5) × 10-3 s-1). When closure is treated as direct from 3 (G/Cl) the rateconstant is (3.39 ± 0.04) × 10-5 s-1. Closure is ca. 10-55-fold faster than that found for 1,2 GG intrastrandcross-link formation by the diaqua form of cisplatin. Changes in the 1H and 15N shifts of the interstrand cross-link 5 indicate that the initially formed conformer (5(i)) converts irreversibly into other product conformer(s)5(f). The NMR data for 5(i) are consistent with a molecular model of the 1,4 GG interstrand cross-link onB-form DNA, which shows that the NH2 protons have no contacts except with solvent. The NMR data for 5(f)show several distinct NH2 environments indicative of interactions between the NH2 protons and the DNA.HPLC characterization of the final product showed only one major product peak that was confirmed by ESI-FTICR mass spectroscopy to be a cross-linked adduct of 15N-1 and the duplex. The potential significance ofthese findings to the antitumor activity of dinuclear platinum complexes is discussed.
f">-NH2(CH2)nNH2)]2+ (1,1/t,t (n = 6), 1), the prototype of a novel class ofplatinum antitumor complexes. The reaction of the self-complementary 12-mer duplex 5'-{d(ATATGTACATAT)2} with 15N-1 has been studied at 298 K, pH 5.4, by [1H,15N] HSQC 2D NMR spectroscopy. Initialelectrostatic interactions with the duplex are observed for 1 and the monoaqua monochloro species (2). Aquationof 1 to yield 2 occurs with a pseudo-first-order rate constant of (4.15 ± 0.04) × 10-5 s-1. 2 then undergoesmonofunctional binding to the guanine N7 of the duplex to form 3 (G/Cl) with a rate constant of 0.47 ± 0.06M-1 s-1. There is an electrostatic interaction between the unbound {PtN3Cl} group of 3 and the duplex, whichis consistent with H-bonding interactions observed in the molecular model of the monofunctional (G/Cl) adduct.Closure of 3 to form the 1,4 GG interstrand cross-link (5) most likely proceeds via the aquated (G/H2O)intermediate (4) (pseudo-first-order rate constant = (3.62 ± 0.04) × 10-5 s-1) followed by closure of 4 toform 5 (rate constant = (2.7 ± 1.5) × 10-3 s-1). When closure is treated as direct from 3 (G/Cl) the rateconstant is (3.39 ± 0.04) × 10-5 s-1. Closure is ca. 10-55-fold faster than that found for 1,2 GG intrastrandcross-link formation by the diaqua form of cisplatin. Changes in the 1H and 15N shifts of the interstrand cross-link 5 indicate that the initially formed conformer (5(i)) converts irreversibly into other product conformer(s)5(f). The NMR data for 5(i) are consistent with a molecular model of the 1,4 GG interstrand cross-link onB-form DNA, which shows that the NH2 protons have no contacts except with solvent. The NMR data for 5(f)show several distinct NH2 environments indicative of interactions between the NH2 protons and the DNA.HPLC characterization of the final product showed only one major product peak that was confirmed by ESI-FTICR mass spectroscopy to be a cross-linked adduct of 15N-1 and the duplex. The potential significance ofthese findings to the antitumor activity of dinuclear platinum complexes is discussed.