Modifications of the DNA secondary structure upon platinum binding: a proposed model

114

Cell killing and mutation induction by cis-and trans-Pt(NH3)2Cl2 in Escherichia coli were examined by studying forward mutagenesis in the lad gene in cells with different repair capacities. Survival experiments showed that repair-proficient cells were slightly more sensitive for the cis isomer than for the trans isomer, whereas repair-deficient RecA and UvrB cells were extremely sensitive only for the cis compound. cis-Pt(NH3)2CI2 induced mutagenesis in both wild-type cells and RecA cells but not in UvrB cells; whereas no detectable mutagenesis was induced by treatment with the trans compound. Examination of the nature of the mutations induced by cis-Pt(NH3)WCI2, by. using the Lad system, revealed that base-pair substitutions leading to nonsense mutants are only induced in wild-type cells, suggesting that the intact products of both the uvrB and the recA gene are necessary for the repair responsible for this type of mutagenesis. Investigation ofthe nonsense mutants reveals that 70% ofthese mutations result from GC --TA or GC --AT substitutions at sites where the guanine is part of a GAG or GCG sequence. These results are discussed in relation to existing theories on the interaction between Pt compounds and DNA. A model for Pt-DNA adducts, leading to base-pair.substitutions, is proposed.About a decade ago, Rosenberg et aL (1) reported that cisPt(NH3)2CI2 shows antitumor activity against sarcoma 180 and leukemia L1210, whereas trans-Pt(NH3)2C12 is ineffective.Since then it has been shown that several other cis Pt(II) and cis Pt(IV) compounds exhibit antitumor activity (2, 3). In mammalian, as well as in bacterial cells, DNA is the preferential target for Pt compounds. For cis-Pt(NH3)2CI2 this interaction results in lesions that selectively block DNA replication (4, 5). In this respect, cis-Pt compounds behave similar to other drugs such as alkylating and radiomimetic agents.In vitro, cis-Pt(NH3)2C12 binds to bases in DNA, and the order of binding affinity has been shown to be guanine>ad-enine>cytosine>>thymine, with a strong preference for the N-7 position of guanine (6). Monofunctional binding to a single base is unlikely to be the principal lesion through which cisPt(NH3)2CI2 exerts its antitumor activity because, at equitoxic doses, more of the inactive trans compound is bound to DNA (7). Therefore, specific bifunctional binding of cis-Pt(NH3)2CI2 to DNA is thought to be responsible for its antitumor activity.For the bifunctional mode of action, several models have been proposed such as intrabase chelation at the 0-6 and N-7 positions of guanine (6,8), interstrand crosslinking between the N-7 positions of guanines in opposite strands (3,7,9), and intrastrand crosslinks between two, presumably adjacent, guanines in the same strand (10-12).In bacteria (13-17), as well as in eukaryotic cells (18)(19)(20), a correlation between mutagenicity and antitumor activity ofseveral Pt compounds has been found, suggesting that lesions leading to mutation events can also be responsible for antitumor activity. Several...

Section: Discussionsupporting

confidence: 90%

mentioning

confidence: 99%

See 1 more Smart Citation

Base-pair substitution hotspots in GAG and GCG nucleotide sequences in Escherichia coli K-12 induced by cis-diamminedichloroplatinum (II).

Brouwer¹,

Putte²,

Fichtinger-Schepman³

et al. 1981

114

“…In the in vitro reaction with nucleic acids the most frequent lesions are a cross-link between two bases and only a small proportion of monofunctional adducts has been detected, mainly for short incubation times (6)(7)(8)(9)(10)(11). In platinated DNA, the lesions prevent the binding of ethidium bromide (21).…”

Section: Resultsmentioning

confidence: 99%

Reaction of nucleic acids and cis-diamminedichloroplatinum(II) in the presence of intercalating agents.

Malinge¹,

Leng²

1986

The reaction of cis-diamminedichloroplatinum(II) and several synthetic or natural double-stranded polydeoxyribonudeotides has been carried out in the presence of such intercalating agents as ethidium bromide, proflavine, and acridine. After incubation of the reaction mixtures at 37C for 24 hr, some ethidium or proflavine, but no acridine, molecules are tightly bound to nucleic acids. Tight binding is defined by resistance to extraction with butanol, assayed by filtration at acid pH or by thin-layer chromatography at basic pH. In the ternary complexes, there is about one tightly bound ethidium (or proflavine) per platinum residue. At 370C, but not at 40C, tightly bound ethidium exchanges with free ethidium, whereas platinum residues do not exchange. The binding and the release of tightly bound ethidium are very slow (several hours). It is suggested that in the ternary complexes, nucleic acid-cis-Pt(NH3)2-intercalating agent, a bidentate adduct (guanine-ethidium or -proflavine)-cis-Pt(NH3)2, is formed. No tightly bound ethidium or proflavine is found when cis-diamminedichloroplatinum(II) is replaced by trans-diamminedichloroplatinum(II). Competition experiments between cis-diamminedichloroplatinum(II), poly(dG-dC), and poly(dG)--poly(dC) or poly(dA-dT) show that the presence of ethidium bromide, proflavine, or acridine interferes with the distribution of platinum between the polynucleotides. These results might help to explain the synergism for drugs used in combination with cis-diamminedichloroplatinum(ll) and in the design of new chemotherapeutic agents.Numerous works have been devoted to the study of the binding of the antitumor drug cis-diamminedichloroplatinum(II) [cis-Pt(NH3)2Cl2] to DNA. cis-Pt(NH3)2CI2 is clinically active against many different neoplasms and it is often used in combination with other anticancer drugs, such as doxorubicin (adriamycin) or bleomycin. Several studies have reported enhanced effects for combination drug therapy, including synergy (1).The mechanism of action of cis-Pt (NH3)2Cl2 is not yet known, but it is often thought that its antitumor activity is related to its binding to DNA (2-5). In the reaction of cis-Pt(NH3)2C12 and DNA, guanine residues are the preferred binding sites but adenine and cytosine residues also react, which leads to the formation of various types of adducts (6-11). On the other hand, for a given DNA, the nature of the adducts depends upon DNA conformation. In the reaction of cis-Pt(NH3)2CI2 and poly(dG-m5dC) in the Z conformation, a monodentate adduct is formed. In the reaction of cisPt(NH3)2CI2 and poly(dG-m5dC) in the B conformation, a bidentate adduct is formed (12).The conformations of B-DNA and Z-DNA are very different (13,14). To know whether smaller conformational changes of DNA could also play a role in the nature of the adducts, we have carried out the platination of nucleic acids in the presence of intercalating compounds. It has been already reported that new cis-Pt(NH3)2Cl2 binding sites were detected by means of exonuclease III when DNA ...

“…The trans isomer is inactive. Because DNA is believed to be the principal cellular target of the drug (3), several attempts have been made to distinguish the DNA-binding properties of the two isomers (4)(5)(6). Although variations in the kinetics of binding have been reported for cis-and trans-DDP, dramatic differences in their reactions with both linear and closed circular DNAs have not been seen.…”

Section: [Pt(cn)4]-2 Subsequent Two-dimensional Gel Electrophoresismentioning

confidence: 99%

Binding of cis - and trans -dichlorodiammineplatinum(II) to the nucleosome core

Lippard

Hoeschele

1979

The binding of the antitumor drug cis-dichlorodiammineplatinum(II) and its inactive trans isomer with the nucleosome core particle has been investigated. Kinetic demonstrated by treating the platinum/nucleosome core reaction mixtures with a protease or with nucleases prior to electrophoresis and observing changes in the gel patterns. Platinum was located in the gels through autoradiography using l9SmPt-labeled complexes. This work clearly demonstrates the greater propensity of trans-dichlorodiammineplatinum(II) to form histone-histone and histone-DNA crosslinks compared with the antitumor active cis isomer, which binds first to the DNA and only forms crosslinks to the histones when the nucleosome core is heavily loaded with platinum.cis-Dichlorodiammineplatinum(II) (cis-DDP) is an anticancer drug currently in widespread use (1, 2). The trans isomer is inactive. Because DNA is believed to be the principal cellular target of the drug (3), several attempts have been made to distinguish the DNA-binding properties of the two isomers (4-6). Although variations in the kinetics of binding have been reported for cis-and trans-DDP, dramatic differences in their reactions with both linear and closed circular DNAs have not been seen.DNA in the nuclei of eukaryotic cells is packaged as chromatin (7,8). This nucleic acid-protein complex may be extracted and digested with nucleases to produce a relatively stable, Mr 200,000 particle known as the nucleosome core. The core consists of "140 base pairs," hereafter referred to as 146 base pairs (9), of DNA wound by 13/4 turns about an octamer aggregate of eight histone proteins to produce a shallow superhelix with a pitch of about 28 A. Nucleosome cores contain two copies each of histone proteins H2a, H2b, H3, and H4. TheThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact. 6091 core particles, combined with one copy of histone H1 and an additional amount (Q6O base pairs in rat liver) of "linker" DNA, form the repeating unit of chromatin. The DNA of chromatin represents a likely cellular target for the antitumor drug cis-DDP. We have therefore initiated a program to study the reactions of cis-and trans-DDP, as well as other known or postulated antitumor drugs, with chromatin. In this initial report we describe the binding of cis-and trans-DDP to the nucleosome core. Striking differences in the binding and crosslinking properties of the two isomers have been observed that correlate nicely with protein-nucleic acid crosslinking results recently described for L1210 mouse leukemia cells treated with cis-and trans-DDP (10). Moreover, the discovery that trans-DDP forms specific, reversible proteinprotein and protein-nucleic acid crosslinks suggests that it might be a useful reagent for probing the structure of the nucleosome core particle and other nucleoprotein complexes.MATERIALS AND METHODS Chemicals and Biochemicals. c...