The reaction between trans-diamminedichloroplatinum(II) and single-stranded oligonucleotides containing the sequence d(GXG) (X being an adenine, cytosine or thymine residue) yields trans-[Pt(NH3)2[(GXG)-GN7,GN7]] intrastrand cross-links. These cross-links do not prevent the pairing of the platinated oligonucleotides with their complementary strands but they decrease the thermal stability of the duplexes. The thermal stability is not much affected by the chemical nature of the X residue and its complementary base. By gel electrophoresis, it is shown that the trans- [Pt(NH3)2[d(GTG)-GN7,GN7]] cross-link bends the DNA double helix (26 degrees) and unwinds it (45 degrees). The pairing of the platinated oligonucleotides with their complementary strands promotes the rearrangement of the 1,3-intrastrand cross-links into interstrand cross-links. At a given temperature, the nature of the X residue, its complementary base and of the base pairs adjacent to the adducts do not dramatically affect the rate of the reaction. To know whether trans-[Pt(NH3)2[d(GXG)-GN7,GN7]] cross-links do not rearrange in some sequences, the location of these adducts was searched in double-stranded DNA after reaction with trans-diamminedichloroplatinum(II) by means of the 3'-5' exonuclease activity of T4 DNA polymerase. At low level of platination, trans-[Pt(NH3)2[d(GXG)-GN7,GN7]] cross-links were not detected. Monofunctional adducts and interstrand cross-links were mainly formed. These results are discussed in relation with the clinical inefficiency of trans-diamminedichloroplatinum(II).
In the reaction between trans-diamminedichloroplatinum(II) and a single-stranded pyrimidinerich oligodeoxyribonucleotide (22-mer) containing the central sequence TGAGT, the 1,3-trans-{Pt(NH3)2[d(GAG)J} crosslink is formed. The 1,3-intrastrand cross-link is inert within the single-stranded oligonucleotide. In contrast, it rearranges to an interstrand cross-link when the platinated oligonucleotide is paired with its complementary deoxyribo-or ribonucleotide strand. The half-life of the 1,3-intrastrand cross-link, -6 h at 370C, is independent of the nature and concentration of the salt (NaCl or NaCI04). It is not dramatically affected when the intervening adenine residue between the chelated guanine residues is replaced by a cytosine or a thymine residue or when the TA base pair adjacent to the 5' or 3' side of the adduct is replaced by a C-G base pair. On the other hand, a mismatch on the 3' or 5' side of the adduct prevents the rearrangement. We propose that the linkage isomerization reaction results from a direct nucleophilic attack of the cytosine residue complementary to the platinated 5' guanine residue on the platinum residue. Among others, the potential use of the DNA RNApromoted reaction is discussed in the context of the antisense strategy to irreversibly cross-link the antisense oligonucleotides to their targets. cis-Diamminedichloroplatinum(II) (cis-DDP) is a chemotherapeutic agent used largely in the clinical treatment of tumors. It is generally accepted that the cytotoxic action of cis-DDP is related to its ability to bind to cellular DNA and form covalent adducts. Although the major lesion is an intrastrand cross-link between two guanine (G) residues, it is not yet established that this lesion is responsible for the antitumor activity of cis-DDP (for general reviews, see refs. 1-4). Acquired resistance to cis-DDP may be associated with the increased gene-specific DNA repair efficiency of the interstrand adducts (5), a minor lesion in which the two G residues in the d(GC)-d(GC) site are cross-linked (6, 7).A question still under debate concerns trans-dichlorodiammineplatinum(II) (trans-DDP), the stereoisomer of cis-DDP. This compound is less mutagenic and less cytotoxic than cis-DDP and is ineffective as an antitumor drug (1)(2)(3)(4). In their reaction with DNA, the two isomers present similarities and differences (for general reviews, see refs. 8 and 9). The DNA modification by cis-or trans-DDP is controlled kinetically and the adducts are formed in two solvent-assisted reactions. The exchange of the chloro groups of both isomers is rate-limiting in both the initial attack of DNA and the closure of monofunctional adducts to bifunctional crosslinks. The preferred site of initial binding of the isomers is the N7 atom of G residues. Subsequently, the monofunctional adducts react with the neighboring bases to form intrastrand and interstrand cross-links. Among the differences between the two isomers, one is that stereochemical limitations preclude trans-DDP from forming intrastrand cross-links b...
In the reaction between trans-diamminedichloroplatinum(II) and single-stranded oligo(2'-O-methyl ribonucleotide)s containing the sequence GNG (N being a nucleotide residue), the 1,3-trans-{Pt-(NH3)2[GNG]} cross-links are formed. The 1,3-intrastrand cross-links are inert within the single-stranded oligonucleotides. By contrast, they rearrange into interstrand cross-links when the platinated oligonucleotides are paired with their complementary RNA strands. The rate of the interstrand cross-linking reaction depends upon the sequence facing the intrastrand cross-links. When the complementary sequences are 5'-CN'C (N' being a nucleotide), the rates are rather slow (T1/2 >/= 3 h at 37 degrees C). The rearrangement of the intrastrand cross-links into interstrand cross-links can be achieved in a few minutes when the triplets facing the intrastrand cross-links are replaced by doublet 5'-UA or 5'-CA. In vitro, the specificity of the cross-linking reaction between a platinated oligo(2'-O-methyl ribonucleotide) and its target sequence (containing the 5'-CA doublet) located within the coding region of Ha-ras mRNA is demonstrated by steric blocking of reverse transcriptase and translation machinery. Within the HBL100ras1 cells, this platinated oligonucleotide binds specifically and irreversibly to the cognate Ha-ras mRNA. It also inhibits the proliferation of the HBL100ras1 cells in a dose-dependent manner. The fast and specific interstrand cross-linking reaction triggered by the formation of a double helix between platinated oligo(2'-O-methyl ribonucleotide)s and RNA enhances the potential of the oligonucleotides which do not induce mRNA cleavage by RNase H, to modulate gene expression by steric blocking of the translation machinery.
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