Alexandre S. Boutorine

DNA-photocleavage agents.

This review describes recent advances in the development of DNA-photocleavage agents. Major mechanisms of photosensitized DNA photocleavage are presented and the most popular categories of compounds are considered, which include metal complexes and many organic functional derivatives. DNA-targeted conjugates of photosensitizers are also described and discussed.

Oligonucleotides and Oligonucleotide Conjugates: A New Approach for Cancer Treatment

In this account we summarise recent studies on oligonucleotides and oligonucleotide derivatives and their utilisation in antigene, antisense and decoy approaches, with particular attention to peptide nucleic acids, locked nucleic acids and oligonucleotide conjugates, the most promising compounds in this field.

Formation of DNA Triple Helices by an Oligonucleotide Conjugated to a Fluorescent Ruthenium Complex

A conjugate of a triple helix forming oligonucleotide (TFO) and the Λ and Δ enantiomers of the ruthenium diphenanthroline dipyridophenazine complex [Ru(phen)2dppz]2+ was synthesized. The ruthenium complex was attached to the 5′‐end of the TFO through the dppz moiety. This conjugate formed a stable triple helix with the polypurine tract (PPT) sequence from HIV proviral DNA. The thermal denaturation temperature of the triplex was increased by 12 °C. One remarkable property of the Δ‐[Ru(phen)2dppz]2+ complex is a strong increase in its fluorescence when it intercalates into DNA. While the fluorescence of the oligonucleotide conjugate was very weak, the formation of a duplex with a complementary sequence or of a triple helix with a target duplex resulted in a large increase in fluorescence of the Δ enantiomer. The increase in fluorescence allowed us to follow the kinetics of duplex and triplex formation by fluorescence spectrometry. In contrast, the Λ enantiomer gave a much smaller fluorescence change when a triplex was formed, even though the stability of the triplex was comparable to that of the Δ enantiomer. The property was ascribed to intercalation of the dipyridophenazine moiety of the Δ enantiomer into DNA and subsequent threading of the ruthenium complex through the DNA double helix. Salt effects were consistent with the involvement of DNA breathing in the formation of the intercalating complex.

Rapid Routes of Synthesis of Oligonucleotide Conjugates from Non-Protected Oligonucleotides and Ligands Possessing Different Nucleophilic or Electrophilic Functional Groups

Abstract Optimized methods are described for post-synthetic conjugation of non-protected oligodeoxyribonucleotides to different ligands. Methods for the terminal functionalization of oligonucleotides by amino, sulfhydryl, thiophosphate or carboxyl groups using different chemical reactions and linkers in both organic and aqueous media are described and compared. Experimental conditions for subsequent coupling of ligands containing aliphatic and aromatic amines, aromatic alcohols, carboxylic, sulfiydryl, alkylating, aldehydic and other reactive nucleophilic and electrophilic groups to oligonucleotides were established, including covalent linkage to other oligonucleotides.

Triplex-Forming Twisted Intercalating Nucleic Acids (TINAs): Design Rules, Stabilization of Antiparallel DNA Triplexes and Inhibition of G-Quartet-Dependent Self- Association

The majority of studies on DNA triple helices have been focused on pH‐sensitive parallel triplexes with Hoogsteen CT‐containing third strands that require protonation of cytosines. Reverse Hoogsteen GT/GA‐containing antiparallel triplex‐forming oligonucleotides (TFOs) do not require an acidic pH but their applicability in triplex technology is limited because of their tendency to form undesired highly stable aggregates such as G‐quadruplexes. In this study, G‐rich oligonucleotides containing 2–4 insertions of twisted intercalating nucleic acid (TINA) monomers are demonstrated to disrupt the formation of G‐quadruplexes and form stable antiparallel triplexes with target DNA duplexes. The structure of TINA‐incorporated oligonucleotides was optimized, the rules of their design were established and the optimal triplex‐forming oligonucleotides were selected. These oligonucleotides show high affinity towards a 16 bp homopurine model sequence from the HIV‐1 genome; dissociation constants as low as 160 nM are observed whereas the unmodified TFO does not show any triplex formation and instead forms an intermolecular G‐quadruplex with Tm exceeding 90 °C in the presence of 50 mM NaCl. Here we present a set of rules that help to reach the full potential of TINA‐TFOs and demonstrate the effect of TINA on the formation and stability of triple helical DNA.

Directing topoisomerase I mediated DNA cleavage to specific sites by camptothecin tethered to minor- and major-groove ligands.

The covalent linkage of a hairpin polyamide, which binds in the minor groove, to camptothecin provides an efficient system to direct topoisomerase I mediated DNA cleavage to specific sites. These conjugates are equally as potent at targeting the enzyme to a single site in a DNA fragment as camptothecin conjugates of ligands that bind in the major groove (triplex-forming oligonucleotides).

Targeting topoisomerase I cleavage to specific sequences of DNA by triple helix-forming oligonucleotide conjugates. A comparison between a rebeccamycin derivative and camptothecin.

Topoisomerase I is an ubiquitous DNA cleaving enzyme and an important therapeutic target in cancer chemotherapy for the camptothecins as well as for indolocarbazole antibiotics such as rebeccamycin and its synthetic derivatives, which stabilize the cleaved DNA-topoisomerase I complex. The covalent linkage of a triple helixforming oligonucleotide to camptothecin or to the indolocarbazole derivative R-6 directs DNA cleavage by topoisomerase I to specific sequences. Sequence-specific recognition of DNA is achieved by the triple helix-forming oligonucleotide, which binds to the major groove of double-helical DNA and positions the drug at a specific site. The efficacy of topoisomerase I-induced DNA cleavage mediated by the rebeccamycin-conjugate and the camptothecin-conjugate was compared and related to the intrinsic potency of the isolated drugs.

Synthesis and Molecular Modeling Studies of Fullerene−5,6,7-Trimethoxyindole−Oligonucleotide Conjugates as Possible Probes for Study of Photochemical Reactions in DNA Triple Helices

The synthesis of novel functionalized fullerene derivatives containing DNA minor groove binders is reported. In order to construct sequence-specific probes for DNA photomodification, the compounds were attached to a triple helix forming oligonucleotide, and the formation of triplexes was monitored. The triplex formation was demonstrated, but the presence of fullerene moieties gives rise to a high degree of instability. Molecular modeling studies on these supramolecular structures provided useful information for the current study and for future developments.

Conjugates of Oligo(2′-O-Methylribonucleotides) with Minor Groove Binders as New Sequence-Specific Agents Recognizing Both Grooves of Double-Stranded DNA

Abstract Design, synthesis and physico-chemical studies of new pyrimidine oligo(2′-O-methylribonucleotide) conjugates with one or two oligo(pyrrolecarboxamide) minor groove binders (MGB) are described.

Synthesis of a hybrid fullerene–trimethoxyindole–oligonucleotide conjugate

The synthesis of novel functionalized fullerene derivatives is reported: a DNA minor groove binder such as trimethoxy- indole-2-carboxylate (TMI) and an oligonucleotide chain have been covalently linked to C60 with the aim of duplicating DNA interactions for increasing sequence selectivity.