UMR9187 / U1196 – Chimie et Modélisation pour la Biologie du Cancer (CMBC)

Publications de l’unité

Année de publication : 2007

Isabelle Ourliac Garnier, Sophie Bombard (2007 Jan 17)

GG sequence of DNA and the human telomeric sequence react with cis-diammine-diaquaplatinum at comparable rates.

Journal of inorganic biochemistry : 101 : 514-24 : DOI : 10.1016/j.jinorgbio.2006.11.005 En savoir plus
Résumé

G-quadruplex structures of telomeric sequences are of growing interest because they inhibit telomerase, an enzyme involved in the maintenance of telomere length of cancer cells. As we have shown previously, the antiparallel structure of G-quadruplexes can be cross-linked in vitro by the anti-tumour drug cisplatin. The question arises whether platination of quadruplex structures of human telomeric sequences by cisplatin could be relevant from a biological point of view. Therefore, we have compared the kinetics of reactions of the diaqua form of cisplatin, cis-[Pt(NH(3))(2)(H(2)O)(2)](2+), with the human telomeric quadruplex structure, a duplex DNA and a single-stranded DNA containing one specific platination GG site. The ratio between the platination rate constants was obtained using two intramolecular competition experiments: either a construct with a junction between duplex DNA containing a unique GG platination site and the quadruplex structure of the human telomeric sequence AG(3)(T(2)AG(3))(3), or a construct with a junction between duplex DNA and a single strand containing each a unique GG platination site. Those competition experiments allowed us to conclude that the platination of the quadruplex is favoured over that of the GG duplex by a factor of about two whereas the GG duplex is platinated three times faster than the GG single strand.

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Année de publication : 2006

David Monchaud, Clémence Allain, Marie-Paule Teulade-Fichou (2006 Sep 15)

Development of a fluorescent intercalator displacement assay (G4-FID) for establishing quadruplex-DNA affinity and selectivity of putative ligands.

Bioorganic & medicinal chemistry letters : 16 : 4842-4845 : DOI : 10.1016/j.bmcl.2006.06.067 En savoir plus
Résumé

A fluorescent intercalator displacement assay (G4-FID) has been designed based on the displacement of thiazole orange (TO) positioned onto a quadruplex-forming oligonucleotide by putative ligands. This technique was validated by the use of a set of representative and fully characterized G-quadruplex binders (ranging from pyridodicarboxamide to macrocyclic ligands). To further extend its applicability, a comparative version has been developed which allows a rapid and viable determination of quadruplex- over duplex-selectivity.

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Clémence Allain, David Monchaud, Marie-Paule Teulade-Fichou (2006 Sep 7)

FRET templated by G-quadruplex DNA: a specific ternary interaction using an original pair of donor/acceptor partners.

Journal of the American Chemical Society : 128 : 11890-11893 : DOI : 10.1021/ja062193h En savoir plus
Résumé

G-quadruplex represents a suitable scaffold for FRET (fluorescence resonance energy transfer) since its two external quartets offer two well-defined binding sites for concomitant trapping of donor/acceptor partners. Combining selective G-quadruplex binders (macrocyclic bis(quinacridine) BOQ1 or monomeric quinacridine MMQ1, donor) with a highly fluorescent DNA probe (thiazole orange, acceptor), we designed a structure-specific FRET-system based on an unprecedented noncovalent ternary complex. This system could be potentially usable as a signature for quadruplex-DNA conformation in solution, but also might offer a unique means for observing cation and ligand binding influence on quadruplex topology.

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Markus Kaiser, Anne De Cian, Matthieu Sainlos, Christian Renner, Jean-Louis Mergny, Marie-Paule Teulade-Fichou (2006 Mar 10)

Neomycin-capped aromatic platforms: quadruplex DNA recognition and telomerase inhibition.

Organic & biomolecular chemistry : 4 : 1049-1057 : DOI : 10.1039/b516378a En savoir plus
Résumé

A series of aminoglycoside-capped macrocyclic structures has been prepared using intramolecular bis-tethering of neomycin on three aromatic platforms (phenanthroline, acridine, quinacridine). Based on NMR and calculations studies, it was found that the cyclic compounds adopt a highly flexible structure without conformational restriction of the aminoglycoside moiety. FRET-melting stabilization measurements showed that the series displays moderate to high affinity for the G4-conformation of human telomeric repeats, this effect being correlated with the size of the aromatic moiety. In addition, a FRET competition assay evidenced the poor binding ability of all macrocycles for duplex DNA and a clear binding preference for loop-containing intramolecular G4 structures compared to tetramolecular parallel G4 DNA. Finally, TRAP experiments demonstrated that the best G4-binder (quinacridine ) is also a potent and selective telomerase inhibitor with an IC(50) in the submicromolar range (200 nM).

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Markus Kaiser, Matthieu Sainlos, Jean-Marie Lehn, Sophie Bombard, Marie-Paule Teulade-Fichou (2006 Jan 13)

Aminoglycoside-quinacridine conjugates: towards recognition of the P6.1 element of telomerase RNA.

Chembiochem : a European journal of chemical biology : 7 : 321-329 : DOI : 10.1002/cbic.200500354 En savoir plus
Résumé

A modular synthesis has been developed which allows easy and rapid attachment of one or two aminoglycoside units to a quinacridine intercalator, thereby leading to monomeric and dimeric conjugates. Melting temperature (Tm) experiments show that the tobramycin dimeric conjugate TD1 exhibits strong binding to the P6.1 element of human telomerase RNA. By contrast, tobramycin alone is much less efficient and the monomeric compound TM1 elicits a poor binding ability. Monitoring of the interaction by an electrophoretic mobility shift assay shows a 1:1 stoichiometry for the binding of the dimeric compound to the hairpin structure and confirms the lower affinity for a control duplex. Protection experiments with RNase T1 indicate interaction of the drug both in the stem and in the loop of the hairpin. Taken together, the data suggest a binding of TD1 inside the hairpin at the stem-loop junction. The same trends are observed with paromomycin and kanamycin analogues but with a lower affinity.

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Année de publication : 2005

A Dupuy, A Shamsaldin, E Quiniou, C Paoletti, M Labbé, M F Avril, D Lefkopoulos, F de Vathaire (2005 Oct 26)

Risk of melanoma following adulthood cancer: a case-control study.

European journal of cancer (Oxford, England : 1990) : 41 : 2904-2910 : DOI : 10.1016/j.ejca.2005.07.020 En savoir plus
Résumé

Melanoma is a severe skin cancer related to sun exposure. Whether this malignancy is linked to exposure to ionising radiation during adulthood is still controversial. This case-control study examined the risk of melanoma following treatment for an adulthood first malignant neoplasm (FMN). Cases were patients who presented with cutaneous melanoma after a first cancer in adulthood. Controls (3 per case) were patients free of melanoma, matched for age, duration of follow-up since the FMN, type of FMN, and followed in the same institution. A total of 57 cases and 171 controls were included. In the final multivariate analysis, no risk of melanoma was associated with radiotherapy (odds ratio (OR) for 1 Gy = 1.01, 95% confidence interval (95%CI) 0.96-1.07) nor hormonotherapy, whereas chemotherapy use (OR = 2.3, 95%CI 0.93-5.6) and having a history of familial cancer (OR = 2.8, 95%CI 1.3-5.9) exhibited a nearly significant risk. In conclusion, unlike the evidence for risk of exposure to ionising radiation during childhood, we did not substantiate a risk for association of melanoma with exposure to ionising radiation during adulthood. The risk associated with chemotherapy should justify the implementation of skin surveillance for early detection of melanoma in these patients.

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Année de publication : 2003

Catherine Tetreau, Yves Blouquit, Eugene Novikov, Eric Quiniou, Daniel Lavalette (2003 Dec 26)

Competition with xenon elicits ligand migration and escape pathways in myoglobin.

Biophysical journal : 86 : 435-447 : DOI : 10.1016/S0006-3495(04)74120-X En savoir plus
Résumé

Evidence for ligand migration toward the xenon-binding cavities in myoglobin comes from a number of laser photolysis studies of MbO2 including mutants and from cryo- and time-resolved crystallography of MbCO. To explore ligand migration in greater detail, we investigated the rebinding kinetics of both MbO2 and MbCO under a xenon partial pressure ranging from 1 to 16 atm over the temperature range (293-77 K). Below 180 K xenon affects to a significant, but minor, extent the thermodynamic parameters for rebinding from the primary docking site in each Mb taxonomic substate. Above 200 K the ligand migrates to the proximal Xe1 site but when the latter is occupied by xenon a new kinetic process appears. It is attributed to rebinding from transient docking sites located on the path between the primary and the secondary docking site of both ligands. Ligand escape exhibits a more complicated pattern than expected. At room temperature O2 and CO escape appears to take place exclusively from the primary site. In contrast, at T approximately 250 K, roughly 50% of the CO molecules that have escaped from the protein originate from the Xe1 secondary site.

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Arnaud David, Nathalie Bleimling, Christine Beuck, Jean-Marie Lehn, Elmar Weinhold, Marie-Paule Teulade-Fichou (2003 Dec 9)

DNA mismatch-specific base flipping by a bisacridine macrocycle.

Chembiochem : a European journal of chemical biology : 4 : 1326-1331 : DOI : 10.1002/cbic.200300693 En savoir plus
Résumé

Most, if not all, enzymes that chemically modify nucleobases in DNA flip their target base from the inside of the double helix into an extrahelical position. This energetically unfavorable conformation is partly stabilized by specific binding of the apparent abasic site being formed. Thus, DNA base-flipping enzymes, like DNA methyltransferases and DNA glycosylases, generally bind very strongly to DNA containing abasic sites or abasic-site analogues. The macrocyclic bisacridine BisA has previously been shown to bind abasic sites. Herein we demonstrate that it is able to specifically recognize DNA base mismatches and most likely induces base flipping. Specific binding of BisA to DNA mismatches was studied by thermal denaturation experiments by using short duplex oligodeoxynucleotides containing central TT, TC, or TG mismatches or a TA match. In the presence of the macrocycle a strong increase in the melting temperature of up to 7.1 degrees C was observed for the mismatch-containing duplexes, whereas the melting temperature of the fully matched duplex was unaffected. Furthermore, BisA binding induced an enhanced reactivity of the mispaired thymine residue in the DNA toward potassium permanganate oxidation. A comparable reactivity has previously been observed for a TT target base mismatch in the presence of DNA methyltransferase M.TaqI. This similarity to a known base-flipping enzyme suggests that insertion of BisA into the DNA helix displaces the mispaired thymine residue into an extrahelical position, where it should be more prone to chemical oxidation. Thus, DNA base flipping does not appear to be limited to DNA-modifying enzymes but it is likely to also be induced by a small synthetic molecule binding to a thermodynamically weakened site in DNA.

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M P Teulade-Fichou, C Hounsou, L Guittat, J L Mergny, P Alberti, C Carrasco, C Bailly, J M Lehn, W D Wilson (2003 Oct 21)

Molecular recognition of quadruplex DNA by quinacridine derivatives.

Nucleosides, nucleotides & nucleic acids : 1483-1485 : DOI : 10.1081/NCN-120023016 En savoir plus
Résumé

The interaction of monomeric and dimeric quinacridines with quadruplex DNA has been investigated using a variety of biophysical methods. Both series of compounds were shown to exhibit a high affinity for the G4 conformation with two equivalent binding sites. As shown from the SPR and dialysis experiments the macrocyclic dimer appears more selective than its monomeric counterpart.

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S Guérin, A Dupuy, H Anderson, A Shamsaldin, G Svahn-Tapper, T Moller, E Quiniou, S Garwicz, M Hawkins, M F Avril, O Oberlin, J Chavaudra, F de Vathaire (2003 Oct 15)

Radiation dose as a risk factor for malignant melanoma following childhood cancer.

European journal of cancer (Oxford, England : 1990) : 39 : 2379-86 : DOI : 10.1016/S0959-8049(03)00663-4 En savoir plus
Résumé

The aim of this study was to determine therapy-related risk factors for the development of melanoma after childhood cancer. Among 4401 3-year survivors of a childhood cancer in eight French and British centres and 25120 patients younger than 20 years old at first malignant neoplasm (FMN) extracted from the Nordic Cancer Registries, 16 patients developed a melanoma as a second malignant neoplasm (SMN). A cohort study of the French and British cohorts was performed. In a nested case-control study, the 16 patients who developed a melanoma as a SMN (cases) were matched with 3-5 controls in their respective cohort according to gender, age at the first cancer, the calendar year of occurrence of the first cancer and follow-up. Radiotherapy appeared to increase the risk of melanoma for local doses >15 Gy, Odds Ratio (OR)=13 (95% Confidence Interval (CI): 0.94-174). Regarding chemotherapy, we observed an increased OR for both alkylating agents and spindle inhibitors, OR=2.7 (95% CI: 0.5-14). Children treated for a gonadal tumour as a FMN were found to be at a higher risk of melanoma, OR=8.7 (95% CI: 0.9-86). The adjusted OR for the local radiation dose was 1.07 (95% CI: 1.00-1.15). In conclusion, radiotherapy may contribute to an increased risk of melanoma as a SMN, but only at very high doses of low linear energy transfer radiation. Common genetic origins between gonadal tumours and malignant melanomas are likely.

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Marie-Paule Teulade-Fichou, Carolina Carrasco, Lionel Guittat, Christian Bailly, Patrizia Alberti, Jean-Louis Mergny, Arnaud David, Jean-Marie Lehn, W David Wilson (2003 Apr 17)

Selective recognition of G-Quadruplex telomeric DNA by a bis(quinacridine) macrocycle.

Journal of the American Chemical Society : 125 : 4732-4740 : DOI : 10.1021/ja021299j En savoir plus
Résumé

The interaction of G-quadruplex DNA with the macrocyclic compound BOQ1, which possesses two dibenzophenanthroline (quinacridine) subunits, has been investigated by a variety of methods. The oligonucleotide 5‘-A(GGGT2A)3G3, which mimics the human telomeric repeat sequence and forms an intramolecular quadruplex, was used as one model system. Equilibrium binding constants measured by biosensor surface plasmon resonance (SPR) methods indicate a high affinity of the macrocycle for the quadruplex conformation (K > 1 × 107 M1) with two equivalent binding sites. The affinity of BOQ1 for DNA duplexes is at least 1 order of magnitude lower. In addition, the macrocycle is more selective than the monomeric control compound (MOQ2), which is not able to discriminate between the two DNA structures (KduplexKquadruplex ≈ 106 M1). Strong binding of BOQ1 to G4 DNA sequences was confirmed by fluorometric titrations with a tetraplex-forming oligonucleotide. Competition dialysis experiments with a panel of different DNA structures, from single strands to quadruplexes, clearly established the quadruplex binding specificity of BOQ1. Fluorescence resonance energy transfer (FRET) Tm experiments with a doubly labeled oligonucleotide also revealed a strong stabilization of the G4 conformation in the presence of BOQ1 (ΔTm = +28 °C). This ΔTm value is one of the highest values measured for a G-quadruplex ligand and is significantly higher than observed for the monomer control compounds (ΔTm = +10−12 °C). Gel mobility shift assays indicated that the macrocycle efficiently induces the formation of G-tetraplexes. Strong inhibition of telomerase was observed in the submicromolar range (IC50 = 0.13 μM). These results indicate that macrocycles represent an exciting new development opportunity for targeting DNA quadruplexes.

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Année de publication : 2002

Sébastien Lyonnais, Candide Hounsou, Marie-Paule Teulade-Fichou, Josette Jeusset, Eric Le Cam, Gilles Mirambeau (2002 Dec 6)

G-quartets assembly within a G-rich DNA flap. A possible event at the center of the HIV-1 genome.

Nucleic acids research : 30 : 5276-5283 : DOI : 10.1093/nar/gkf644 En savoir plus
Résumé

Stretches of guanines can associate in vitro through Hoogsteen hydrogen bonding to form four-stranded structures. In the HIV-1 central DNA flap, generated by reverse transcriptase at the end of retrotranscription, both the two 99 nt-long overlapping (+) strands contain two adjacent tracts of guanines. This study demonstrates that oligonucleotides containing these G-clusters form highly stable G-quadruplexes of various structures in vitro, whose formation was controlled by an easy and reversible protocol using sodium hydroxide. Among these sequences, a G’2 hairpin dimer was the most stable structure adopted by the 5′-tail of the (+) downstream strand. Since the two (+) strands of the HIV-1 central DNA flap hold these G-clusters, and based on the properties of reverse branch migration in DNA flaps, constructions using HIV-1 sequences were assembled to mimic small DNA flaps where the G-clusters are neighbors. G-quartets were successfully probed in such flaps. They were induced by potassium and by a dibenzophenanthroline derivative already known to stabilize them. Such results suggest some function(s) for G-quartets associated with a DNA flap in the HIV-1 pre-integration steps, and argue for their transient formation during the processing of G-rich DNA flaps at the time of replication and/or repair.

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Jean-Louis Mergny, Jean-François Riou, Patrick Mailliet, Marie-Paule Teulade-Fichou, Eric Gilson (2002 Feb 14)

Natural and pharmacological regulation of telomerase.

Nucleic acids research : 30 : 839-865 : DOI : 10.1093/nar/30.4.839 En savoir plus
Résumé

The extremities of eukaryotic chromosomes are called telomeres. They have a structure unlike the bulk of the chromosome, which allows the cell DNA repair machinery to distinguish them from ‘broken’ DNA ends. But these specialised structures present a problem when it comes to replicating the DNA. Indeed, telomeric DNA progressively erodes with each round of cell division in cells that do not express telomerase, a specialised reverse transcriptase necessary to fully duplicate the telomeric DNA. Telomerase is expressed in tumour cells but not in most somatic cells and thus telomeres and telomerase may be proposed as attractive targets for the discovery of new anticancer agents.

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P Alberti, J Ren, M P Teulade-Fichou, L Guittat, J F Riou, J Chaires, C Hélène, J P Vigneron, J M Lehn, J L Mergny (2002 Jan 16)

Interaction of an acridine dimer with DNA quadruplex structures.

Journal of biomolecular structure & dynamics : 19 : 505-513 : DOI : 10.1080/07391102.2001.10506758 En savoir plus
Résumé

The reactivation of telomerase activity in most cancer cells supports the concept that telomerase is a relevant target in oncology, and telomerase inhibitors have been proposed as new potential anticancer agents. The telomeric G-rich single-stranded DNA can adopt an intramolecular G-quadruplex structure in vitro, which has been shown to inhibit telomerase activity. The C-rich sequence can also adopt a quadruplex (intercalated) structure (i-DNA). Two acridine derivatives were shown to increase the melting temperature of the G- quadruplex and the C-quadruplex at 1 microM dye concentration. The increase in Tm value of the G-quadruplex was associated with telomerase inhibition in vitro. The most active compound, « BisA », showed an IC(50) value of 0.75 microM in a standard TRAP assay.

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Année de publication : 2001

M P Teulade-Fichou, D Perrin, A Boutorine, D Polverari, J P Vigneron, J M Lehn, J S Sun, T Garestier, C Helene (2001 Sep 20)

Direct photocleavage of HIV-DNA by quinacridine derivatives triggered by triplex formation.

Journal of the American Chemical Society : 123 : 9283-9292 : DOI : 10.1021/ja0109040 En savoir plus
Résumé

Amino-p-quinacridine compounds (PQs) have been shown to stabilize strongly and specifically triple-helical DNA. Moreover, these derivatives display photoactive properties that make them efficient DNA cleavage agents. We exploited these two properties (triplex-specific binding and photoactivity) to selectively cleave a double-stranded (ds)DNA sequence present in the HIV-1 genome. Cleavage was first carried out on a linearized plasmid (3300 bp) containing the HIV polypurine tract (PPT) that allowed targeting by a triplex-forming oligonucleotide (TFO). PQ3, the most active compound of the series, efficiently cleaved double-stranded DNA in the vicinity of the PPT when this sequence had formed a triplex with a 16-mer TFO. Investigation of the cleavage at the molecular level was addressed on a short DNA fragment (56 bp); the photoinduced cleavage by PQ3 occurred only in the presence of the triple helix. Nevertheless, unusual cleavage patterns were observed:  damage was observed at guanines located 6−9 bp away from the end of the triple helical site. This cleavage is very efficient (up to 60%), does not require alkaline treatment, and is observed on both strands. A quinacridine−TFO conjugate produced the same cleavage pattern. This observation, along with others, excludes the hypothesis of a triplex-induced allosteric binding site of PQ3 adjacent to the damaged sequence and indicates that PQ3 preferentially binds in the vicinity of the 5‘-triplex junction. Irradiation in the presence of TFO-conjugates with acridine (an intercalative agent) and with the tripeptide lys-tryp-lys led to a complete inhibition of the photocleavage reaction. These results are interpreted in terms of competitive binding and of electron-transfer quenching. Together with the findings of simple mechanistic investigations, they led to the conclusion that the photoinduced damage proceeds through a direct electron transfer between the quinacridine and the guanines. This study addresses the chemical mechanism leading to strand breakage and characterizes the particular photosensitivity of the HIV−DNA target sequence which could be an oxidative hot spot for addressed photoinduced strand scission by photosensitizers.

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