RESUMEN
Its broad spectrum of biological activity makes benzimidazole a fundamental pharmacophore in pharmaceutics. The paper describes newly synthesized AT-specific fluorescent bis-benzimidazole molecules DB2Py(n) that contain a pyrrolcarboxamide fragment of the antibiotic drug netropsin. Physico-chemical methods using absorption, fluorescence, and circular dichroism spectra have shown the ability of bis-benzimidazole- pyrroles to form complexes with DNA. The new DB2Py(n) series have turned out to be more toxic to human tumor lines and less vulnerable to non-tumor cell lines. Bis-benzimidazole-pyrroles penetrated the cell nucleus, affected the cell-cycle synthesis (S) phase, and inhibited eukaryotic topoisomerase I in a cellfree model at low concentrations. A real-time tumor cell proliferation test confirmed the molecule's enhanced toxic properties upon dimerization. Preliminary cytotoxicity data for the bis-benzimidazole-pyrroles tested in a cell model with a MDR phenotype showed that monomeric compounds can overcome MDR, while dimerization weakens this ability to its intermediate values as compared to doxorubicin. In this respect, the newly synthesized cytotoxic structures seem promising for further, in-depth study of their properties and action mechanism in relation to human tumor cells, as well as for designing new AT-specific ligands.
RESUMEN
In this work, we synthesized and characterized the properties of a series of new fluorescent DB3(n) narrow-groove ligands. DB3(n) compounds based on dimeric trisbenzimidazoles have the ability to bind to the AT regions of DNA. The synthesis of DB3(n), whose trisbenzimidazole fragments are linked by oligomethylene linkers of different lengths (n = 1, 5, 9), is based on the condensation of the MB3 monomeric trisbenzimidazole with α,ω-alkyldicarboxylic acids. DB3 (n) proved to be effective inhibitors of the catalytic activity of HIV-1 integrase at submicromolar concentrations (0.20-0.30 µM). DB3(n) was found to inhibit the catalytic activity of DNA topoisomerase I at low micromolar concentrations.
Asunto(s)
Replicación del ADN , ADN , Secuencia de Bases , Ligandos , ADN/química , ColorantesRESUMEN
Antiviral activity of new AТ-specific fluorescent symmetric dimeric bisbenzimidazoles of DBÐ(n) series was assessed in the cell models of infections caused by type 1 herpes simplex virus (HSV1) and human cytomegalovirus (CMV). In DBA(n) molecules bisbenzimidazole fragments are bound to an oligomethylene liner with varied number of methylene groups in the linker (n = 1, 3, 5, 7, 9, 11). In contrast to DB(n) dimeric bisbenzimidazoles, in DBA(n) series terminal fragments of macromolecules contain N-dimethylaminopropylcarboxamide groups instead of N-methylpiperazine groups. DBÐ(n) compounds better dissolve in water, pass across plasma and nuclear membrane, and stain DNA in living cells. DBA(1) and DBA(7) produced therapeutic effects in HSV1 infection; DBA(7) completely suppressed the infection. DBA(11) displayed in vitro therapeutic activity in HSV1 and CMV infections. In addition, DBA(7) and DBA(1) showed microbicidal activity. Thus, DBA(11), which is active against two viruses causing severe diseases with serious health consequences for immunodeficient individuals, should be further investigated. High antiviral activity of DBA(7) in all test models indicates that this compound is a promising active agent for innovative antiviral drugs.
Asunto(s)
Antivirales/farmacología , Bisbenzimidazol/farmacología , Citomegalovirus/efectos de los fármacos , Simplexvirus/efectos de los fármacos , Infecciones por Citomegalovirus/tratamiento farmacológico , Herpes Simple , Infecciones por Herpesviridae , HumanosRESUMEN
Cancer cells are characterized by the hypermethylation of promoter regions of tumor suppressor genes. DNA methyltransferase inhibitors cause re-activation of these genes that allows considering DNA methyltransferases as targets for anticancer therapy. As it was previously shown by us, dimeric bisbenzimidazoles, DB(n), differing in length of the oligomethylene linker between the two bisbenzimidazole fragments (n--number of methylene groups in linker) effectively inhibit the methylation of DNA duplexes by murine methyltransferase Dnmt3a. Here, the cytotoxicity of some of these compounds, their penetration into cells and influence on the methylation of genomic DNA in fetal lung fibroblasts line F-977 and cervical cancer cells HeLa have been studied. In the 0-60 microM concentration range, only the DB(11) displayed a significant toxic effect on the normal cells, whereas the effect of DB(n) investigated on the cancer cells was not significant. Interestingly, the DB(1) and DB(3) to a small extent stimulate the proliferation of HeLa and F-977 cells, respectively. DB(1) and DB(3) display ability to penetrate into the nucleus of HeLa and F-977 cells and accumulate in various parts of the nuclei. DB(11) is not able to penetrate into the nuclei of these cells. The incubation of F-977 cells with 26 microM of DB(1) or DB(3) led to a decrease of the methylation of 18S rRNA gene, which is located in the region of DB(1) and DB(3) accumulation. A similar effect produces the same concentration of DB (3) in the F-977 cells. However, the overall level of genomic DNA methylation was not changed. These data suggest that DB(n) can be directed to act on specific genes demethylation and in the future may selectively inhibit the proliferation of cancer cells.
Asunto(s)
Bisbenzimidazol/farmacología , ADN (Citosina-5-)-Metiltransferasas/antagonistas & inhibidores , Metilación de ADN/efectos de los fármacos , Neoplasias/genética , Animales , Bisbenzimidazol/química , Proliferación Celular/efectos de los fármacos , ADN (Citosina-5-)-Metiltransferasas/genética , ADN (Citosina-5-)-Metiltransferasas/metabolismo , ADN Metiltransferasa 3A , Femenino , Células HeLa , Humanos , Ratones , Terapia Molecular Dirigida , Neoplasias/tratamiento farmacológico , ARN Ribosómico 18S/genéticaRESUMEN
Double-stranded DNA is a one of the most important intracellular anticancer agent targets. Disturbance of DNA functions as well as DNA structure lead to disorder of such processes as transcription and/or translation thus inducing tumor cells death. Complex formation between novel dimeric bisbenzimidazole DB(7) and poly(dA-dT) duplex in comparison with known monomeric bisbenzimidazole MB(Ac) was investigated in this study. DB(7)-poly(dA-dT) binding constant was determined by fluorescence spectroscopy using Scatchard plot and it values 1.18 x 10(8) M(-1) that is two orders of magnitude larger than MB(Ac) one (2.06 x 10(6) M(-1)). Thus, from findings mentioned above it could be concluded that the presence of two bisbenzimidazole moieties in the ligand structure significantly increases its affinity to the polynucleotide which motivates the synthesis of new potential anticancer drugs based on dimeric bisbenzimidazoles.
Asunto(s)
Antineoplásicos/química , Bisbenzimidazol/análogos & derivados , Bisbenzimidazol/química , ADN/química , Colorantes Fluorescentes/química , Oligodesoxirribonucleótidos/química , Antineoplásicos/farmacología , Bisbenzimidazol/farmacología , Muerte Celular/efectos de los fármacos , Colorantes Fluorescentes/farmacología , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismoRESUMEN
Five fluorescent symmetric dimeric bisbenzimidazoles DB(n) have been synthesized containing four 2,6-substited benzimidazole fragments and differ in length of oligomethylene linker (n=3, 4, 5, 7, 11) between the two bisbenzimidazole blocks. The ability of these dimeric bisbenzimidazoles to form complexes with double-stranded DNA (dsDNA) was shown by spectral methods. Upon binding to dsDNA DB(n) are localized in the minor groove. The DNA-methyltransferase Dnmt3a inhibition data are demonstrate the site-specific binding of dimeric bisbenzimidazoles DB(3) and DB(11) with oligonucleotide duplex.
Asunto(s)
Emparejamiento Base , Bisbenzimidazol/síntesis química , ADN/química , Colorantes Fluorescentes/síntesis química , Animales , Sitios de Unión , Bisbenzimidazol/química , Bovinos , ADN (Citosina-5-)-Metiltransferasas/antagonistas & inhibidores , ADN (Citosina-5-)-Metiltransferasas/química , Metilación de ADN , ADN Metiltransferasa 3A , ADN-Topoisomerasas de Tipo I/química , Colorantes Fluorescentes/química , Ligandos , Ratones , Sensibilidad y EspecificidadRESUMEN
When located in the DNA minor groove, dimeric bisbenzimidazoles DB(n) effectively inhibited in vitro the Dnmt3a catalytic domain (IC50 5-77 µM). The lowest IC50 value was observed for compound DB(11) with an 11-unit methylene linker joining the bisbenzimidazole fragments. Increased time of incubation of DNA with DB(n) as well as the presence of AT-clusters in DNA enhances the inhibitory effect.
Asunto(s)
Bisbenzimidazol/farmacología , Metilación de ADN/efectos de los fármacos , Metilasas de Modificación del ADN/química , Metilasas de Modificación del ADN/metabolismo , Animales , Bisbenzimidazol/síntesis química , Bisbenzimidazol/química , Dominio Catalítico , Espectroscopía de Resonancia Magnética , Ratones , Estructura MolecularRESUMEN
Here we studied the inhibition of the catalytic domain of Dnmt3a methyltransferase (Dnmt3a-CD) by DNA duplexes containing the mechanism-based inhibitor pyrimidine-2(1H)-one (P) instead of the target cytosine. It has been shown that conjugates of Dnmt3a-CD with P-DNA (DNA containing pyrimidine-2(1H)-one) are not stable to heating at 65°C in 0.1% SDS. The yield of covalent intermediate increases in the presence of the regulatory factor Dnmt3L. The importance of the DNA minor groove for covalent intermediate formation during the methylation reaction catalyzed by Dnmt3a-CD has been revealed. P-DNA was shown to inhibit Dnmt3a-CD; the IC(50) is 830 nM. The competitive mechanism of inhibition of Dnmt3a-CD by P-DNA has been elucidated. It is suggested that therapeutic effect of zebularine could be achieved by inhibition of not only Dnmt1 but also Dnmt3a.
Asunto(s)
ADN (Citosina-5-)-Metiltransferasas/antagonistas & inhibidores , Inhibidores Enzimáticos/química , Pirimidinonas/química , Animales , Unión Competitiva , Dominio Catalítico , ADN/metabolismo , ADN (Citosina-5-)-Metiltransferasas/metabolismo , Metilación de ADN , ADN Metiltransferasa 3A , Inhibidores Enzimáticos/farmacología , Ratones , Pirimidinonas/farmacología , Temperatura de TransiciónRESUMEN
HIV-1 integrase is responsible for one of the key steps of the viral replication, integration of the viral cDNA into the host cell genome. Integration inhibition leads to complete block of the virus replication. In this study inhibition of integration by dimeric bisbenzimidazoles DBBI(7) with heptamethylene and DBBI(8) with tri(ethylene glycol) spacers was examined, and it was learned out that IC50 for DBBI(7) was about 0.03 microM, and IC50 for DBBI(8) was about 10 microM. By using cross-linking assays, it was shown that both compounds impeded a proper disposition of DNA-substrate at the active centre of integrase. Dissociation constants for complexes between either DBBI and DNA-substrate of integrase were determined. Calculated Kd values were 270 nM and 140 nM for complexes formed by DBBI(7) and DBBI(8), respectively. Therefore, inhibition of integration does not directly result from the binding of DBBIs with DNA. The dependence of initial rates of enzymatic reaction on the DNA-substrate concentration in presence of different concentrations of inhibitors was found, and inhibition constants were determined. All the data obtained allow us to suppose that the different inhibition activity of DBBI(7) and DBBI(8) results from the different mechanism of their binding: DBBI(7) is a competitive inhibitor of integrase whereas DBBI(8) is assumed to show a more complex mechanism of inhibition.
Asunto(s)
Bisbenzimidazol/química , ADN/química , Inhibidores de Integrasa VIH/química , Integrasa de VIH/química , VIH-1/enzimología , Bisbenzimidazol/análogos & derivados , Bisbenzimidazol/metabolismo , Dominio Catalítico , ADN/metabolismo , Integrasa de VIH/metabolismo , Cinética , Unión ProteicaRESUMEN
The potential of six dimeric bisbenzimidazoles bound to scDNA to inhibit eukaryotic DNA topoisomerase (topo-I) was studied chemically; the tested compounds differed in linker structure and length. All the compounds inhibited topo-I, DB(7) being the most efficient; its inhibitory activity in vitro was 50-fold higher than that of camptothecin. It is noteworthy that inhibitory properties of nearly all the tested compounds increased many times if they were preincubated with scDNA for three days.
Asunto(s)
Bisbenzimidazol/farmacología , ADN-Topoisomerasas de Tipo I/metabolismo , ADN/metabolismo , Bisbenzimidazol/química , Dimerización , Unión Proteica , Inhibidores de Topoisomerasa IRESUMEN
It was found recently that Hoechst 33258, a dsDNA fluorescent dye used in cytological studies, is an efficient inhibitor of the interaction of TATA-box-binding protein with DNA, DNA topoisomerase I, and DNA helicases. In addition it proved to be a radioprotector. Biological activity of Hoechst 33258 may be associated with dsDNA complexes of not only monomeric, but also dimeric type. In this work, the Hoechst 33258 interaction with poly(dG-dC).poly(dG-dC) was studied using UV-vis and fluorescent spectroscopy, circular and flow-type linear dichroism. It was found that Hoechst 33258 formed with poly(dG-dC).poly(dG-dC) complexes of three types, namely, monomeric, dimeric, and, apparently, tetrameric, and their spectral properties were studied. Complexes of monomeric and dimeric types competed with distamycin A, a minor groove ligand, for binding to poly(dG-dC).poly(dG-dC). We proposed that Hoechst 33258 both monomers and dimers form complexes of the external type with poly(dG-dC).poly(dG-dC) from the side of the minor groove.
Asunto(s)
Bisbenzimidazol/metabolismo , ADN-Topoisomerasas de Tipo I/metabolismo , Colorantes Fluorescentes/metabolismo , Poli C/metabolismo , Poli G/metabolismo , Polinucleótidos/metabolismo , Dicroismo Circular , Dimerización , Modelos Moleculares , Conformación de Ácido Nucleico , Poli C/química , Poli G/química , Polidesoxirribonucleótidos , Espectrometría de Fluorescencia , Espectrofotometría UltravioletaRESUMEN
Dimeric bisbenzimidazole DB(5) fluorescing in the blue spectral area (lambda(em) 476 nm) within the DNA complex was synthesized. DB(5) is bound by a double-strand DNA and can differentially stain human and plant (flax) chromosomes. According to preliminary data, it provides considerably more intensely contrasting chromosome staining than DAPI and Hoechst 33258 dyes. It was also found that DB(5) is an in vitro inhibitor of HIV-1 integrase in the reaction of 3'-processing. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2008, vol. 34, no. 2; see also http://www.maik.ru.
Asunto(s)
Bencimidazoles/síntesis química , ADN/química , Colorantes Fluorescentes/síntesis química , Bencimidazoles/química , Bencimidazoles/farmacología , Bisbenzimidazol/química , Pintura Cromosómica , Cromosomas Humanos , Cromosomas de las Plantas , Dimerización , Lino , Colorantes Fluorescentes/química , Colorantes Fluorescentes/farmacología , Integrasa de VIH/química , Inhibidores de Integrasa VIH/síntesis química , Inhibidores de Integrasa VIH/química , Humanos , Indoles/química , LigandosRESUMEN
Dimeric Hoechst 33258 molecules [dimeric bisbenzimidazoles (DBBIs)] that, upon binding, occupy one turn of the B form of DNA in the narrow groove were constructed by computer simulation. Three fluorescent DBBIs were synthesized; they consist of two bisbenzimidazole units tail-to-tail linked to phenolic hydroxy groups via penta- or heptamethylene or tri(ethylene glycol) spacers and have terminal positively charged N.N-dimethylaminopropyl carboxamide groups in the molecule. The absorption spectra of the DBBIs in the presence of different DNA concentrations showed a hypochromic effect and a small shift of the absorption band to longer wavelengths, which indicated the formation of a complex with DNA. The presence of an isobestic point in the spectrum indicates the formation of one type of DBBI-DNA complexes. The interaction of DBBIs with DNA was studied by CD using a cholesteric liquid-crystalline dispersion (CLD) of DNA. The appearance of a positive band in the absorption region of ligand chromophores in the CD spectrum of the DNA CLD indicates the formation of a DBBI-DNA complex in which ligand chromophores are arranged at an angle close to 54 degrees relative to the helix axis of DNA, which suggests the localization of the DBBI in the narrow groove of DNA. All the DBBIs were found to be in vitro inhibitors of HIV-1 DNA integrase in the 3'-processing reaction, and, of the three DBBIs, two dimers inhibit HIV-1 integrase even in submicromolar concentrations.
Asunto(s)
Bencimidazoles/química , ADN/química , Colorantes Fluorescentes/química , Inhibidores de Integrasa VIH/química , Integrasa de VIH/efectos de los fármacos , Secuencia de Bases , Bencimidazoles/síntesis química , Bencimidazoles/farmacología , Bisbenzimidazol/química , Simulación por Computador , Dimerización , Integrasa de VIH/química , Inhibidores de Integrasa VIH/síntesis química , Inhibidores de Integrasa VIH/farmacología , Humanos , Ligandos , Estructura MolecularRESUMEN
With the goal to design ligands recognizing extended regions on dsDNA, a covalent dimer of the fluorescent dye Hoechst 33258 [bis-HT(NMe)] composed of two dye molecules linked via the phenol oxygen atoms with a (CH2)3-N+ H(CH3)-(CH2)3 fragment was constructed using computer modeling and then synthesized. Its interactions with the double-stranded DNA (dsDNA) were studied by fluorescent and UV-Vis spectroscopy and circular (CD) and linear dichroism (LD). Based on variations in the affinity to the dsDNA, it was shown that complexes of three types are formed. The first type complexes result from binding of a bis-HT(NMe) monomer in the open conformation; in this case the ligand covers the total dsDNA turn and is located in the minor groove according to the positive value of CD at 370 nm. In addition, the ability to form bis-HT(NMe)-bridges between two dsDNA molecules, i.e., each of the two bis-HT(NMe) ends binds to two different dsDNA molecules, was demonstrated for the first type complexes. Spectral characteristics (maximal absorption at 362 nm, positive sign, and maximal value of CD at 370 nm) of the first type complexes conform to those of the specific Hoechst 33258 complex with poly[d(A-T)] x poly[d(A-T]. The second type complexes correspond to the bis-HT(NMe) sandwich (as an inter- or intramolecular) binding to dsDNA with stoichiometry > or = 5 bp. Thereby, a negative LD at 360 nm and the location of bis-HT(NMe) sandwiches in the minor groove of B form dsDNA seems contradictory. Spectral characteristics (maximal positive CD at 345 nm, a dramatic decrease in fluorescence intensity and the shift of its maximum to 490 nm) of these complexes favor a suggestion that this binding correlates to the formation of nonspecific dimeric Hoechst 33258 complex with dsDNA. The third type complexes are characterized by stoichiometry of one bis-HT(NMe) molecule per approximately 2 bp and the tendency to zero of LD values at 270 and 360 nm. We assume that in these complexes bis-HT(NMe) sandwich dimers are formed on dsDNA. The complexes of this type conform to the aggregation type complex of Hoechst 33258 with dsDNA. The ability of bis-HT(NMe) to cover the whole dsDNA turn or form bridges with two dsDNA upon the formation of the first type complexes essentially distinguishes it from Hoechst 33258, which can only occupy 5 bp and does not form such bridges. This specific property of bis-HT(NMe) may support new biological activities.
Asunto(s)
Bisbenzimidazol/química , ADN/química , Secuencia de Bases , Sitios de Unión , Dicroismo Circular , Dimerización , Colorantes Fluorescentes , Cinética , Modelos Moleculares , Conformación de Ácido Nucleico , Oligodesoxirribonucleótidos , Espectrometría de Fluorescencia , EspectrofotometríaRESUMEN
We synthesized dimeric Hoechst dye molecules composed of two moieties of the Hoechst 33258 fluorescent dye phenolic hydroxy groups of which were tethered via pentamethylene, heptamethylene, or triethylene oxide linkers. A characteristic pattern of differential staining of chromosome preparations from human premonocytic leukemia HL60 cells was observed for all the three fluorescent dyes. The most contrast pattern was obtained for the bis-Hoechst analogue with the heptamethylene linker; its quality was comparable with the picture obtained in the case of chromosome staining with 4',6-diamidino-2-phenylindole. The ability to penetrate into the live human fibroblasts was studied for the three bis-Hoechst compounds. The fluorescence intensity of nuclei of live and fixed cells stained with the penta- and heptamethylene-linked bis-Hoechst analogues was found to differ only slightly, whereas the fluorescence of the nuclei of live cells stained with triethylene oxide-linked bis-Hoechst was considerably weaker than that of the fixed cells. The bis-Hoechst molecules are new promising fluorescent dyes that can both differentially stain chromosome preparations and penetrate through cell and nuclear membranes and effectively stain cell nuclei.
Asunto(s)
Bisbenzimidazol/síntesis química , ADN/química , Colorantes Fluorescentes/síntesis química , Emparejamiento Base , Sitios de Unión , Bisbenzimidazol/química , Cromosomas Humanos/ultraestructura , Fibroblastos/metabolismo , Fibroblastos/ultraestructura , Colorantes Fluorescentes/química , Células HL-60 , Humanos , LigandosRESUMEN
A dimeric analogue of the fluorescent dye Hoechst 33258 was synthesized. It was shown to differentially stain human chromosome preparations and bind to double-stranded DNAs.
Asunto(s)
Bencimidazoles/química , ADN/química , Colorantes Fluorescentes/síntesis química , Cromosomas Humanos/ultraestructura , Dimerización , Colorantes Fluorescentes/química , HumanosRESUMEN
The kinetics of the hydrolysis of the lactone ring of a hybrid molecule containing the molecules of the antitumor drug camptothecin and a derivative of the antibiotic netropsines, which is highly affine and specific to the DNA A-T sequences was investigated. It was shown that intramolecular interaction significantly slows down the rate of hydrolysis but does not change the equilibrium ratio of concentrations of the lactone and carboxylate forms of the camptothecin fragment of the hybrid molecule, which corresponds to the pH value. The use of intramolecular interaction for controlling the kinetics of the lactone/carboxylate transition makes it possible to create the drugs of the camptothecin family, which preserve the biologically active lactone form under the physiological conditions for a longer time and, therefore, are more effective as anticancer agents.
Asunto(s)
Camptotecina/química , Ácidos Carboxílicos/química , Lactonas/química , Netropsina/química , Hidrólisis , CinéticaRESUMEN
Temperature, concentration, and time dependence for the emergence of breaks in the sugar-phosphate backbone in a circular supercoiled DNA (scDNA) was studied for the first time in the presence of topotecan (TPT) and in the absence of human DNA topoisomerase I (topo I). Because TPT is a comptothecin (CPT) derivative, it is the first example for the ability of molecules of CPT family to cause double-stranded breaks in scDNA in the absence of the enzyme. The experiments were carried out in low ionic strength solutions (10 mM sodium cacodylate) at neutral pH (6.8). Incubation time necessary for the appearance of double-stranded breaks in scDNA in the presence of TPT correlated with the time of formation of strong TPT-DNA complex. A model was suggested for the complex composed of two crossed DNA duplexes bound through a bridge of two dimers of TPT lactone form. According to this model, two carbonyl groups of D rings of different TPT dimers form hydrogen bonds with 2-amino groups of guanines located in the neighboring base pairs of diverse strands of one of DNA duplexes. At the same time, two other carbonyl groups of D rings of TPT dimers form hydrogen bonds with 2-amino groups of guanines spaced five bp apart in the same strand of the second DNA duplex.
Asunto(s)
Daño del ADN , Desoxirribonucleótidos/metabolismo , Inhibidores Enzimáticos/metabolismo , Inhibidores de Topoisomerasa I , Topotecan/metabolismo , Inhibidores Enzimáticos/farmacología , Temperatura , Topotecan/farmacologíaRESUMEN
Interaction of topotecan (TPT) with synthetic double-stranded polydeoxyribonucleotides has been studied in solutions of low ionic strength at pH = 6.8 by linear flow dichroism (LD), circular dichroism (CD), UV-Vis absorption and Raman spectroscopy. The complexes of TPT with poly(dG-dC).poly(dG-dC), poly(dG).poly(dC), poly(dA-dC).poly(dG-dT), poly(dA).poly(dT) and previously studied by us complexes of TPT with calf thymus DNA and coliphage T4 DNA have been shown to have negative LD in the long-wavelength absorption band of TPT, whereas the complex of TPT with poly(dA-dT).poly(dA-dT) has positive LD in this absorption band of TPT. Thus, there are two different types of TPT complexes with the polymers. TPT has been established to bind preferably to GC base pairs because its affinity to the polymers of different GC composition decreases in the following order: poly(dG-dC).poly(dG-dC) > poly(dG).poly(dC) > poly(dA-dC).poly(dG-dT) > poly(dA).poly(dT). The presence of DNA has been shown to shift monomer-dimer equilibrium in TPT solutions toward dimer formation. Several duplexes of the synthetic polynucleotides bound together by the bridges of TPT dimers may participate in the formation of the studied type of TPT-polynucleotide complexes. Molecular models of TPT complex with linear and ring supercoiled DNAs and with deoxyguanosine have been considered. TPT (and presumably all camptothecin family) proved to be a representative of a new class of DNA-specific ligands whose biological action is associated with formation of dimeric bridges between two DNA duplexes.
Asunto(s)
ADN/química , ADN/metabolismo , Inhibidores Enzimáticos/metabolismo , Polidesoxirribonucleótidos/química , Polidesoxirribonucleótidos/metabolismo , Topotecan/metabolismo , Emparejamiento Base , Sitios de Unión , Dicroismo Circular , Modelos Moleculares , Conformación de Ácido Nucleico , Concentración Osmolar , Soluciones , Espectrometría Raman , Inhibidores de Topoisomerasa I , Topotecan/químicaRESUMEN
Interaction of topotecan (TPT) with calf thymus DNA, coliphage T4 DNA, and poly(dG-dC). poly(dG-dC) was studied by optical (linear flow dichroism, UV-vis spectroscopy) and quantum chemical methods. The linear dichroism (LD) signal of TPT bound to DNA was shown to have positive sign in the range 260-295 nm. This means that the plane of quinoline fragment (rings A and B) of TPT molecule form an angle lower 54 degrees with the long axis of DNA, and hence TPT molecule can not intercalate between DNA base pairs. TPT was established to bind to calf thymus DNA as readily as to coliphage T4 DNA whose all cytosines in the major groove were glycosylated at the 5th position. Consequently, the DNA major groove does not participate in TPT binding. TPT molecule was shown to compete with distamycin for binding sites in the minor groove of DNA and poly(dG-dC). poly(dG-dC). Thus, it was demonstrated for the first time that TPT binds to DNA at its minor groove.