C3-Spacer-containing circular oligonucleotides as inhibitors of human topoisomerase I

Some dumbbell-shaped circular oligonucleotides containing internal C3-spacers and Topo I-binding sites were designed and synthesized which displayed high inhibitory efficiency on the activity of human Topo I as well as resisted the degradation by some DNA repair enzymes.     

Nick-containing oligonucleotides as human topoisomerase I inhibitors

A series of oligonucleotides with various lengths that contain nick and topoisomerase I-binding sites were designed. The interactions between these oligonucleotides and human topoisomerase I were investigated and the most efficient one among them has displayed IC₅₀ value of 6.3 nM. Our studies have also demonstrated that the position of the nick as well as the length of the oligonucleotides were crucial factors for the inhibition of this nuclear enzyme.     

Quadruplex-duplex motifs as new topoisomerase I inhibitors

In this article, 13 short chains that can form G-quadruplex and quadruplex-duplex motif have been designed. Fourteen oligonucleotides, including 13 short chains as well as a reference short chain all have certain level of inhibition to topoisomerase I, whether or not they form G-quadruplex and quadruplex-duplex motif, and the G-quadruplex and quadruplex-duplex motif show better activity than single short chain. The result confirmed that after forming G-quadruplex and quadruplex-duplex motif these 14 oligonucleotides are competitive inhibition, that is, through the priority binding with the topoisomerase I and precluding from its binding with the normal substrate pBR322 and, therefore, inhibiting the next reaction.     

3-Arylisoquinolines as novel topoisomerase I inhibitors

Topoisomerase I (topo I) is an essential enzyme for vital cellular processes. Inhibition of topo I activities is lethal and leads to cell death, thus establishing topo I as a promising target for cancer treatment. Camptothecin, a natural alkaloid, inhibits topo I. Topotecan and irinotecan, synthetic derivatives of camptothecin, are the most potent anticancer drugs in clinical use. However, several limitations of camptothecins such as solubility, toxicity, stability, resistance and the required high drug dose have encouraged the development of non-camptothecin topo I inhibitors. Natural alkaloid benzo[c]phenanthridines and synthetic indenoisoquinolines have been extensively studied as alternatives to camptothecin. Interestingly, these non-camptothecin topo I inhibitors share a common 3-arylisoquinoline scaffold. This review will describe the development of novel indeno[1,2-c]isoquinolines, isoindolo[2,1-b]isoquinolines, 12-oxobenzo[c]phenanthridines and benz[b]oxepines with a 3-arylisoquinoline nucleus as topo I inhibitors.     

Heterocyclic bibenzimidazole derivatives as topoisomerase I inhibitors

A series of 2'-heterocyclic derivatives of 5-phenyl-2,5'-1H-bibenzimidazoles were evaluated for topoisomerase I poisoning activity and cytotoxicity. Topo I poisoning activity was associated with 2'-derivatives that possessed a hydrogen atom capable of hydrogen bond formation, suggesting that the interatomic distances between such hydrogen atoms and the heteroatoms on the adjacent benzimidazole influence activity.     

Synthesis of nitrated indenoisoquinolines as topoisomerase I inhibitors

Indenoisoquinolines and dihydroindenoisoquinolines have been synthesized possessing a nitro-substituted isoquinoline ring in an effort to explore the effects of electron-withdrawing substituents on biological activity. The in vitro anticancer activities of these molecules have been tested in the National Cancer Institute's screen of 55 cell lines. The compounds have also been tested for topoisomerase I (top1) inhibition. The results indicate that these substances are a potent class of top1 inhibitors with sub-micromolar cytotoxicity mean graph midpoints (MGM) and top1 inhibition equal to camptothecin.     

1H-Benzimidazole derivatives as mammalian DNA topoisomerase I inhibitors

Benzimidazole is one of the most important heterocyclic groups manifesting various biological properties, such as antibacterial, antifungal, antimicrobial, antiprotozoal and antihelmintic activities. Several benzimidazole derivatives are also active as inhibitors of type I DNA topoisomerases. In this study, three 1H-benzimidazole derivatives with different electronic characteristics at position 5-, namely 5-chloro-4-(1H-benzimidazole-2-yl)phenol (Cpd I), 5-methyl-4-(1H-benzimidazole-2-yl)phenol (Cpd II) and 4-(1H-benzimidazole-2-yl)phenol (Cpd III), were synthesized and evaluated for their effects on mammalian type I DNA topoisomerase activity using quantitative in vitro plasmid supercoil relaxation assays. For the structure elucidation of the compounds, melting points, UV, IR, 1H NMR, 13C NMR, mass spectral data and elemental analyses were interpreted. Among the compounds, 5-methyl-4-(1H-benzimidazole-2-yl)phenol (Cpd II) manifested relatively potent topoisomerase I inhibition.     

Inhibition of Mycobacterium smegmatis topoisomerase I by specific oligonucleotides

DNA topoisomerase I from Mycobacterium smegmatis unlike many other type I topoisomerases is a site specific DNA binding protein. We have investigated the sequence specific DNA binding characteristics of the enzyme using specific oligonucleotides of varied length. DNA binding, oligonucleotide competition and covalent complex assays show that the substrate length requirement for interaction is much longer ( approximately 20 nucleotides) in contrast to short length substrates (eight nucleotides) reported for Escherichia coli topoisomerase I and III. P1 nuclease and KMnO(4) footprinting experiments indicate a large protected region spanning about 20 nucleotides upstream and 2-3 nucleotides downstream of the cleavage site. Binding characteristics indicate that the enzyme interacts efficiently with both single-stranded and double-stranded substrates containing strong topoisomerase I sites (STS), a unique property not shared by any other type I topoisomerase. The oligonucleotides containing STS effectively inhibit the M. smegmatis topoisomerase I DNA relaxation activity.     

A series of enediynes as novel inhibitors of topoisomerase I.

A series of acyclic enediynes, 2-((6-substituted)-3-hexen-1,5-diynyl)benzonitriles (8--11), display potent inhibition against topoisomerase I without the formation of active biradical intermediates and show inhibitory activity against topoisomerase I at 10 microM, which is five times that of camptothecin from the results of agarose gel electrophoresis.     

Synthesis and biological evaluations of novel indenoisoquinolines as topoisomerase I inhibitors

A series of novel indenoisoquinoline derivatives were synthesized. The anticancer activities of these molecules were tested in human cancer cell lines A549, HepG2, and HCT-116. These compounds were also tested for their activity of topoisomerase I (top1) inhibition. Among them, compound 25 was found to be 10-times more potent in cell-killing activity for both cell lines HepG2 and HCT-116 than reported compound 11, with IC(50) of 0.019 and 0.093μM, respectively. Compound 25 was also found to have stronger top1 inhibition activity than 11 in our inhibition assay. Further in vivo evaluations of compound 25 are in progress and will be reported in due course.     

Some benzoxazoles and benzimidazoles as DNA topoisomerase I and II inhibitors

Some novel fused heterocyclic compounds of 2, 5-disubstituted-benzoxazole and benzimidazole derivatives, which were previously synthesized by our group, were investigated for their inhibitory activity on both eukaryotic DNA topoisomerase I and II in a cell free system. 2-Phenoxymethylbenzimidazole (17), 5-amino-2-(p-fluorophenyl)benzoxazole (3), 5-amino-2-(p-bromophenyl)benzoxazole (5), 5-nitro-2-phenoxymethyl-benzimidazole (18), 2-(p-chlorobenzyl)benzoxazole (10) and 5-amino-2-phenylbenzoxazole (2) were found to be more potent as eukaryotic DNA topoisomerase I poisons than the reference drug camptothecin having IC(50) values of 14.1, 132.3, 134.1, 248, 443.5, and 495 microM, respectively. 5-Chloro-2-(p-methylphenyl)benzoxazole (4), 2-(p-nitrobenzyl)benzoxazole (6) and 5-nitro-2-(p-nitrobenzyl)benzoxazole (8) exhibited significant activity as eukaryotic DNA topoisomerase II inhibitors, having IC(50) values of 22.3, 17.4, 91.41 microM, respectively, showing higher potency than the reference drug etoposide.     

Oxiranylmethyloxy or thiiranylmethyloxy-azaxanthones and -acridone analogues as potential topoisomerase I inhibitors

A total of seven new oxyranylmethyloxy or thiiranylmethyloxy group substituted 5-azaxanthones and -acridones analogues were synthesized and tested for their biological activities for cancer cell lines and topoisomerases. Among the compounds, compound 5, 3-thiiranylmethyloxy-1-hydroxy-5-azaxanthone, showed effective topoisomerase I inhibitory activity, 50% and 27% inhibition ratio at 100 and 20 μM, respectively. This result is the first finding of the function of 5-azaxanthone compounds for topoisomerase I inhibition and can provide a novel skeleton for the anticancer drug development process.     

DNA topoisomerase I inhibitors from Rinorea anguifera

An organic extract prepared from Rinorea anguifera was investigated in order to identify the natural principle(s) responsible for stabilization of a topoisomerase I-DNA covalent binary complex. Bioassay-guided fractionation resulted in the isolation of mauritianin and (+)-syringaresinol as new topoisomerase I inhibitors, and also of the known inhibitor camptothecin.     

Quinoline alkaloids as intercalative topoisomerase inhibitors

Quinoline alkaloids are abundant in the Rutaceae, and many have exhibited cytotoxic activity. Because structurally related antitumor alkaloids such as camptothecin and fagaronine are known to function as intercalative topoisomerase poisons, it is hypothesized that cytotoxic Stauranthus alkaloids may also serve as intercalative topoisomerase inhibitors. To test this hypothesis theoretically, ten Stauranthus quinoline alkaloids were examined for potential intercalation into DNA using a molecular docking approach. Four of the alkaloids (stauranthine, skimmianine, 3′,6′-dihydroxy-3′,6′-dihydrostauranthine, and trans-3′,4′-dihydroxy-3′,4′-dihydrostauranthine) were able to intercalatively dock consistently into DNA. In order to probe the intermolecular interactions that may be responsible for intercalation of these quinoline alkaloids, density functional calculations have been carried out using both the B3LYP and M06 functionals. M06 calculations indicated favorable π–π interactions between either skimmianine or stauranthine and the guanine–cytosine base pair. Furthermore, the lowest-energy face-to-face orientation of stauranthine with guanine is consistent with favorable dipole–dipole orientations, favorable electrostatic interactions, and favorable frontier molecular orbital interactions. Likewise, the lowest-energy face-to-face orientation of stauranthine with the guanine–cytosine base pair reveals favorable electrostatic interactions as well as frontier molecular orbital interactions. Thus, not only can quinoline alkaloids dock intercalatively into DNA, but the docked orientations are also electronically favorable.      

Synthesis and biological evaluation of 7-alkenyl homocamptothecins as potent topoisomerase I inhibitors

Homocamptothecin (hCPT) is a camptothecin (CPT) derivative with a seven-membered β-hydroxylactone E ring, which shows higher lactone stability and improves topoisomerase I (Topo I) inhibition activity. In an attempt to improve the antitumor activity of homocamptothecins, a series of 7-alkenyl-homocamptothecin derivatives was designed and synthesized based on a semisynthetic route starting from CPT. Most of the synthesized compounds exhibit higher cytotoxic activities on the A-549 tumor cell line than topotecan (TPT). Some compounds such as 2a and 2o show a broad in vitro antitumor spectrum and exhibit superior Topo I-inhibition activity.     

Natural inhibitors of DNA topoisomerase I with cytotoxicities

DNA Topoisomerase I can cause DNA breaks and play a key role during cell proliferation and differentiation. It is an important target for anticancer agents. While screening for anticancer compounds, seven natural compounds, 1-7, showed potent cytotoxicities against a panel of ten cancer cell lines. Moreover, an inhibition assay demonstrated that they are also DNA topoisomerase I inhibitors, in which inhibitors 1-5 are new ones.     

Solid phase synthesis of anthraquinone peptides and their evaluation as topoisomerase I inhibitors.

Anthraquinone peptide derivatives have previously been shown to inhibit the enzyme topoisomerase I (topo I), a pharmaceutical target for the prevention of malignant carcinomas. A highly efficient procedure for the attachment of the anthraquinone moiety to the N-terminus of a peptide on a solid support is reported. This methodology provides a convenient method for the synthesis of labelled peptides, with potential applications for chemotherapy, DNA detection and protein purification. As the synthetic strategy utilizes the solid phase, it should also be amenable to the generation of combinatorial libraries. The utility of the method by synthesizing a pool of peptides and assaying for topo I inhibition is demonstrated.     

DNA topoisomerase I from human placenta

In this study we report that human placenta is an excellent source of DNA topoisomerase I. The enzyme can be isolated in the fully intact 100 kDa form although lower molecular mass species are also observed. Occasionally, the enzyme can be resolved into two peaks of activity by chromatography on phosphocellulose. As expected, the enzyme promotes marked cleavage of DNA in response to the anticancer drug camptothecin. Because of this property and the ready availability of human placenta, the enzyme should prove to be useful in the development and testing of new anticancer drugs that target topoisomerase I.     

Flavonoids as DNA topoisomerase I poisons

The therapeutic anticancer potential of flavonoids shown by recent research needs a greater understanding of these compounds. They are antioxidants and antimutagenic agents that can inhibit tumor promotion and transformation and can modify the activity of a large number of mammalian enzyme systems, such as human DNA-topoisomerases. Poisons of topoisomerases generate toxic DNA damage by stabilization of the covalent DNA-topoisomerase cleavage complex and some of them have therapeutic efficacy in human cancer. The present investigation has assayed ten flavonoids, isolated in our laboratory, as topoisomerase I poisons obtaining myricetin and myricetin-3-galactoside as two new topoiosomerase I poisons. These two flavonoids, and the plant extract from which they were isolated, were assayed for cytotoxic activity against three human cancer cell lines using the SRB assay. Taking into account our previous research, structural requisites implicated in the topoisomerase poisoning are discussed.