No abstract

Some G-quadruplex-hemin complexes can be used as peroxidase-mimicking DNAzymes, catalyzing H(2)O(2)-mediated reactions such as the oxidation of 2,2'-azinobis (3-ethylbenzothiozoline)-6-sulfonic acid (ABTS) by H(2)O(2). However, some challenges, for example, the relatively low catalytic activity and the disproportionation of the reaction product ABTS*(+), may seriously restrict further development and applications of these complexes. Here, we demonstrated the positive effect of adenosine triphosphate (ATP) on G-quadruplex-hemin DNAzyme-mediated catalytic reactions. The presence of ATP not only improved the catalytic activity of G-quadruplex-hemin DNAzymes, but also inhibited the disproportionation of ABTS*(+). These observations may improve the performance of existing G-quadruplex-hemin DNAzyme-based chemical sensors, for example, the Ag(+)-detection method that uses G-quadruplex-hemin DNAzymes, and widen the application range of G-quadruplex-hemin DNAzymes. We also demonstrated that the phosphate groups, nucleobase, and sugar of ATP determine the reaction-promoting ability of ATP. These observations may be helpful in the design of highly efficient enhancers for G-quadruplex-hemin DNAzymes.

Activation of human telomerase reverse transcriptase (hTERT) is necessary for limitless replication in tumorigenesis. Whereas hTERT is transcriptionally silenced in normal cells, most tumor cells reactivate hTERT expression by alleviating transcriptional repression through diverse genetic and epigenetic mechanisms. Transcription-activating hTERT promoter mutations have been found to occur at high frequencies in multiple cancer types. These mutations have been shown to form new transcription factor binding sites that drive hTERT expression, but this model cannot fully account for differences in wildtype (WT) and mutant promoter activation and has not yet enabled a selective therapeutic strategy. Here we demonstrate a novel mechanism by which promoter mutations activate hTERT transcription, which also sheds light on a unique therapeutic opportunity. Promoter mutations occur in a core promoter region that forms tertiary structures consisting of a pair of G-quadruplexes involved in transcriptional silencing.We show that promoter mutations exert a detrimental effect on the folding of one of these Gquadruplexes, resulting in a nonfunctional silencer element that alleviates transcriptional repression. We have also identified a small drug-like pharmacological chaperone (pharmacoperone) molecule, GTC365, that acts at an early step in the G-quadruplex folding pathway to redirect mutant promoter G-quadruplex misfolding, partially reinstate the correct folding pathway, and reduce hTERT activity through transcriptional repression. This transcription-mediated repression effects cancer cell death through multiple routes including both induction of apoptosis through inhibition of hTERT's role in regulating apoptosis-related proteins and inducing senescence by decreasing telomerase activity and telomere length.We demonstrate the selective therapeutic potential of this strategy in melanoma cells that overexpress hTERT.3

Some G-quadruplex-hemin complexes are DNAzyme peroxidases that efficiently catalyze H(2)O(2)-mediated reactions, such as the oxidation of ABTS (2,2'-azinobis(3-ethylbenzothiozoline)-6-sulfonic acid) by H(2)O(2). Since Ag(+) chelates guanine bases at the binding sites are involved in G-quadruplex formation, the presence of Ag(+) may disrupt these structures and inhibit the peroxidase activity of G-quadruplex-hemin DNAzymes. On the basis of this principle, a highly sensitive and selective Ag(+)-detection method was developed. The method allows simple detection of aqueous Ag(+) with a detection limit of 64 nM and a linear range of 50-3000 nM. Cysteine (Cys) is a strong Ag(+)-binder and competes with quadruplex-forming G-rich oligonucleotides for Ag(+)-binding, promoting the reformation of G-quadruplexes and increasing their peroxidase activity. Therefore, the Ag(+)-sensing system was also developed as a Cys-sensing system. This "turn-on" process allowed the detection of Cys at concentrations as low as 50 nM using a simple colorimetric technique. The Cys-sensing system could also be used for the detection of reduced glutathione (GSH). Neither the Ag(+)-sensing nor the Cys-sensing systems required labeled oligonucleotides. In addition, both gave large changes in absorbance signal that could be observed by the naked eye. Thus, a simple visual method for Ag(+)- or Cys-detection was developed.