| A noncanonical secondary DNA structure,named G-quadruplex DNA,is formed from G-rich sequences under physiological conditions and is widespread in biologically important regions of the human genome.A great deal of research has been reported on its potential role as a telomerase inhibitor,which makes it a potential target for the development of new anticancer therapies.However,a great deal DNA sequences within the human genome have the potential to form G-quadruplex structures,which makes it very important to selectively target the specific conformation of the G-quadruplex.Currently,a small amount of multimeric G-quadruplexes DNA and RNA were only found in telomeric regions and non-coding region of C9orf72 gene.Therefore,it is very significant to design binders or fluorescent probes that selectively recognize and bind multimeric G-quadruplexes towards the development of potential anticancer agents.In this paper,we designed two kinds of compounds to selectively recognize and bind multimeric G-quadruplexes of the human telomere.In the first part of the work,three new polyether-tethered bisquinolinium dimers(1a-c)were synthesized and studied their binding affinities,selectivities and thermal stabilization towards dimeric G-quadruplex DNA(G2T1)in human telomeric regions.Bisquinolinium dimer with the medium polyether linker(1b)showed 30-425-fold higher binding affinity and selectivity towards antiparallel G2T1 over monomeric quadruplexes including human monomeric G-quadruplexes(G1),c-kit 1,c-kit 2 and c-myc,respectively.In addition,compound lb induced the formation of quadruplexes and displayed highest thermal stabilization(?Tm>28.1?)among all reported multimeric G-quadruplex binders.Compound lb also displayed higher selectivities towards antiparallel G2T1 than monomer 360A and bisquinolinium dimers la and lc.These results show that polyether linkers in these dimeric G-quadruplex binders play a crucial role in regulating the binding affinities,selectivities and thermal stabilization towards G2T1.The possible binding mode is that two 360A moieties in compound lb might stack with the two adjacent G-quadruplex units in G2T1.In addition,compound lb with high selectivity towards antiparallel G2T1 also showed strong telomerase inhibition(IC50 = 5.0 ± 0.2 ?M)and potent anticancer activities,especially against Hela cells(IC50 = 9.5±2.0 ?m).In the second part of the work,two berberine-bisquinolinium conjugates Ber-360A(2a)and Ber-PDS(2b)were synthesized and evaluated their abilities of fluorescence response,binding affinities,selectivities,and stabilization towards human telomeric G2T1.Compounds 2a-b showed stronger fluorescence response,higher binding affinity(Ka>107 M-1)and selectivity towards Anti-G2T1 than towards other DNA sequences.Notably,compound 2b showed stronger fluorescence response and higher sensitivity with a detection limit of 0.44 nM than compound 2a.In addition,compounds 2a-b induced the formation of quadruplexes and displayed the highest level of thermal stabilization(?Tm>20 ?)towards Anti-G2T1 among all DNA sequences.The binding mode is that both Ber-360A and Ber-PDS interacted with the two adjacent G-quartet in G2T1,but had weaker binding towards the tetrad under Ap7 and Ap19 bases than other three tetrads in G2T1.Interestingly,cell imaging indicate that two conjugates could enter into the cells,especially conjugate Ber-PDS could enter into the nucleoli and fluorescently detect rDNA G-quadruplexes.In addition,compounds 2a-b showed strong telomerase inhibition(0.92± 0.14?M,1.34± 0.19?M,respectively)and potent anticancer activities.In summary,we have found a class of excellent binders and a class of excellent fluorescent probes by conjugating fluorescent ligand and G-quadruplex binder towards dimeric G-quadruplexes.These research provides new insights for the development of special recognition binders and anticancer drugs targeting telomeric dimeric G-quadruplexes. |
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