<i>In situ</i> formation of transcriptional modulators using non-canonical DNA i-motifs

Abstract

Non-canonical DNA i-motifs and G-quadruplexes are postulated as genetic switches for the transcriptional regulation of proto-oncogenes. However, in comparison to G-quadruplexes, the therapeutic potential of i-motifs is less explored. The development of i-motif selective ligands by conventional approaches is challenging due to the structural complexity of i-motifs. The target guided synthetic (TGS) approach involving <i>in situ</i> cycloaddition could provide specific ligands for these dynamic DNA structures. Herein, we have used i-motif forming C-rich DNA and their complementary G-quadruplex forming DNA sequences of <i>c-MYC</i> and <i>BCL2</i> promoter regions as well as a control self-complementary duplex DNA sequence as the templates to generate selective ligands from a pool of reactive azide-alkyne building blocks. In our approach, thiolated DNA targets are immobilized on the surface of gold-coated iron nanoparticles to enable efficient isolation of the newly generated ligands from the solution mixture by simple magnetic decantation. The combinatorial <i>in situ</i> cycloaddition generated cell-membrane permeable triazole leads for respective DNA targets (<i>c-MYC</i> and <i>BCL2</i> i-motifs and G-quadruplexes) that selectively promote their formation. <i>In vitro</i> cellular studies reveal that the <i>c-MYC</i> i-motif and G-quadruplex leads downregulate <i>c-MYC</i> gene expression whereas the <i>BCL2</i> i-motif lead upregulates and the <i>BCL2</i> G-quadruplex lead represses <i>BCL2</i> gene expression. The TGS strategy using i-motif DNA nanotemplates represents a promising platform for the direct <i>in situ</i> formation of i-motif specific ligands for therapeutic intervention.