An Electroanalytical Approach for Advancing Cancer Diagnosis and Therapy by Tracking Globally DNA G‑Quadruplex Noncanonical Structures

Abstract

This work reports the first electroanalytical methodology developed to determine global levels of G-quadruplex (G4) motifs in DNA, hitherto underestimated noncanonical structures of increasing relevance both in cancer diagnostics and precision therapy. The methodology developed takes advantage of the benefits provided by magnetic microcarriers to implement a competitive immunoassay involving a synthetic DNA sequence carrying a single G4 motif and a commercial recombinant antibody that can recognize these motifs. This antibody was enzymatically labeled with an HRP-conjugated secondary antibody, and amperometric transduction was performed using disposable electrode substrates. The careful selection of the bioassay format and the optimization of the experimental variables has led to a groundbreaking technology allowing the determination in only 10 min of synthetic sequences carrying a single G4 motif at the low nM level. The methodology was applied to the analysis of 50 ng of genomic DNA extracted from different cancer cells, highlighting its potential to accurately determine the content of such structures in these extracts and to relate their expression to the silencing of key genes or to the cell’s ability to interact with potential oncotherapeutic agents capable of stabilizing these noncanonical structures.