Distinguishing High from Low Concentrations of RNA Cancer Marker: Five-Input DNAzyme Thresholding Gate

RNA-cleaving DNAzymes are promising agents for gene therapy that can overcome the low selectivity of other therapeutic nucleic acid agents, including CRISPR/Cas9, antisense oligonucleotides, and RNA interference agents, in protein-independent RNA cleavage. To achieve this, DNAzyme gene therapy agents should accurately distinguish cancer from healthy cells. Often, cancer RNA markers are expressed in healthy tissue at lower concentrations. Previously, we developed a DNAzyme thresholding gate (DTh), which must bind to three cancer marker molecules prior to cleaving the RNA target. This article presents the next step in the development of DTh by converting a signal from five microRNA inputs into an output cleavage of the RNA model using an advanced system of molecular logic. We demonstrate that it is possible to broaden the available threshold range by increasing the number of marker recognition modules. The threshold gates can be easily adjusted to cleave a broad range of RNA targets. This approach can be used in cancer diagnostics as well as the computational component of DNA nanorobots, autonomous DNA devices for anti-cancer gene therapy. Graphic Abstract: We have developed a 5-input deoxyribozyme-based Threshold from a binary construct, allowing to be activated after reaching a certain concentration window. This construct can easily discriminate between high and low concentrations of activator.image

Automated summary

This article presents a new method for RNA cleavage using DNAzyme threshold gates, which can accurately distinguish cancer cells from healthy cells by increasing the number of marker recognition modules. It has potential applications in cancer diagnostics and DNA nanorobots.