Using DNA devices to track anticancer drug activity

Abstract

It is beneficial to develop systems that reproduce complex reactions of biological systems while maintaining control over specific factors involved in such processes. We demonstrated a DNA device for following the repair of DNA damage produced by a redox-cycling anticancer drug, beta-lapachone (β-lap). These chips supported ß-lap-induced biological redox cycle and tracked subsequent DNA damage repair activity with redox-modified DNA monolayers on gold. We observed drug-specific changes in square wave voltammetry from these chips at therapeutic ß-lap concentrations of high statistical significance over drug-free control. We also demonstrated a high correlation of this change with the specific ß-lap-induced redox cycle using rational controls. The concentration dependence of ß-lap revealed significant signal changes at levels of high clinical significance as well as sensitivity to sub-lethal levels of ß-lap. Catalase, an enzyme decomposing peroxide, was found to suppress DNA damage at a NQO1/catalase ratio found in healthy cells, but was clearly overcome at a higher NQO1/catalase ratio consistent with cancer cells. We found that it was necessary to reproduce key features of the cellular environment to observe this activity. Thus, this chip-based platform enabled tracking of ß-lap-induced DNA damage repair when biological criteria were met, providing a unique synthetic platform for uncovering activity normally confined to inside cells.

Following anticancer drug activity in cell lysates with DNA devices

“…This lower signal change is anticipated as the lower NQO1 concentration cannot catalyse the same degree of hydrogen peroxide production from β-lap (see Figure 3d), and hence lower damage is produced. This 370% signal change difference and statistical confidence is significantly better than the ~70% signal change difference that we previously observed in buffer solution for NQO1 free controls (Kahanda et al 2016). This is likely owed to the greater diversity of repair proteins and higher enzymatic turnover in the lysates versus the single repair protein and limited enzymes used in buffer solution.…”

“…We added both the cytoplasmic lysate and nuclear lysate directly to the chip, then observed the signal change upon addition of the β-lap was very small, within zero in consideration of the standard error. The reason for this is not fully clear, but some inferences can be drawn from our previous study in buffer mixtures on chips (Kahanda et al 2016). In these experiments, only NQO1, NADH, iron, transferrin, formamidopyrimidine DNA glycosylase (FPG, a repair protein), and β-lap were introduced to the chip.…”

“…In these devices, DNA monolayers on electrodes can succumb to damage by reactive oxygen species like H 2 O 2 . In turn, this damage can be accessed by repair proteins in the surrounding solution (Kahanda et al 2016). These DNA monolayers thus function as a natural recognition element of biological DNA damage repair.…”

“…Previously, we demonstrated that these electrochemical DNA devices could be used to track β-lap activity in buffer solutions containing NQO1 and various cofactors for drug activity (Kahanda et al 2016). However, cellular samples are considerably more complex, with competitive DNA binding proteins, additional protein-protein interactions, and competing biochemical reactions and pathways, all of which are important to consider if these devices are to be used for practical medical applications or biological assays.…”

Electrochemical Sensor Based on Poly(Azure B)-DNA Composite for Doxorubicin Determination

“…DNA sensors have found increasing attention during the past decade for the fast and sensitive determination of complementary sequences such as hybridization events [1] and antitumor drugs [2,3], as well as for the detection of DNA damage caused by physical [4] and chemical [5,6,7] factors. The interest in DNA sensors has increased due to its advantages, including compatibility with conventional measurement equipment, low-cost, possibility to use for point-of-care diagnostics, in-flow analysis mode, and intuitive and understandable design and data interpretation.…”