Drawing inspiration from natural cell signaling cascades, chemists at Université de Montréal (UdeM) have developed a DNA-based signaling cascade that allows them to quantify and report via an easily measurable electrochemical signal the concentration of various molecules in a drop of blood, all within five minutes.
The approach developed by the Vallée-Bélisle team might aid efforts to build point-of-care devices for monitoring and optimizing the treatment of various diseases, the team suggested. The team, headed by UdeM chemistry professor Alexis Vallée-Bélisle, PhD, reported on their findings, validated by experiments in mice, in the Journal of the American Chemical Society, in a paper titled “Kinetically Programmed Signaling Cascades for Molecular Detection.” In their paper, they stated, “Notably, the cascade operated directly in a 5 μL drop of unprocessed blood using a simple one-step, 5 min procedure, eliminating the need for separation, washing, or purification steps typically required by centralized laboratory methods.”
The breakthrough with the new technology came from observing how cells detect and quantify the concentration of molecules in their surroundings. Vallée-Belisle, who is a holder of a Canada Research Chair in Bioengineering and Bio-nanotechnology, has spent many years exploring how biological systems monitor the concentration of molecules in their surroundings in real time.
Signaling cascades represent a key biochemical mechanism, the authors explained. “These finely tuned molecular network reactions, which typically display modular and hierarchical architectures including input, receptor, processor, and output modules, have evolved to detect a wide array of chemical inputs to control and optimize cell activity, division, and differentiation.” Such molecular networks have also inspired the development of strategies in areas including synthetic biology, molecular computing, and drug delivery, the team continued.
“One of the key factors in successfully treating various diseases is to provide and maintain a therapeutic drug dosage throughout treatment,” said Vallée-Bélisle. “Sub-optimal therapeutic exposure reduces efficiency and typically leads to drug resistance, while overexposure increases side effects.”
However, maintaining the right concentration of drugs in the blood remains a major challenge in modern medicine. Since each patient has a distinct pharmacokinetic profile, the concentration of medications in their blood varies significantly. In chemotherapy, for example, many cancer patients do not get an optimal dosage of drugs, and few or no tests are currently rapid enough to flag this issue.
The sensing principle of the newly developed sensors is straightforward. The molecular target or drug to be monitored can interact with a specific DNA molecule, called an aptamer. Upon binding to the molecular target, the aptamer DNA can no longer inhibit another electro-active DNA, which can then reach the surface of an electrode and generate an electrochemical current that is easily detectable with an inexpensive reader.
“Cells have developed nanoscale ‘signaling cascades’ made of biomolecules that are programmed to interact together to activate specific cellular activities in the presence of a specific amount of external stimuli or molecules,” commented first author Guichi Zhu, PhD, a postdoctoral fellow at UdeM. “Inspired by the modularity of nature’s signaling systems and by the ease with which they can evolve to detect novel molecular targets, we have developed similar DNA-based signaling cascades that can detect and quantify specific molecules via the generation of an easy, measurable electrochemical signal.”
The scientists further explained, “We designed our DNA-based signaling cascade by drawing inspiration from the modularity and hierarchical arrangement found in intracellular signaling cascades, which are typically composed of input, receptor, processor, and output modules …”
This novel signaling mechanism produces sufficient change in electrical current to be measured using inexpensive electronics similar to those in the home glucose meters used by diabetics to check their blood sugar. “Using this DNA-based assay, we have been able to develop sensors for multiple blood molecules even if their concentration was sometimes less than 100,000 times less concentrated than glucose,” said co-author Bal-Ram Adhikari, PhD, another UdeM postdoctoral fellow.
To illustrate how this novel signaling mechanism can be adapted into an easy-to-use home test to help patients monitor and optimize their chemical therapy, the authors demonstrated the real-time monitoring of the anti-malaria drug, quinine, in living mice. The current gold-standard tests employed to do so typically require hours of procedures and an expensive instrumental setting. “Convenient strategies enabling the monitoring of quinine and other antimalarial drugs at home would represent a significant breakthrough to improve the efficacy and effectiveness of these treatments,” they commented in their paper.
“A great advantage of these DNA-based electrochemical tests is that their sensing principle can also be generalized to many different targets, allowing us to build inexpensive devices that could detect many different molecules in five minutes in the doctor’s office or even at home,” said Vallée-Bélisle, whose team validated their novel mechanism by developing additional sensors for detecting ATP, thrombin, and platelet-derived growth factor. “… ATP molecules, thrombin, and PDGF were all successfully detected using a 5 min reaction time,” the team wrote when reporting on their experiments.
“Overall, these results show that the kinetically programmed signaling cascades perform well in a one-pot format, which allows a simple one-step workflow that can be performed by untrained users.” The technology could have broad applications, the investigators concluded. “We believe that similar kinetically programmed signaling cascades could be developed for a wide range of chemical applications, allowing complex, multistep workflows to be streamlined into a rapid, single-step reaction.
A patent for the invention has been licensed by the Montreal-based company Anasens in order to expedite its commercialization.
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