Celebrating DNA aptamers on DNA Day!
April 25th is DNA day. This day celebrates the completion of the Human Genome Project 20 years ago and the discovery of DNA’s double helix 70 years ago, so it’s a big anniversary for DNA!
Over this time we’ve learned so much about the power of DNA and how we can work with it and adapt it to make new breakthroughs. These advances range from paternity testing and forensics, to modifying plants or microbes for better crops or as a new source of fuel. Then of course, there’s our favourite DNA innovation – DNA aptamers.
DNA aptamers are cheaper and simpler to manufacture and more robust than their RNA counterparts. Because of this, DNA-based aptamers are often used as research reagents and affinity tools within diagnostics, though we are also starting to see DNA aptamers increase in the clinical pipeline too.
At Aptamer Group, we use both DNA and RNA aptamer libraries to deliver the best Optimer for your specific project, so we are pleased to celebrate DNA day and the contribution that DNA-based aptamers have made to science by highlighting a few of our favourite DNA aptamer innovations so far.
DNA aptamer therapies
Of the nine current aptamers in clinical trial, two are unmodified DNA molecules. DNA aptamers were not initially explored as therapeutic molecules, as RNA aptamers are more amenable to modification for longer in vivo half-lives. However, this feature of DNA aptamers has now been embraced allowing ‘hit-and-run’ strategies for aptamer therapies, where the DNA aptamer can quickly reach its target, induce an effect and be excreted or degraded. In this case the small size of the aptamer and accessibility of the DNA molecule to endogenous enzymes facilitates the desired shorter half-life.
Aptoll is a DNA aptamer therapy in phase II trials for acute ischaemic stroke and phase I trials for long COVID. This aptamer is a Toll-like receptor 4 antagonist. In cases of ischaemic stroke it acts to inhibit inflammation, reduce infarct volumes and improve functional outcomes. Trials with Aptoll have shown excellent safety profiles, and when used within 6 hours of stroke onset in combination with endovascular treatment showed reduced mortality and disability at 90 days.
BC007 is a second DNA aptamer therapy that is currently in phase II trials for heart failure . This DNA aptamer neutralises functionally active, pathological auto-antibodies directed against G protein-coupled receptors. In the body functional auto-antibodies trigger a hormone-like function through binding to cell-surface G-protein coupled receptors. Pathological versions of these auto-antibodies have been found to function in pulmonary hypertension, glaucoma, Chronic Fatigue Syndrome (CFS), long COVID and heart failure. BC007 binds to and eliminates auto-antibodies against the beta-1-adrenoceptor that regulates heart rate and contraction strength. Ongoing trials with this DNA aptamer show good patient tolerability and efficacy for heart failure.
Unlocking the proteome with DNA aptamers
The interrogation of proteomes (“proteomics”) in a highly multiplexed and efficient manner remains a coveted and challenging goal in biology and medicine. The major goal of proteomics is the identification and quantification of every single protein species in complex biological mixtures. This will require the ability to detect and monitor all proteins and their post-translational modifications. While there are an estimated 20,000 genes in the genome this translates to an estimates of between 20,000 to several million different proteoforms covering a broad dynamic range.
Many methods have been used to explore the proteome from antibody-based methods to mass spectrometry. However, the current leader within the space with the largest coverage is Somascan. This is an aptamer-based proteomic technology for biomarker discovery capable of simultaneously measuring thousands of proteins from small sample volumes (~15 µL). The Somascan platform currently allows detection of up to 7,000 proteins – more than double the number of proteins compared to any other platform, covering more biological pathways than any other proteomic assay. Built on DNA aptamer technology with protein-like side chains the impressive target range of this proteomic platform is supported through the speed of aptamer generation and lack of reliance on the immune system to generate aptamers, whereas antibodies typically rely on immune responses from a host animal.
Taking the heat out of hot-starts
Taq DNA polymerase used in PCR reactions can add bases to ssDNA in a non-template-dependent manner. At the high temperatures used during PCR thermal cycling, this activity is reduced, and annealing becomes more stringent. Yet, at lower temperatures, including room temperature, this activity can impact the reaction specificity, leading to erroneous results. Preventing the low-temperature activity and retaining the reaction specificity is crucial to the experiment.
‘Hot-start’ PCR offered a way to overcome the specificity issues. Approaches include excluding the Taq DNA polymerase enzyme and spiking it into the mixture once thermal cycling begins at high temperatures or using antibodies specific to Taq that could block the enzyme’s activity at room temperature. In the first approach, process issues include the labor-intensive requirements of opening all the tubes during the reaction and adding small volumes of reagents, risking contamination. Antibodies are still used in ‘hot start’ reactions, but NEB, the life science reagent supplier now offers a best in class alternative, that is based on DNA aptamers.
Just as the antibody-Taq inhibitory solution worked, the Taq aptamer used by NEB inhibits Taq polymerase at room temperature. This DNA aptamer solution has proven to yield highly specific PCR reactions with additional advantages over the antibody-Taq inhibitor:
- The aptamer inhibition/activation process is fully reversible, preventing any further Taq activity at the end of thermal cycling.
- The aptamer inhibition is released at lower temperatures (~45°C) than the antibody (94°C), removing the high temperature activation step and enabling faster protocols.
From deciphering the structure of double-stranded DNA to cracking the human genome, over the past 70 years our understanding of DNA has advanced hugely. We are now engineering DNA molecules to offer better research reagents, improved diagnostics and novel therapeutic approaches using aptamer technology.
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