Get more from your sequencer: Increased demand and new technology mean going beyond DNA analysis

As more and more healthcare decisions are based on laboratory-generated data, clinical lab teams are under constant pressure to do more with less. Tight budgets and limited staff sizes are well-known challenges; another comes from the physical footprint of the laboratory itself. Physicians may want more tests on the menu, but adding new testing platforms to enable menu expansion is not always feasible.

Size limitations are a major issue for clinical lab managers, and there is every expectation that space will continue to be at a premium going forward. For that reason, long-term laboratory planning should favor technology platforms that are more flexible, providing the ability to generate a wider variety of data from a small-footprint device.

Consider the ubiquitous DNA sequencer. Already, most labs have adopted some kind of sequencing platform, and those that haven’t yet will likely do so in the near future. However, with so many types of sequencers to choose from — small and large, rapid and slow, high-throughput and low-throughput, with a wide range of cost profiles across them — it can be difficult to determine which sequencer is the right choice. This becomes even more challenging when trying to predict what the lab’s sequencing needs will be in the coming years.

For long-term planning initiatives, the best way to evaluate sequencers may be less about brand and other well-known data points and more about instrument flexibility. A DNA sequencer will be useful for a certain group of testing needs, but if that same sequencer could also directly analyze RNA, methylation and other modifications, or possibly even proteins, it would offer much broader utility in the same physical footprint. Packing more capabilities into a platform is a handy way to expand testing options within the confines of a clinical lab.

Future test types

Technology planning is often based on new test types that are emerging from academia now and will be implemented in clinical labs in the next several years. On the sequencing front, there are several exciting areas of investigation that are likely to underpin future clinical tests.

Take modifications, for example. Certain DNA modifications are currently measured with methylation arrays for specific clinical testing needs, such as for imprinting disorders including Prader-Willi syndrome. However, arrays can be less sensitive, and they only generate results for known methylation patterns. A sequencing-based approach that directly detects DNA modifications in an unbiased manner could prove far more useful.

Beyond DNA, modifications of RNA will also likely become clinically relevant. These have been very difficult to investigate since most sequencing-based explorations of RNA are actually performed with cDNA, in which modifications have been stripped out by the conversion process. But as scientists begin to analyze RNA directly, they are finding that a clear view of expressed transcripts with their modifications intact is important for understanding how certain biomarkers are associated with disease states. Modifications can lead to aberrant splicing or translation, both of which can affect the final protein product. As such, the need for sequencing modifications within long-read RNA transcripts was a key focus of a recent report from the National Academies of Sciences, Engineering, and Medicine on the future of RNA sequencing.¹

Progress is also coming from new analyses of transfer RNAs (tRNAs). These highly structured molecules, containing many modified nucleotides, are difficult to convert to cDNA, so they have been understudied in the past. However, with better technical approaches, scientists are learning more about how tRNAs are linked to disease, and they are classifying isotypes based on modifications and other profile information. Ultimately, this work should lead to the development of clinically useful biomarkers.

Cell-free DNA studies have been around for years, and have already led to clinical tests related to cancer and prenatal health, with plenty of interest in other areas as well. Now, though, scientists are making inroads with the analysis of cell-free RNA. The conventional wisdom said that cell-free RNA would not be worth investigation; these transitory molecules would be too short and too degraded to provide any useful information. But with more advanced technologies, scientists have found that it is possible to generate longer reads from cell-free RNA, and that these molecules may be useful as biomarkers for cancer.

In one final example, there have been promising studies — and even some early clinical applications — of the use of sequencing data to better understand a person’s response to newer classes of treatment, such as cell and gene therapies or immunotherapies. One day, it could become routine to deploy sequencing tools (especially for RNA) to monitor treatment response.

All of these cases are in too early of a stage for clinical laboratory implementation, but it is important to be aware that they are on the horizon. Already, labs are integrating sequencing-based approaches for clinical applications such as germline sequencing, somatic oncology, and infectious disease diagnostics.

Technology considerations

Preparing for this expected spate of sequencing-based clinical tests requires a broader evaluation of technologies than simply what works for current needs. Here’s a brief list of considerations for longer-term technology planning.

Read length. Most widely used sequencing technologies generate short reads of a few hundred base pairs, making this the accepted norm. Some newer platforms offer much longer reads — extending to kilobases or even megabases captured in a single read. With traditional short-read sequencers, labs often must sacrifice key capabilities: rare disease testing with short reads can force compromises on detecting structural variants, methylation (unless a separate run is conducted), and speed. Long reads are also crucial for transcriptomic sequencing as they provide the complete information of an RNA transcript, allowing for better isoform quantification, isoform reconstruction, and fusion transcript detection, all of which are important in determining certain diseases.

Molecules supported. Looking ahead, sequencing DNA alone will not be sufficient for future testing needs. Some sequencers are already capable of directly sequencing RNA, eliminating the need for the cDNA conversion step and providing a clearer view of the native biology. Separately, certain sequencers are able to directly sequence some modified bases and their positional information within a full-length transcript without the hassle of bisulfite conversion, as well as other RNA nucleotide modifications that underpin certain disease biomarkers but for which orthogonal conversion methods to identify them within intact transcripts are not available. To support the broadest range of future tests — and to make the most of a single piece of equipment to ease space constraints — plan for a sequencing platform that can directly analyze DNA and RNA along with their modifications. If possible, look for a technology that is extensible to protein analysis as well.

Cost. This is an obvious one, but for long-term planning, it can be a challenge to predict platform pricing years in advance. It may be simpler to look at cost profiles (Does the instrument require a large outlay up front, or is it a lower capital expenditure?) and acceptable costs per sample, allowing clinical lab teams to focus on the cost elements most important to their own operations.

Size. The need to work within space constraints makes instrument size a very important element. In general, sequencing platforms have gotten smaller over time; manufacturers know that lab space is at a premium. For long-term planning, look for platforms that are smaller than their competitors now, or for sequencing manufacturers that have focused on reducing the size of their products.

With so much potential for future sequencing-based clinical tests, now is the time to begin planning for a platform that is flexible enough to meet a broad range of testing types. As sequencing technologies continue to improve, there should be some very good options suitable to clinical lab analysis of DNA, RNA, and even proteins.

Reference

1. Toward Sequencing and Mapping of RNA Modifications Committee, Board on Life Sciences, Board on Health Sciences Policy, Division on Earth and Life Studies, Health and Medicine Division, National Academies of Sciences, Engineering, and Medicine. Charting a future for sequencing RNA and its modifications: A New Era for biology and medicine. Published online July 22, 2024. doi:10.17226/27165.