TAmiRNA welcomes ISEV study highlighting need for more precise EV characterization
Biotech and diagnostic innovator TAmiRNA have welcomed a new review article indicating the importance of advanced RNA-sequencing methods in characterizing extracellular vesicles (EVs) as therapeutic agents or diagnostic targets.
The study ‘A critical systematic review of extracellular vesicle clinical trials’ is authored by Rachel Mizenko, Madison Feaver, Batuhan Bozkurt, Neona Lowe, Bryan Nguyen, Kuan-Wei Huang, AijunWang, and Randy Carney from the Departments of Biomedical Engineering and Surgery at University of California, Davis. It is published by the Journal of Extracellular Vesicles (See Resources).
EV characterization: still learning
This systematic review examines the landscape of extracellular vesicle (EV)-related clinical trials to identify clinical application and methodology trends in a field that the authors note is ‘still in its infancy’, with an additional focus on the acknowledgement of EV subpopulations.
The team analyzed data from public reporting repositories and catalogued more than 471 EV-related clinical trials since 1999, with indications for more than 200 diseases. Diagnostics and companion diagnostics represented the bulk of these clinical trials, with cancer being the most frequent application and mesenchymal stromal cell (MSC) EVs the most frequently used as potential therapeutics.
RNA sequencing: method of choice but lack of transparency
Significantly, the study identifies RNA-sequencing (RNA-seq) as the leading method used to characterize EVs, ahead of Western Blot, nanoparticle tracking analysis (NTA), and qRT-PCR as other characterization methods.
The characterization of EV cargo by RNA-seq is a field in which TAmiRNA has become a world leader, using both small RNA sequencing based on its miND® platform as well as whole transcriptome sequencing to quantify a diverse RNA population from short non-coding RNAs such as microRNAs to long non-coding RNAs and messenger RNAs. Tailored bioinformatic and multi-omic analysis tools allow deep insight into the molecular composition of isolated EV populations and can support the prediction of biological functions of EVs based on their composition. Thus, RNA-seq can identify specific molecular signatures in EVs, which may be useful for diagnostics or understanding the mechanism of action of EV therapeutics.
However, Mizenko et al. determined a general lack of explicitly defined methodologies in clinical trials, which includes insufficient reporting on the specifics of EV characterization techniques like RNA-seq. TAmiRNA is fully transparent on the methods used: in 2022 Kseniya Khamina-Kotisch, senior scientist at TAmiRNA, published the small RNA-sequencing workflow including the development of spike-in controls for quality control and absolute quantification1. This was followed by the published bioinformatic analysis to interpret small RNA-sequencing data, lead authored by TAmiRNA’s head of computational biology, Dr. Andreas Diendorfer2.
Chasing out subpopulations
Mizenko and coauthors further determine in their review that “Most of the reported characterization relied on bulk characterization of EV isolates, with only 11% utilizing EV subpopulations in their experimental design. While this may be connected to a lack of available techniques suitable for clinical implementation, it also highlights the opportunity for use of EV subpopulations to improve translational efforts,” the study comments.
Indeed, the detection of intraluminal miRNAs and other RNAs in a small subpopulation of EVs can help to uncover specific RNA species that could have functional or diagnostic importance. At TAmiRNA, we recently succeeded in increasing the sensitivity of small RNA-sequencing to accommodate single-cell analysis. This enhanced sensitivity of RNA-seq generates new opportunities for examining the RNA cargo of homogeneous EV subpopulations.
Key milestones
They end by identifying several key milestones towards future progress, including:
- RNA-seq, as a characterization technique, can identify disease-specific molecular signatures in EVs. These RNA signatures may be useful for diagnostics or understanding the disease state.
- Improved methodological reporting and classification are required.
- Incorporation of EV subpopulations: Employing advanced techniques such as RNA-seq characterize specific EV subpopulations can enhance the sensitivity and efficacy of clinical applications.
References:
1. Khamina, K., Diendorfer, A.B., Skalicky, S., Weigl, M., Pultar, M., Krammer, T.L., Fournier, C.A., Schofield, A.L., Otto, C., Smith, A.T., Buchtele, N., Schoergenhofer, C., Jilma, B., Frank, B.J.H., Hofstaetter, J.G., Grillari, R., Grillari, J., Ruprecht, K., Goldring, C.E. and Rehrauer, H. (2022). A MicroRNA Next-Generation-Sequencing Discovery Assay (miND) for Genome-Scale Analysis and Absolute Quantitation of Circulating MicroRNA Biomarkers. International Journal of Molecular Sciences, [online] 23(3), p.1226. doi: https://doi.org/10.3390/ijms23031226
2. Diendorfer, A., Khamina, K., Pultar, M. and Hackl, M. (2022). miND (miRNA NGS Discovery pipeline): a small RNA-seq analysis pipeline and report generator for microRNA biomarker discovery studies. F1000Research, 11, p.233. doi: https://doi.org/10.12688/f1000research.94159.1
About TAmiRNA
TAmiRNA specializes in technologies for profiling levels of blood-circulating miRNAs and developing multi-parametric classification algorithms (“signatures”). TAmiRNA uses these technologies to develop minimal-invasive diagnostic tests for drug development, early diagnosis, and prognosis of disease, and as companion diagnostic tests to support treatment decisions. Through its CRO business, TAmiRNA enables clients from academic to private sectors to perform high-quality RNA profiling of various sample types and offers a fully integrated service from RNA extraction to data analysis. TAmiRNA has optimized its workflows for the comprehensive analysis of EV small and long RNA cargo.
TAmiRNA is among Europe´s first companies that have CE-marked a microRNA-based test – hepatomiR® – which is intended for the diagnosis of liver function in liver cancer patients.
For TAmiRNA´s work on novel drug safety biomarkers, TAmiRNA receives funding from the Innovative Medicines Initiative 2 Joint Undertaking “TransBioLine” under grant agreement No. 821283. This Joint Undertaking receives support from the European Union’s Horizon 2020 research and innovation programme and EFPIA. For its work on senescence-associated biomarkers, TAmiRNA is receiving funding from Eureka-Eurostars grant Ab-SENS. For its work on novel liver function biomarkers, TAmiRNA is receiving funding from the Vienna Business Agency as well as FFG.
More information available at: www.tamirna.com
Resources
Click on A critical systematic review of extracellular (Mizenko et al.) to access the full study.
Click on miND® pipeline for small RNA-sequencing to learn more about NextGen multiomic EV and small RNA characterization.
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