The Nu.Q NETs test, developed by epigenetics diagnostics firm VolitionRx to aid the detection and evaluation of diseases associated with NETosis such as COVID-19, sepsis, and other conditions, received CE marking in May. It is the first approved clinical assay to assess disease severity by quantifying circulating nucleosomes which make up plasma neutrophil extracellular traps, or NETs — snares of DNA expelled by neutrophils to trap and kill invading pathogens.
The process by which neutrophils unravel their chromosomes and expel chromatin outside the cell in response to infection — dying in the process — was first described in Science in 2004 but is still not widely understood. In a 2016 paper published in the Journal of Clinical Investigation, Scandinavian researchers concluded that there was an association between neutrophil extracellular traps with acute respiratory distress, sepsis, and thrombosis. However, the clinical potential for monitoring this process, called NETosis, has only recently been considered by physicians.
“There was an academic curiosity that NETs had a downside,” said Jake Micallef, chief scientific officer of Volition, one of the company’s two co-founding scientists. “The appreciation that they were involved in important diseases was really limited to those few people that were specifically interested in NETs clinically and scientifically.”
However, as NETosis levels have been correlated with the severity of immune response, Micallef said, NETs could be an important predictor of whether someone with a respiratory infection or sepsis is likely to need intensive care in hospital, for organ support, ventilation, or other treatments needing high-tech equipment.
Now, Volition has implemented its nucleosome quantification, or Nu.Q, technology to detect NETosis levels in patients. “The advent of COVID-19 made us realize that this is a whole new area that's of huge interest to us and completely amenable to our technology,” said Micallef. “The good thing for Volition is that our Nu.Q NETs assay — undertaken in a simple blood test — is currently the only validated and approved test for NETosis in the world.”
The Significance of NETs
With respiratory infections caused by a virus such as COVID-19 or influenza, NETs are produced in the lungs, but an excess can clog up the capillaries and damage their cellular lining. This in turn damages the lining of the alveoli in the lungs which lose their semi-permeable properties and eventually start to leak body fluids. At this point, the lungs start to fail and breathing becomes difficult. The worst cases lead to multiple organ failure, septic shock, lowered blood pressure, and death.
“Knowing which patients have high levels of NETs early would be good for the patient because they will get the treatment they require earlier, such as antivirals, antibiotics, ventilation or other organ support to manage them better,” Micallef explained.
“NETs are such a critical part of the inflammation pathway, they are relevant in any inflammatory disease, whether that's thrombosis, COVID-19, sepsis, cancer, and transplants,” said Andrew Aswani, consultant in critical care and anesthesia at Guys and St. Thomas’ NHS Foundation Trust in London. “The unique and powerful potential of measuring them — and potentially reducing them — could really empower human health.”
Currently, in sepsis patients, clinicians use SOFA (sequential organ failure assessment) scores from 0-24 to determine severity of disease, looking at six areas — respiratory, cardiovascular, hepatic, coagulation, renal and neurological — through daily clinical exams and blood tests. Aswani said it would potentially be extremely useful if a prognostic test could be integrated into that blood panel.
Right at the start of COVID-19, Volition started researching the possibilities of a NETs biomarker test using its existing Nu.Q technology. Volition collaborated with University Hospital Liege in Belgium and Technical University Munich, Germany, which tested COVID-19 patients for NETs. The results of the study, published in 2021, showed an association between the severity of disease and NETs.
Next, Volition worked with researchers at King’s College London, studying NET levels in patients on admission to hospital with COVID-19; a non-severe cohort and a severe cohort requiring organ support. The study, presented in a poster at the International Society on Thrombosis and Haemostasis (ISTH) in 2021, found that a high NETs level on admission was predictive of which patients would need organ support in ITU, with the highest levels associated with the poorest outcomes, demonstrating the test’s predictive value.
A more recent study of 118 subjects published at the ISTH Congress in July 2022 provided a direct comparison of NETosis biomarkers including nucleosomes in two infectious conditions (septic shock and COVID-19). A key finding from this project was the correlation between the Nu.Q NETs level and the currently used SOFA score.
Now the company is awaiting the results of studies by researchers at a leading UK hospital for NET levels in up to 180 sepsis, COVID-19, and cardiac patients in ITU.
In addition, Volition is collaborating with the University of Texas MD Anderson Cancer Center on a research study looking at the correlation of NETs in cancer patients with sepsis.
Aswani says more research is needed, but the results from the published data and now his own hospital are adding up. The Nu.Q NETs assay is, he says, a simple, low cost and effective measure of NETs that may be used to identify patients with clinically relevant elevated circulating levels of NETs and enable physicians to rapidly treat these patients.
Can NETs be Treated?
Since COVID-19 and the realization of the importance of NETs, researchers and companies are now developing a variety of drug therapies and medical devices directly targeting excessive NETosis. Aswani is also the chief medical officer of Santersus, a company focused on the development of devices used to cleanse patient blood of NETs.
An animal study published by Aswani et al last year found that their NucleoCapture technology removed NETs from the blood of pigs with sepsis. After a safety trial of 10 patients, the researchers from Santersus are recruiting for phase two human trials later this year.