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A novel single-tube multi-marker assay that quantifies six serum proteins exhibited high sensitivity and specificity in identifying preeclampsia (PE), besting a currently used immunoassay in prognostic yield. An article in The Journal of Applied Laboratory Medicine describes these findings.

PE impacts 2%–8% of all pregnancies across the globe. New onset hypertension and proteinuria during the second half of gestation are core clinical symptoms, but this condition poses significant risks for pregnant women and their babies, including organ failure and even death. “Thus, advances in screening options of PE are needed for early prognosis, preventive management, and clinical decision-making,” study authors Maris Laan, PhD, and Kaspar Ratnik, MSc, of the University of Tartu, Estonia, told CLN Stat in a joint statement. Currently, the predictive value of maternal serum fms-like tyrosine kinase-1/ placental growth factor (sFlt-1/PlGF) tests is less than 40% for PE onset within 4 weeks.

Identifying PE at the early stages can reduce harm to the mother, make it easier to plan for deliveries, and improve clinical management. Laan, Ratnik, and the other authors sought to develop a multiplex assay to improve PE prediction.

The result, the Luminex 6PLEX assay, combines from a single sample the measurements of sFlt-1, PlGF, soluble endoglin (sENG), leptin, disintegrin and metalloproteinase domain-containing protein 12 (ADAM12), adiponectin, and Pentraxin 3 (PTX3). “The measured biomarker concentrations along with maternal characteristics are incorporated into the developed prognostic algorithm to rule in/rule out PE development during the ongoing pregnancy,” explained Laan and Ratnik.

Investigators drew 61 serum samples from 53 pregnant women between 180 and 275 gestational days to evaluate the assay’s performance. In all, 22 women eventually developed PE, while 31 gave birth without complications. PE was diagnosed based on International Society for the Study of Hypertension in Pregnancy (ISSHP) criteria.

The 6PLEX multiplex assay performs equally well using fresh and frozen-thawed blood serum samples. The investigators used logistic regression models to look at associations between clinical PE onset and biomarker measurements. Overall, they produced 30 PE prediction models from the 6PLEX measurements taken from women who developed PE and from those who did not.

The area under the curve (AUC) with the threshold AUC > 0.7 was employed to assess the predictive power of models. “For every model, a corresponding formula was developed along with the calculated threshold value for PE prediction and coefficients for the included biomarkers and clinical characteristics,” the investigators wrote.

Comparing the novel assay with a conventional test, Thermo Fisher’s B·R·A·H·M·S sFlt-1 KRYPTOR assay, they found a high correlation in sFlt-1/PlGF estimated for individual sera among both assays. “Due to higher sensitivity of the method, only the Luminex 6PLEX assay was able to measure statistically significant increase in sFlt-1 levels already 28 and more days before the onset of PE,” reported the investigators.

The 6PLEX combined with gestational age and maternal weight at sampling reached AUC 0.99, achieving 100% sensitivity and 96.9% specificity. It also achieved a higher prognostic yield (96.5%) compared to the B·R·A·H·M·S assay (73.7%).

A need exists to develop a noninvasive test that’s easily accessible, easy to interpret, and is cost-effective. With additional technical vetting and validation, the investigators believe the Luminex 6PLEX could potentially fulfill these gaps. This novel assay shows promise, yet the study itself had some limitations concerning its sample size, Octavia M. Peck Palmer, PhD, and Saswati Das, MD, wrote in a related editorial. The results only reflect singlet pregnancies of mainly Estonian origin, and the ISSHP diagnosis criteria used in the study dates back to 2014, they noted.

“Importantly, criteria for the diagnoses of preeclampsia differs among the ISSHP and the American College of Obstetricians and Gynecologists. Patients may be classified differently using the two guidelines. Importantly, laboratory testing thresholds differ among the guidelines for renal insufficiency and platelet count,” wrote Palmer and Das. Study participants also lacked diversity, they added. “Studies show ethnic differences in the pathogenesis of preeclampsia and the incidence of preeclampsia.”

Palmer and Das recommended validating the 6PLEX assay in a larger population before adopting it in clinical practice.

The study’s authors acknowledged that the sample size didn’t adequately consider the heterogeneity of PE, nor did it address the assay’s performance on patients receiving anti-hypertension therapy or other treatments. In future research, the investigators plan to use this test as an early screening tool during the first half of pregnancy. “For broader impact, the novel-developed solution requires large-scale studies involving several clinical centers to fine-tune the prediction algorithms and introduce the test in routine clinical practice,” said Laan and Ratnik.