Circulating extracellular vesicles (EVs) such as cholera toxin B chain (CTB)- or annexin V (AV)-binding EVs were previously shown to be rich sources of biomarkers. Here we test if previously identified pre-eclampsia (PE) candidate biomarkers, TIMP-1 in CTB-EVs (CTB-TIMP) and PAI-1 in AV-EVs (AV-PAI) complement plasma PlGF in predicting PE in a low-risk obstetric population. Eight hundred and forty-three prospectively banked plasma samples collected at 28 + 0 to 32 + 0 gestation weeks in the Neonatal and Obstetrics Risk Assessment (NORA) cohort study were assayed by sandwich ELISAs for plasma PlGF, CTB-TIMP1 and AV-PAI1. Nineteen patients subsequently developed PE 7.3 (±2.9) weeks later at a mean gestational age of 36.1 ± 3.5 weeks. The biomarkers were assessed for their predictive accuracy for PE using stepwise multivariate logistic regression analysis with Firth correction and Areas under the curve (AUC). To achieve 100% sensitivity in predicting PE, the cut-off for plasma PlGF, CTB-TIMP1 & AV-PAI1 were set at <1235, ≤300 or >1300 and <10,550 pg/mL plasma, respectively. The corresponding AUCs, specificity and PPV at a 95% confidence interval were 0.92, 52.1% and 4.7%; 0.72, 44.5% and 4.0%; and 0.69, 21.5% and 2.9%, respectively. At 100% sensitivity, the three biomarkers had a combined AUC of 0.96, specificity of 78.6%, and PPV of 9.9%. This is the first large cohort validation of the utility of EV-associated analytes as disease biomarkers. Specifically, EV biomarkers enhanced the predictive robustness of an existing PE biomarker sufficiently to justify PE screening in a low-risk general obstetric population.
Keywords: Exosomes; PAI1; PIGF; TIMP1; annexin V; biomarkers; cholera toxin B chain; pre-eclampsia.