Plasma cell-free extrachromosomal circular DNA is a molecular hallmark of preeclampsia.

Background: Extrachromosomal circular DNA (eccDNA) is emerging as a novel biomarker for multiple diseases, but its role in reproductive diseases remains underexplored. Preeclampsia (PE) is associated with increased maternal and perinatal morbidity and mortality. Identifying new screening biomarkers may improve PE-affected pregnancies' outcomes. Methods: To investigate eccDNAs in PE, we collected plasma biopsies from 24 PE-affected and 23 normotensive (Norm) pregnancies, including 14 paired samples collected before and after diagnosis. eccDNA was profiled through linear DNA removal, circular DNA enrichment, and sequencing. The reliability of eccDNA detection was validated using Integrative Genomics Viewer plots and Sanger sequencing. Results: Our analysis identified significantly more eccDNAs in the plasma of PE-affected pregnancies than that in Norm pregnancies (p = 1.67 × 10-6). Notably, eccDNAs in PE were significantly enriched for genes previously found to be associated with PE, including COMT, RORC, CAPZA1, PPP1R12C, AIRE, CYP11A1, CYP11B1, and HSD11B2. Additionally, eccDNA carries four of these genes (AIRE, CYP11A1, CYP11B1, and HSD11B2) are associated with decreased circulating aldosterone levels, a known endocrine feature of PE. Statistically, COMT-containing eccDNA was the most abundant and recurrent in PE. Among the 14 paired plasma samples from pre-diagnosis and diagnosed PE cases, we observed a significant increase in eccDNA abundance in PE (p = 4.14 × 10-10), with 7 of the 9 gene-carrying eccDNAs uniquely identified in PE plasma. Conclusions: This study presents the first eccDNA landscape in the plasma of PE and indicates that plasma eccDNA may serve as a distinguishing hallmark between PE-affected and Norm pregnancies.