Comparison of total and endometrial circulating cell-free DNA in women with and without endometriosis.

Abstract Research question: Do women with laparoscopically confirmed endometriosis have higher plasma concentrations of circulating cell-free DNA (cirDNA) than those without endometriosis? Design: This prospective study recruited women aged 18-45 years undergoing benign gynaecological laparoscopy at two tertiary hospitals. Venous blood was collected immediately prior to surgery, and women were allocated to the endometriosis or control groups based on surgical findings. Total plasma cirDNA and cirDNA integrity were measured by qPCR targeting short (115 bases) and long (247 bases) ALU segments, while endometrial-derived cirDNA was measured by qPCR of bisulfite treated cirDNA using primers selective for a FAM101A sequence uniquely unmethylated in endometrial tissue. Five cirDNA parameters were compared between the control and endometriosis cohorts: total cirDNA concentration, long-stranded cirDNA concentration, integrity ratio, endometrial cirDNA concentration and endometrial cirDNA proportion. Results: Twenty-eight endometriosis and 15 control samples were included in analysis. Women with and without endometriosis had cirDNA concentrations of 2.24 ± 0.89 ng/mL and 2.56 ± 0.92 ng/mL respectively. Analysis by phenotype of endometriosis revealed a significantly higher endometrial cirDNA concentration in women with superficial disease (n = 10) compared to deep endometriosis (n = 18) (mean difference 0.14 ng/mL; 95% confidence interval 0.15-0.26; p = 0.025), but not compared to controls. Conclusions: There were no significant differences in any of the cirDNA parameters between women with and without endometriosis. The low statistical power and heterogenous pelvic pathology in the control group render it difficult to determine whether the negative results demonstrated in this study reflect a true lack of increase in cirDNA in endometriosis.