Circulating cell-free DNA is elevated in postmenopausal compared with pre- and perimenopausal women.

Objective: With the rising use of circulating cell-free DNA (cirDNA) liquid biopsies for disease screening, it is important to understand biological differences that may impact the accuracy of cirDNA-based clinical tests. Although a number of biological factors have been researched, the relationship between menopause and cirDNA has not been thoroughly investigated. We aimed to compare plasma cirDNA concentration and DNA fragment integrity in healthy women pre- and postmenopause.

Methods: Blood was collected from healthy female volunteers 40 years and older. cirDNA was extracted from plasma (n = 52) and quantified by quantitative polymerase chain reaction (n = 47; 26 premenopause, mean age-46 y; 21 postmenopause, mean age-59 y). cirDNA concentration was quantitated using an ALU repetitive sequence with a 115-base-pair (bp) product (ALU-115), and long cirDNA fragments were quantitated using an ALU repetitive sequence with a 247-bp product (ALU-247). cirDNA integrity was expressed as a ratio of ALU-247 over ALU-115. Mann-Whitney U test was used to compare pre- and postmenopause qPCR results, and a two-tailed, unpaired t test was undertaken to compare the integrity ratio between the two groups.

Results: Postmenopause plasma samples were found to have a significantly higher cirDNA concentration (P < 0.0001, premenopause: mean, 3.10 ± 1.84 ng/mL; median, 2.90 ng/mL; postmenopause: mean, 5.28 ± 2.76 ng/mL; median, 4.56 ng/mL) and significantly higher concentration of long-stranded cirDNA fragments (P = 0.0033, premenopause: mean, 1.06 ± 0.48 ng/mL; median, 0.96 ng/mL; postmenopause: mean, 1.69 ± 0.89 ng/mL; median, 1.48 ng/mL). There was no significant difference in the integrity ratio between the groups (P = 0.1788).

Conclusions: Plasma cirDNA concentrations are higher in postmenopausal women. This has important implications in cirDNA liquid biopsy development and screening, especially for diseases such as cancer where the majority of cases are diagnosed postmenopause.