Detection of endometrial cancer using tampon-based collection and methylated DNA markers

Jamie N. Bakkum-Gamez, Mark E. Sherman, Seth W. Slettedahl, Douglas W. Mahoney, Maureen A. Lemens, Shannon K. Laughlin-Tommaso, Matthew R. Hopkins, Ann VanOosten, Viji Shridhar, Julie K. Staub, Xiaoming Cao, Patrick H. Foote, Megan A. Clarke, Kelli N. Burger, Calise K. Berger, Maria C. O'Connell, Karen A. Doering, Karl C. Podratz, Christopher C. DeStephano, J. Kenneth SchoolmeesterSarah E. Kerr, Nicolas Wentzensen, William R. Taylor, John B. Kisiel

Research output: Contribution to journalArticlepeer-review


Objective: Alterations in DNA methylation are early events in endometrial cancer (EC) development and may have utility in EC detection via tampon-collected vaginal fluid. Methods: For discovery, DNA from frozen EC, benign endometrium (BE), and benign cervicovaginal (BCV) tissues underwent reduced representation bisulfite sequencing (RRBS) to identify differentially methylated regions (DMRs). Candidate DMRs were selected based on receiver operating characteristic (ROC) discrimination, methylation level fold-change between cancers and controls, and absence of background CpG methylation. Methylated DNA marker (MDM) validation was performed using qMSP on DNA from independent EC and BE FFPE tissue sets. Women ≥45 years of age with abnormal uterine bleeding (AUB) or postmenopausal bleeding (PMB) or any age with biopsy-proven EC self-collected vaginal fluid using a tampon prior to clinically indicated endometrial sampling or hysterectomy. Vaginal fluid DNA was assayed by qMSP for EC-associated MDMs. Random forest modeling analysis was performed to generate predictive probability of underlying disease; results were 500-fold in-silico cross-validated. Results: Thirty-three candidate MDMs met performance criteria in tissue. For the tampon pilot, 100 EC cases were frequency matched by menopausal status and tampon collection date to 92 BE controls. A 28-MDM panel highly discriminated between EC and BE (96% (95%CI 89–99%) specificity; 76% (66–84%) sensitivity (AUC 0.88). In PBS/EDTA tampon buffer, the panel yielded 96% (95% CI 87–99%) specificity and 82% (70–91%) sensitivity (AUC 0.91). Conclusion: Next generation methylome sequencing, stringent filtering criteria, and independent validation yielded excellent candidate MDMs for EC. EC-associated MDMs performed with promisingly high sensitivity and specificity in tampon-collected vaginal fluid; PBS-based tampon buffer with added EDTA improved sensitivity. Larger tampon-based EC MDM testing studies are warranted.

Original languageEnglish (US)
Pages (from-to)11-20
Number of pages10
JournalGynecologic oncology
StatePublished - Jul 2023


  • Cell-free nucleic acids
  • DNA methylation
  • Endometrial cancer
  • Endometrial neoplasm/diagnosis
  • Endometrial neoplasm/prevention & control
  • Liquid biopsy

ASJC Scopus subject areas

  • Oncology
  • Obstetrics and Gynecology


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