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Journal Articles Clinical Chemistry Year : 2022

Highly Specific Droplet-Digital PCR Detection of Universally Methylated Circulating Tumor DNA in Endometrial Carcinoma

Guillaume Beinse
  • Function : Author
Bruno Borghese
  • Function : Author
Marie Métairie
  • Function : Author
Pierre-Alexandre Just
  • Function : Author
Geoffroy Poulet
  • Function : Author
Simon Garinet
  • Function : Author
Beatrice Parfait
  • Function : Author
Audrey Didelot
  • Function : Author
Camille Bourreau
  • Function : Author
Natacha Agueeff
  • Function : Author
Alexandre Lavollé
  • Function : Author
Benoit Terris
  • Function : Author
Charles Chapron
  • Function : Author
François Goldwasser
  • Function : Author
Karen Leroy
  • Function : Author
Helene Blons
  • Function : Author
Pierre Laurent-Puig
  • Function : Author
Valérie Taly
Jérôme Alexandre
  • Function : Author

Abstract

BACKGROUND: No circulating biomarker is available for endometrial carcinoma (EC). We aimed to identify DNA positions universally hypermethylated in EC, and to develop a digital droplet PCR (ddPCR) assay for detection of hypermethylated circulating tumor DNA (meth-ctDNA) in plasma from patients with EC. METHODS: DNA positions hypermethylated in EC, and without unspecific hypermethylation in tissue/cell types releasing circulating cell-free DNA in plasma, were identified in silico from TCGA/Gene Expression Omnibus (GEO) data. A methylation-specific ddPCR (meth-ddPCR) assay following bisulfite conversion of DNA extracted from plasma was optimized for detection of meth-ctDNA according to dMIQE guidelines. Performances were validated on a retrospective cohort (n ¼ 78 tumors, n ¼ 30 tumor-adjacent tissues), a prospective pilot cohort (n ¼ 33 stage I-IV patients), and 55 patients/donors without cancer. RESULTS: Hypermethylation of zinc finger and SCAN domain containing 12 (ZSCAN12) and/or oxytocin (OXT) classified EC samples from multiple noncancer samples with high diagnostic specificity/sensitivity [>97%; area under the curve (AUC) ¼ 0.99; TCGA/ GEO tissues/blood samples]. These results were confirmed in the independent retrospective cohort (AUC ¼ 0.99). Meth-ddPCR showed a high analytical specificity (limit of blank ¼ 2) and sensitivity (absolute lower threshold of detection ¼ 50 pg methDNA /mL plasma). In the pilot cohort, meth-ctDNA was detected in pretreatment plasma samples from 9/11 and 5/20 patients with advanced and non-advanced EC, respectively. 2 of 9 patients had ctDNA detected after macroscopic complete surgery and experienced progression within 6 months. No healthy donors had any copy of hypermethylated DNA detected in plasma. CONCLUSIONS: Meth-ddPCR of ZSCAN12/OXT allows a highly specific and sensitive detection of ctDNA in plasma from patients with EC and appears promising for personalized approaches for these patients.
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Dates and versions

hal-04301316 , version 1 (22-11-2023)

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Guillaume Beinse, Bruno Borghese, Marie Métairie, Pierre-Alexandre Just, Geoffroy Poulet, et al.. Highly Specific Droplet-Digital PCR Detection of Universally Methylated Circulating Tumor DNA in Endometrial Carcinoma. Clinical Chemistry, 2022, 68, pp.782 - 793. ⟨10.1093/clinchem/hvac020⟩. ⟨hal-04301316⟩
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