Highly Specific Droplet-Digital PCR Detection of Universally Methylated Circulating Tumor DNA in Endometrial Carcinoma
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 patien...
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| Veröffentlicht in: | Clinical chemistry (Baltimore, Md.) Md.), 2022-06, Vol.68 (6), p.782-793 |
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| creator | Beinse, Guillaume Borghese, Bruno Métairie, Marie Just, Pierre-Alexandre Poulet, Geoffroy Garinet, Simon Parfait, Beatrice Didelot, Audrey Bourreau, Camille Agueeff, Natacha Lavollé, Alexandre Terris, Benoit Chapron, Charles Goldwasser, François Leroy, Karen Blons, Helene Laurent-Puig, Pierre Taly, Valérie Alexandre, Jérôme |
| description | 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 = 78 tumors, n = 30 tumor-adjacent tissues), a prospective pilot cohort (n = 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) = 0.99; TCGA/GEO tissues/blood samples]. These results were confirmed in the independent retrospective cohort (AUC = 0.99). Meth-ddPCR showed a high analytical specificity (limit of blank = 2) and sensitivity (absolute lower threshold of detection = 50 pgmethDNA/mLplasma). 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. |
| doi_str_mv | 10.1093/clinchem/hvac020 |
| format | Article |
| fullrecord | <record><control><sourceid>proquest_hal_p</sourceid><recordid>TN_cdi_hal_primary_oai_HAL_hal_04301316v1</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><oup_id>10.1093/clinchem/hvac020</oup_id><sourcerecordid>2642898214</sourcerecordid><originalsourceid>FETCH-LOGICAL-c411t-3e4502d513f152bcef79a2ea56d1f43d88cd428572ed1adab05ffca8c91a9ad43</originalsourceid><addsrcrecordid>eNqFkUuP0zAURi0EYjoDe1bISxAK40ecJssqHaZI5SGYWVuufd0YOXGwnUr996RqZ7asrq91vnMXH0LvKPlMScNvtXeD7qC_7Q5KE0ZeoAUVnBS1qOhLtCCENEVDy-UVuk7pz7yWy7p6ja644Iw3rFqgw8btO3_Ev0fQzjqN1zGMHnKxdnuXlcc_2194DRl0dmHAweLHwR0gJuXn1DfI3dGrDAa3Luppfrphjx-mPkS8_r7CbsB3gwk95OhmWauidkPo1Rv0yiqf4O1l3qDHL3cP7abY_rj_2q62hS4pzQWHUhBmBOWWCrbTYJeNYqBEZagtualrbUpWiyUDQ5VROyKs1arWDVWNMiW_QR_P3k55OUbXq3iUQTm5WW3l6Y-UnFBOqwOd2Q9ndozh7wQpy94lDd6rAcKUJKvmU03N6ElLzqiOIaUI9tlNiTw1I5-akZdm5sj7i33a9WCeA09VzMCnMxCm8f-6fxoanJg</addsrcrecordid><sourcetype>Open Access Repository</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>2642898214</pqid></control><display><type>article</type><title>Highly Specific Droplet-Digital PCR Detection of Universally Methylated Circulating Tumor DNA in Endometrial Carcinoma</title><source>Oxford University Press Journals All Titles (1996-Current)</source><creator>Beinse, Guillaume ; Borghese, Bruno ; Métairie, Marie ; Just, Pierre-Alexandre ; Poulet, Geoffroy ; Garinet, Simon ; Parfait, Beatrice ; Didelot, Audrey ; Bourreau, Camille ; Agueeff, Natacha ; Lavollé, Alexandre ; Terris, Benoit ; Chapron, Charles ; Goldwasser, François ; Leroy, Karen ; Blons, Helene ; Laurent-Puig, Pierre ; Taly, Valérie ; Alexandre, Jérôme</creator><creatorcontrib>Beinse, Guillaume ; Borghese, Bruno ; Métairie, Marie ; Just, Pierre-Alexandre ; Poulet, Geoffroy ; Garinet, Simon ; Parfait, Beatrice ; Didelot, Audrey ; Bourreau, Camille ; Agueeff, Natacha ; Lavollé, Alexandre ; Terris, Benoit ; Chapron, Charles ; Goldwasser, François ; Leroy, Karen ; Blons, Helene ; Laurent-Puig, Pierre ; Taly, Valérie ; Alexandre, Jérôme</creatorcontrib><description>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 = 78 tumors, n = 30 tumor-adjacent tissues), a prospective pilot cohort (n = 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) = 0.99; TCGA/GEO tissues/blood samples]. These results were confirmed in the independent retrospective cohort (AUC = 0.99). Meth-ddPCR showed a high analytical specificity (limit of blank = 2) and sensitivity (absolute lower threshold of detection = 50 pgmethDNA/mLplasma). 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.</description><identifier>ISSN: 0009-9147</identifier><identifier>EISSN: 1530-8561</identifier><identifier>DOI: 10.1093/clinchem/hvac020</identifier><identifier>PMID: 35323926</identifier><language>eng</language><publisher>England: Oxford University Press</publisher><subject>Life Sciences</subject><ispartof>Clinical chemistry (Baltimore, Md.), 2022-06, Vol.68 (6), p.782-793</ispartof><rights>American Association for Clinical Chemistry 2022. All rights reserved. For permissions, please email: journals.permissions@oup.com. 2022</rights><rights>American Association for Clinical Chemistry 2022. All rights reserved. For permissions, please email: journals.permissions@oup.com.</rights><rights>Distributed under a Creative Commons Attribution 4.0 International License</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c411t-3e4502d513f152bcef79a2ea56d1f43d88cd428572ed1adab05ffca8c91a9ad43</citedby><cites>FETCH-LOGICAL-c411t-3e4502d513f152bcef79a2ea56d1f43d88cd428572ed1adab05ffca8c91a9ad43</cites><orcidid>0000-0002-3116-8602</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>230,314,780,784,885,1584,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/35323926$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink><backlink>$$Uhttps://hal.science/hal-04301316$$DView record in HAL$$Hfree_for_read</backlink></links><search><creatorcontrib>Beinse, Guillaume</creatorcontrib><creatorcontrib>Borghese, Bruno</creatorcontrib><creatorcontrib>Métairie, Marie</creatorcontrib><creatorcontrib>Just, Pierre-Alexandre</creatorcontrib><creatorcontrib>Poulet, Geoffroy</creatorcontrib><creatorcontrib>Garinet, Simon</creatorcontrib><creatorcontrib>Parfait, Beatrice</creatorcontrib><creatorcontrib>Didelot, Audrey</creatorcontrib><creatorcontrib>Bourreau, Camille</creatorcontrib><creatorcontrib>Agueeff, Natacha</creatorcontrib><creatorcontrib>Lavollé, Alexandre</creatorcontrib><creatorcontrib>Terris, Benoit</creatorcontrib><creatorcontrib>Chapron, Charles</creatorcontrib><creatorcontrib>Goldwasser, François</creatorcontrib><creatorcontrib>Leroy, Karen</creatorcontrib><creatorcontrib>Blons, Helene</creatorcontrib><creatorcontrib>Laurent-Puig, Pierre</creatorcontrib><creatorcontrib>Taly, Valérie</creatorcontrib><creatorcontrib>Alexandre, Jérôme</creatorcontrib><title>Highly Specific Droplet-Digital PCR Detection of Universally Methylated Circulating Tumor DNA in Endometrial Carcinoma</title><title>Clinical chemistry (Baltimore, Md.)</title><addtitle>Clin Chem</addtitle><description>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 = 78 tumors, n = 30 tumor-adjacent tissues), a prospective pilot cohort (n = 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) = 0.99; TCGA/GEO tissues/blood samples]. These results were confirmed in the independent retrospective cohort (AUC = 0.99). Meth-ddPCR showed a high analytical specificity (limit of blank = 2) and sensitivity (absolute lower threshold of detection = 50 pgmethDNA/mLplasma). 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.</description><subject>Life Sciences</subject><issn>0009-9147</issn><issn>1530-8561</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2022</creationdate><recordtype>article</recordtype><recordid>eNqFkUuP0zAURi0EYjoDe1bISxAK40ecJssqHaZI5SGYWVuufd0YOXGwnUr996RqZ7asrq91vnMXH0LvKPlMScNvtXeD7qC_7Q5KE0ZeoAUVnBS1qOhLtCCENEVDy-UVuk7pz7yWy7p6ja644Iw3rFqgw8btO3_Ev0fQzjqN1zGMHnKxdnuXlcc_2194DRl0dmHAweLHwR0gJuXn1DfI3dGrDAa3Luppfrphjx-mPkS8_r7CbsB3gwk95OhmWauidkPo1Rv0yiqf4O1l3qDHL3cP7abY_rj_2q62hS4pzQWHUhBmBOWWCrbTYJeNYqBEZagtualrbUpWiyUDQ5VROyKs1arWDVWNMiW_QR_P3k55OUbXq3iUQTm5WW3l6Y-UnFBOqwOd2Q9ndozh7wQpy94lDd6rAcKUJKvmU03N6ElLzqiOIaUI9tlNiTw1I5-akZdm5sj7i33a9WCeA09VzMCnMxCm8f-6fxoanJg</recordid><startdate>20220601</startdate><enddate>20220601</enddate><creator>Beinse, Guillaume</creator><creator>Borghese, Bruno</creator><creator>Métairie, Marie</creator><creator>Just, Pierre-Alexandre</creator><creator>Poulet, Geoffroy</creator><creator>Garinet, Simon</creator><creator>Parfait, Beatrice</creator><creator>Didelot, Audrey</creator><creator>Bourreau, Camille</creator><creator>Agueeff, Natacha</creator><creator>Lavollé, Alexandre</creator><creator>Terris, Benoit</creator><creator>Chapron, Charles</creator><creator>Goldwasser, François</creator><creator>Leroy, Karen</creator><creator>Blons, Helene</creator><creator>Laurent-Puig, Pierre</creator><creator>Taly, Valérie</creator><creator>Alexandre, Jérôme</creator><general>Oxford University Press</general><general>American Association for Clinical Chemistry</general><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope><scope>1XC</scope><scope>VOOES</scope><orcidid>https://orcid.org/0000-0002-3116-8602</orcidid></search><sort><creationdate>20220601</creationdate><title>Highly Specific Droplet-Digital PCR Detection of Universally Methylated Circulating Tumor DNA in Endometrial Carcinoma</title><author>Beinse, Guillaume ; Borghese, Bruno ; Métairie, Marie ; Just, Pierre-Alexandre ; Poulet, Geoffroy ; Garinet, Simon ; Parfait, Beatrice ; Didelot, Audrey ; Bourreau, Camille ; Agueeff, Natacha ; Lavollé, Alexandre ; Terris, Benoit ; Chapron, Charles ; Goldwasser, François ; Leroy, Karen ; Blons, Helene ; Laurent-Puig, Pierre ; Taly, Valérie ; Alexandre, Jérôme</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c411t-3e4502d513f152bcef79a2ea56d1f43d88cd428572ed1adab05ffca8c91a9ad43</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2022</creationdate><topic>Life Sciences</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Beinse, Guillaume</creatorcontrib><creatorcontrib>Borghese, Bruno</creatorcontrib><creatorcontrib>Métairie, Marie</creatorcontrib><creatorcontrib>Just, Pierre-Alexandre</creatorcontrib><creatorcontrib>Poulet, Geoffroy</creatorcontrib><creatorcontrib>Garinet, Simon</creatorcontrib><creatorcontrib>Parfait, Beatrice</creatorcontrib><creatorcontrib>Didelot, Audrey</creatorcontrib><creatorcontrib>Bourreau, Camille</creatorcontrib><creatorcontrib>Agueeff, Natacha</creatorcontrib><creatorcontrib>Lavollé, Alexandre</creatorcontrib><creatorcontrib>Terris, Benoit</creatorcontrib><creatorcontrib>Chapron, Charles</creatorcontrib><creatorcontrib>Goldwasser, François</creatorcontrib><creatorcontrib>Leroy, Karen</creatorcontrib><creatorcontrib>Blons, Helene</creatorcontrib><creatorcontrib>Laurent-Puig, Pierre</creatorcontrib><creatorcontrib>Taly, Valérie</creatorcontrib><creatorcontrib>Alexandre, Jérôme</creatorcontrib><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><collection>Hyper Article en Ligne (HAL)</collection><collection>Hyper Article en Ligne (HAL) (Open Access)</collection><jtitle>Clinical chemistry (Baltimore, Md.)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Beinse, Guillaume</au><au>Borghese, Bruno</au><au>Métairie, Marie</au><au>Just, Pierre-Alexandre</au><au>Poulet, Geoffroy</au><au>Garinet, Simon</au><au>Parfait, Beatrice</au><au>Didelot, Audrey</au><au>Bourreau, Camille</au><au>Agueeff, Natacha</au><au>Lavollé, Alexandre</au><au>Terris, Benoit</au><au>Chapron, Charles</au><au>Goldwasser, François</au><au>Leroy, Karen</au><au>Blons, Helene</au><au>Laurent-Puig, Pierre</au><au>Taly, Valérie</au><au>Alexandre, Jérôme</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Highly Specific Droplet-Digital PCR Detection of Universally Methylated Circulating Tumor DNA in Endometrial Carcinoma</atitle><jtitle>Clinical chemistry (Baltimore, Md.)</jtitle><addtitle>Clin Chem</addtitle><date>2022-06-01</date><risdate>2022</risdate><volume>68</volume><issue>6</issue><spage>782</spage><epage>793</epage><pages>782-793</pages><issn>0009-9147</issn><eissn>1530-8561</eissn><abstract>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 = 78 tumors, n = 30 tumor-adjacent tissues), a prospective pilot cohort (n = 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) = 0.99; TCGA/GEO tissues/blood samples]. These results were confirmed in the independent retrospective cohort (AUC = 0.99). Meth-ddPCR showed a high analytical specificity (limit of blank = 2) and sensitivity (absolute lower threshold of detection = 50 pgmethDNA/mLplasma). 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.</abstract><cop>England</cop><pub>Oxford University Press</pub><pmid>35323926</pmid><doi>10.1093/clinchem/hvac020</doi><tpages>12</tpages><orcidid>https://orcid.org/0000-0002-3116-8602</orcidid><oa>free_for_read</oa></addata></record> |
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| source | Oxford University Press Journals All Titles (1996-Current) |
| subjects | Life Sciences |
| title | Highly Specific Droplet-Digital PCR Detection of Universally Methylated Circulating Tumor DNA in Endometrial Carcinoma |
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