Evaluation of commercial kits for isolation and bisulfite conversion of circulating cell-free tumor DNA from blood
Tóm tắt
DNA methylation biomarkers in circulating cell-free DNA (cfDNA) have great clinical potential for cancer management. Most methods for DNA methylation analysis require bisulfite conversion, causing DNA degradation and loss. This is particularly challenging for cfDNA, which is naturally fragmented and normally present in low amounts. The aim of the present study was to identify an optimal combination of cfDNA isolation and bisulfite conversion kits for downstream analysis of DNA methylation biomarkers in plasma. Of the five tested bisulfite conversion kits (EpiJET Bisulfite Conversion Kit, EpiTect Plus DNA Bisulfite Kit (EpiTect), EZ DNA Methylation-Direct Kit, Imprint DNA Modification Kit (Imprint) and Premium Bisulfite Kit), the highest and lowest DNA yield and recovery were achieved using the EpiTect kit and the Imprint kit, respectively, with more than double the amount of DNA for the EpiTect kit. Of the three tested cfDNA isolation kits (Maxwell RSC ccfDNA Plasma Kit, QIAamp Circulating Nucleic Acid Kit (CNA) and QIAamp MinElute ccfDNA Mini Kit), the CNA kit yielded around twice as much cfDNA compared to the two others kits, although with more high molecular weight DNA present. When comparing various combinations of cfDNA isolation kits and bisulfite conversion kits, the CNA kit and the EpiTect kit were identified as the best-performing combination, resulting in the highest yield of bisulfite converted cfDNA from normal plasma, as measured by droplet digital PCR (ddPCR). As a proof of principle, this kit combination was used to process plasma samples from 13 colorectal cancer patients for subsequent ddPCR methylation analysis of BCAT1 and IKZF1. Methylation of BCAT1 and/or IKZF1 was identified in 6/10 (60%) stage IV patients and 1/3 (33%) stage III patients. Based on a thorough evaluation of five bisulfite conversion kits and three cfDNA isolation kits, both individually and in combination, the CNA kit and the EpiTect kit were identified as the best-performing kit combination, with highest DNA yield and recovery across a range of DNA input amounts. The combination was successfully used for detection of clinically relevant DNA methylation biomarkers in plasma from cancer patients.
Tài liệu tham khảo
citation_journal_title=Mol Cancer; citation_title=Liquid biopsy: a step closer to transform diagnosis, prognosis and future of cancer treatments; citation_author=SN Lone, S Nisar, T Masoodi, M Singh, A Rizwan, S Hashem; citation_volume=21; citation_issue=1; citation_publication_date=2022; citation_pages=79; citation_doi=10.1186/s12943-022-01543-7; citation_id=CR1
citation_journal_title=Expert Rev Mol Diagn; citation_title=Concentration of cell-free DNA in different tumor types; citation_author=OE Bryzgunova, MY Konoshenko, PP Laktionov; citation_volume=21; citation_issue=1; citation_publication_date=2021; citation_pages=63-75; citation_doi=10.1080/14737159.2020.1860021; citation_id=CR2
citation_journal_title=Anal Chem; citation_title=High-throughput droplet digital PCR system for absolute quantitation of DNA copy number; citation_author=BJ Hindson, KD Ness, DA Masquelier, P Belgrader, NJ Heredia, AJ Makarewicz; citation_volume=83; citation_issue=22; citation_publication_date=2011; citation_pages=8604-8610; citation_doi=10.1021/ac202028g; citation_id=CR3
citation_journal_title=Trends Genet; citation_title=Navigating the DNA methylation landscape of cancer; citation_author=A Nishiyama, M Nakanishi; citation_volume=37; citation_issue=11; citation_publication_date=2021; citation_pages=1012-1027; citation_doi=10.1016/j.tig.2021.05.002; citation_id=CR4
citation_journal_title=Bioorg Med Chem Lett; citation_title=Degradation of DNA by bisulfite treatment; citation_author=K Tanaka, A Okamoto; citation_volume=17; citation_issue=7; citation_publication_date=2007; citation_pages=1912-1915; citation_doi=10.1016/j.bmcl.2007.01.040; citation_id=CR5
citation_journal_title=Science (New York, NY).; citation_title=Epigenetics, fragmentomics, and topology of cell-free DNA in liquid biopsies; citation_author=YMD Lo, DSC Han, P Jiang, RWK Chiu; citation_volume=372; citation_publication_date=2021; citation_pages=6538; citation_doi=10.1126/science.aaw3616; citation_id=CR6
citation_journal_title=PLoS ONE; citation_title=Performance evaluation of kits for bisulfite-conversion of DNA from tissues, cell lines, FFPE tissues, aspirates, lavages, effusions, plasma, serum, and urine; citation_author=EE Holmes, M Jung, S Meller, A Leisse, V Sailer, J Zech; citation_volume=9; citation_issue=4; citation_publication_date=2014; citation_pages=e93933; citation_doi=10.1371/journal.pone.0093933; citation_id=CR7
citation_journal_title=PLoS ONE; citation_title=Evaluation of bisulfite kits for DNA methylation profiling in terms of DNA fragmentation and DNA recovery using digital PCR; citation_author=S Kint, W Spiegelaere, J Kesel, L Vandekerckhove, W Criekinge; citation_volume=13; citation_issue=6; citation_publication_date=2018; citation_pages=e0199091; citation_doi=10.1371/journal.pone.0199091; citation_id=CR8
citation_journal_title=PLoS ONE; citation_title=Bisulfite conversion of DNA: performance comparison of different kits and methylation quantitation of epigenetic biomarkers that have the potential to be used in non-invasive prenatal testing; citation_author=CA Leontiou, MD Hadjidaniel, P Mina, P Antoniou, M Ioannides, PC Patsalis; citation_volume=10; citation_issue=8; citation_publication_date=2015; citation_pages=e0135058; citation_doi=10.1371/journal.pone.0135058; citation_id=CR9
citation_journal_title=Epigenetics; citation_title=Comparative analysis of 12 different kits for bisulfite conversion of circulating cell-free DNA; citation_author=MBW Orntoft, SO Jensen, TB Hansen, JB Bramsen, CL Andersen; citation_volume=12; citation_issue=8; citation_publication_date=2017; citation_pages=626-636; citation_doi=10.1080/15592294.2017.1334024; citation_id=CR10
citation_journal_title=Genet Epigenet; citation_title=Comprehensive evaluation of commercial bisulfite-based DNA methylation kits and development of an alternative protocol with improved conversion performance; citation_author=S Tierling, B Schmitt, J Walter; citation_volume=10; citation_publication_date=2018; citation_pages=1179237X18766097; citation_doi=10.1177/1179237X18766097; citation_id=CR11
citation_journal_title=Forensic Sci Int Genet; citation_title=Development and optimization of the VISAGE basic prototype tool for forensic age estimation; citation_author=A Heidegger, C Xavier, H Niederstatter, M Puente, E Pospiech, A Pisarek; citation_volume=48; citation_publication_date=2020; citation_pages=102322; citation_doi=10.1016/j.fsigen.2020.102322; citation_id=CR12
citation_journal_title=Front Genet; citation_title=Bisulfite-converted DNA quantity evaluation: a multiplex quantitative real-time PCR system for evaluation of bisulfite conversion; citation_author=SR Hong, KJ Shin; citation_volume=12; citation_publication_date=2021; citation_pages=618955; citation_doi=10.3389/fgene.2021.618955; citation_id=CR13
citation_journal_title=J Clin Microbiol; citation_title=Does size matter? Comparison of extraction yields for different-sized DNA fragments by seven different routine and four new circulating cell-free extraction methods; citation_author=L Cook, K Starr, J Boonyaratanakornkit, R Hayden, SS Sam, AM Caliendo; citation_publication_date=2018; citation_doi=10.1128/JCM.01061-18; citation_id=CR14
citation_journal_title=Cancer Genet; citation_title=Evaluation of commercial kits for purification of circulating free DNA; citation_author=RJ Diefenbach, JH Lee, RF Kefford, H Rizos; citation_volume=228–229; citation_publication_date=2018; citation_pages=21-27; citation_doi=10.1016/j.cancergen.2018.08.005; citation_id=CR15
citation_journal_title=Cancers (Basel); citation_title=Comparison of circulating cell-free DNA extraction methods for downstream analysis in cancer patients; citation_author=PV Leest, PA Boonstra, AT Elst, LCV Kempen, M Tibbesma, J Koopmans; citation_publication_date=2020; citation_doi=10.3390/cancers12051222; citation_id=CR16
citation_journal_title=J Mol Diagn; citation_title=A comparison of cell-free DNA isolation kits: isolation and quantification of cell-free DNA in plasma; citation_author=L Sorber, K Zwaenepoel, V Deschoolmeester, G Roeyen, F Lardon, C Rolfo; citation_volume=19; citation_issue=1; citation_publication_date=2017; citation_pages=162-168; citation_doi=10.1016/j.jmoldx.2016.09.009; citation_id=CR17
citation_journal_title=Biopreserv Biobank; citation_title=Effects of quantification methods, isolation kits, plasma biobanking, and hemolysis on cell-free DNA analysis in plasma; citation_author=G Streleckiene, M Forster, R Inciuraite, R Lukosevicius, J Skieceviciene; citation_volume=17; citation_issue=6; citation_publication_date=2019; citation_pages=553-561; citation_doi=10.1089/bio.2019.0026; citation_id=CR18
citation_journal_title=Cancer Genet-Ny; citation_title=Comparison of 4 commercial kits for the extraction of circulating DNA from plasma; citation_author=K Warton, LJ Graham, N Yuwono, G Samimi; citation_volume=228; citation_publication_date=2018; citation_pages=143-150; citation_doi=10.1016/j.cancergen.2018.02.004; citation_id=CR19
citation_journal_title=Anal Bioanal Chem; citation_title=Comprehensive evaluation of methods to isolate, quantify, and characterize circulating cell-free DNA from small volumes of plasma; citation_author=F Mauger, C Dulary, C Daviaud, JF Deleuze, J Tost; citation_volume=407; citation_issue=22; citation_publication_date=2015; citation_pages=6873-6878; citation_doi=10.1007/s00216-015-8846-4; citation_id=CR20
citation_journal_title=Diagnostics (Basel); citation_title=Isolation and quantification of plasma cell-free DNA using different manual and automated methods; citation_author=E Polatoglou, Z Mayer, V Ungerer, AJ Bronkhorst, S Holdenrieder; citation_publication_date=2022; citation_doi=10.3390/diagnostics12102550; citation_id=CR21
citation_journal_title=Scand J Clin Lab Invest; citation_title=Performance comparison of commercial kits for isolating and detecting circulating tumor DNA; citation_author=M Wang, X Huang, X Li, Q Guo, W Xu, M Zhao; citation_volume=81; citation_issue=4; citation_publication_date=2021; citation_pages=276-281; citation_doi=10.1080/00365513.2020.1821394; citation_id=CR22
citation_journal_title=Clin Epigenetics; citation_title=A robust internal control for high-precision DNA methylation analyses by droplet digital PCR; citation_author=HD Pharo, K Andresen, KCG Berg, RA Lothe, M Jeanmougin, GE Lind; citation_volume=10; citation_publication_date=2018; citation_pages=24; citation_doi=10.1186/s13148-018-0456-5; citation_id=CR23
citation_journal_title=Cancer; citation_title=Detection of recurrent colorectal cancer with high specificity using a reporting threshold for circulating tumor DNA methylated in BCAT1 and IKZF1; citation_author=SK Pedersen, BL Musher, LC LaPointe, MK Tuck, EL Symonds, N Loayza; citation_volume=128; citation_issue=10; citation_publication_date=2022; citation_pages=1921-1928; citation_doi=10.1002/cncr.34159; citation_id=CR24
citation_journal_title=BMC Cancer; citation_title=Evaluation of an assay for methylated BCAT1 and IKZF1 in plasma for detection of colorectal neoplasia; citation_author=SK Pedersen, EL Symonds, RT Baker, DH Murray, A McEvoy, SC Doorn; citation_volume=15; citation_publication_date=2015; citation_pages=654; citation_doi=10.1186/s12885-015-1674-2; citation_id=CR25
citation_journal_title=Comput Biol Bioinf; citation_title=Efficacy of bisulfite modification and DNA recovery using commercial kits from samples of genomic and circulating DNA; citation_author=O Bryzgunova, P Laktionov, T Skvortsova, A Bondar, E Morozkin, A Lebedeva; citation_volume=1; citation_issue=6; citation_publication_date=2013; citation_pages=28-36; citation_id=CR26
citation_journal_title=Genome Res; citation_title=Nondestructive enzymatic deamination enables single-molecule long-read amplicon sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution; citation_author=Z Sun, R Vaisvila, LM Hussong, B Yan, C Baum, L Saleh; citation_volume=31; citation_issue=2; citation_publication_date=2021; citation_pages=291-300; citation_doi=10.1101/gr.265306.120; citation_id=CR27
citation_journal_title=Genome Res; citation_title=Enzymatic methyl sequencing detects DNA methylation at single-base resolution from picograms of DNA; citation_author=R Vaisvila, VKC Ponnaluri, Z Sun, BW Langhorst, L Saleh, S Guan; citation_volume=31; citation_issue=7; citation_publication_date=2021; citation_pages=1280-1289; citation_doi=10.1101/gr.266551.120; citation_id=CR28
citation_journal_title=Epigenetics Chromatin; citation_title=Evaluation of whole-genome DNA methylation sequencing library preparation protocols; citation_author=J Morrison, JM Koeman, BK Johnson, KK Foy, I Beddows, W Zhou; citation_volume=14; citation_issue=1; citation_publication_date=2021; citation_pages=28; citation_doi=10.1186/s13072-021-00401-y; citation_id=CR29
citation_journal_title=Clin Chem; citation_title=Effects of blood-processing protocols on fetal and total DNA quantification in maternal plasma; citation_author=RWK Chiu, LLM Poon, TK Lau, TN Leung, EMC Wong, YMD Lo; citation_volume=47; citation_issue=9; citation_publication_date=2001; citation_pages=1607-1613; citation_doi=10.1093/clinchem/47.9.1607; citation_id=CR30
citation_journal_title=Clin Chem; citation_title=Effects of blood-processing protocols on cell-free DNA quantification in plasma [4]; citation_author=DW Swinkels, E Wiegerinck, EAP Steegers, JB Kok; citation_volume=49; citation_issue=3; citation_publication_date=2003; citation_pages=525-526; citation_doi=10.1373/49.3.525; citation_id=CR31
citation_journal_title=Clin Cancer Res; citation_title=Harmonizing cell-free DNA collection and processing practices through evidence-based guidance; citation_author=SR Greytak, KB Engel, S Parpart-Li, M Murtaza, AJ Bronkhorst, MD Pertile; citation_volume=26; citation_issue=13; citation_publication_date=2020; citation_pages=3104-3109; citation_doi=10.1158/1078-0432.CCR-19-3015; citation_id=CR32
citation_journal_title=Bioinformatics; citation_title=PoDCall: positive droplet calling and normalization of droplet digital PCR DNA methylation data; citation_author=M Jeanmougin, HP Brodal, H Dietrichson Pharo, HM Vedeld, GE Lind; citation_publication_date=2023; citation_doi=10.1093/bioinformatics/btac766; citation_id=CR33
citation_journal_title=Hepatology; citation_title=Early and accurate detection of cholangiocarcinoma in patients with primary sclerosing cholangitis by methylation markers in bile; citation_author=HM Vedeld, MM Grimsrud, K Andresen, HD Pharo, E Seth, TH Karlsen; citation_volume=75; citation_issue=1; citation_publication_date=2022; citation_pages=59-73; citation_doi=10.1002/hep.32125; citation_id=CR34
citation_journal_title=Clin Chem; citation_title=The digital MIQE guidelines update: minimum information for publication of quantitative digital PCR experiments for 2020; citation_author=JF Huggett; citation_volume=66; citation_issue=8; citation_publication_date=2020; citation_pages=1012-1029; citation_doi=10.1093/clinchem/hvaa125; citation_id=CR35