S Durgut, L Salihefendić, D Pećar, I Čeko, N Mulahuseinović, M Izmirlija, R Konjhodžić
{"title":"液滴数字 PCR 作为一种分子工具,用于检测具有表皮生长因子受体活化突变的 NSCLC 患者的表皮生长因子受体 T790M 突变。","authors":"S Durgut, L Salihefendić, D Pećar, I Čeko, N Mulahuseinović, M Izmirlija, R Konjhodžić","doi":"10.2478/bjmg-2023-0020","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Almost 50% of NSCLC patients who initially show a successful response to tyrosine kinase inhibitors targeted therapy (TKI therapy) eventually develop acquired <i>EGFR</i> T790M mutation. The T790M secondary mutation can cause resistance to the targeted therapy and disease relapse. Since this mutation can be present at very low frequencies in liquid biopsy samples, droplet digital PCR (ddPCR), due to its high sensitivity, has opened the possibility for minimally invasive monitoring of the disease during TKI targeted therapy.</p><p><strong>Materials and methods: </strong>For this study, a total of 45 plasma samples from NSCLC patients with previously detected <i>EGFR</i>-activating mutations were analyzed. Extracted circulating free DNA was amplified and examined for the presence of T790M mutation using ddPCR technology. For the data analysis, QuantaSoft Software was used.</p><p><strong>Results: </strong>Of 45 tested plasma samples, a total of 14 samples were identified as positive for the T790M mutation. The same samples eventually showed the presence of T790M mutation in FFPE. Droplet digital PCR showed its great advantage in high sensitivity detection of rare allele variants. Our ddPCR assay detected T790M mutant allele in frequencies from 0.1%. The average number of droplets generated by ddPCR was 9571.</p><p><strong>Conclusion: </strong>Monitoring of the T790M mutation has an important role in the examination of the effects of the prescribed TKI therapy. Since monitoring of potential changes during TKI therapy requires repeated sampling, our results showed that ddPCR technology has made it possible to use liquid biopsy as an adequate minimally invasive alternative for single nucleotide polymorphisms (SNP) detection.</p>","PeriodicalId":55403,"journal":{"name":"Balkan Journal of Medical Genetics","volume":null,"pages":null},"PeriodicalIF":0.5000,"publicationDate":"2024-03-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10932593/pdf/","citationCount":"0","resultStr":"{\"title\":\"Droplet Digital PCR as a Molecular Tool for the Detection of the <i>EGFR</i> T790M Mutation in NSCLC Patients with the <i>EGFR</i> Activating Mutations.\",\"authors\":\"S Durgut, L Salihefendić, D Pećar, I Čeko, N Mulahuseinović, M Izmirlija, R Konjhodžić\",\"doi\":\"10.2478/bjmg-2023-0020\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background: </strong>Almost 50% of NSCLC patients who initially show a successful response to tyrosine kinase inhibitors targeted therapy (TKI therapy) eventually develop acquired <i>EGFR</i> T790M mutation. The T790M secondary mutation can cause resistance to the targeted therapy and disease relapse. Since this mutation can be present at very low frequencies in liquid biopsy samples, droplet digital PCR (ddPCR), due to its high sensitivity, has opened the possibility for minimally invasive monitoring of the disease during TKI targeted therapy.</p><p><strong>Materials and methods: </strong>For this study, a total of 45 plasma samples from NSCLC patients with previously detected <i>EGFR</i>-activating mutations were analyzed. Extracted circulating free DNA was amplified and examined for the presence of T790M mutation using ddPCR technology. For the data analysis, QuantaSoft Software was used.</p><p><strong>Results: </strong>Of 45 tested plasma samples, a total of 14 samples were identified as positive for the T790M mutation. The same samples eventually showed the presence of T790M mutation in FFPE. Droplet digital PCR showed its great advantage in high sensitivity detection of rare allele variants. Our ddPCR assay detected T790M mutant allele in frequencies from 0.1%. The average number of droplets generated by ddPCR was 9571.</p><p><strong>Conclusion: </strong>Monitoring of the T790M mutation has an important role in the examination of the effects of the prescribed TKI therapy. Since monitoring of potential changes during TKI therapy requires repeated sampling, our results showed that ddPCR technology has made it possible to use liquid biopsy as an adequate minimally invasive alternative for single nucleotide polymorphisms (SNP) detection.</p>\",\"PeriodicalId\":55403,\"journal\":{\"name\":\"Balkan Journal of Medical Genetics\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":0.5000,\"publicationDate\":\"2024-03-12\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10932593/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Balkan Journal of Medical Genetics\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.2478/bjmg-2023-0020\",\"RegionNum\":4,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2023/12/1 0:00:00\",\"PubModel\":\"eCollection\",\"JCR\":\"Q4\",\"JCRName\":\"GENETICS & HEREDITY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Balkan Journal of Medical Genetics","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.2478/bjmg-2023-0020","RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2023/12/1 0:00:00","PubModel":"eCollection","JCR":"Q4","JCRName":"GENETICS & HEREDITY","Score":null,"Total":0}
Droplet Digital PCR as a Molecular Tool for the Detection of the EGFR T790M Mutation in NSCLC Patients with the EGFR Activating Mutations.
Background: Almost 50% of NSCLC patients who initially show a successful response to tyrosine kinase inhibitors targeted therapy (TKI therapy) eventually develop acquired EGFR T790M mutation. The T790M secondary mutation can cause resistance to the targeted therapy and disease relapse. Since this mutation can be present at very low frequencies in liquid biopsy samples, droplet digital PCR (ddPCR), due to its high sensitivity, has opened the possibility for minimally invasive monitoring of the disease during TKI targeted therapy.
Materials and methods: For this study, a total of 45 plasma samples from NSCLC patients with previously detected EGFR-activating mutations were analyzed. Extracted circulating free DNA was amplified and examined for the presence of T790M mutation using ddPCR technology. For the data analysis, QuantaSoft Software was used.
Results: Of 45 tested plasma samples, a total of 14 samples were identified as positive for the T790M mutation. The same samples eventually showed the presence of T790M mutation in FFPE. Droplet digital PCR showed its great advantage in high sensitivity detection of rare allele variants. Our ddPCR assay detected T790M mutant allele in frequencies from 0.1%. The average number of droplets generated by ddPCR was 9571.
Conclusion: Monitoring of the T790M mutation has an important role in the examination of the effects of the prescribed TKI therapy. Since monitoring of potential changes during TKI therapy requires repeated sampling, our results showed that ddPCR technology has made it possible to use liquid biopsy as an adequate minimally invasive alternative for single nucleotide polymorphisms (SNP) detection.
期刊介绍:
Balkan Journal of Medical Genetics is a journal in the English language for publication of articles involving all branches of medical genetics: human cytogenetics, molecular genetics, clinical genetics, immunogenetics, oncogenetics, pharmacogenetics, population genetics, genetic screening and diagnosis of monogenic and polygenic diseases, prenatal and preimplantation genetic diagnosis, genetic counselling, advances in treatment and prevention.