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SAT630 Analytical Validation Of A Telomerase Reverse Transcriptase (TERT) Promoter Mutation Assay

Disclosure: P. Iyer: None. R. Dadu: Advisory Board Member; Self; Exelixis, Inc., Bayer, Inc. Research Investigator; Self; Merck, Exelixis, Inc., AstraZeneca. A. Barque: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. Z. Cleslei: Employee; Self; Veracyte, Inc. Stock Owner; Self; Vera...

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Autores principales: Iyer, Priyanka, Dadu, Ramona, Barque, Anna, Cleslei, Zanelli, Jiang, Huimin, Walsh, Sean, Zheng, Xingyu, Hao, Yangyang, Huang, Jing, Klopper, Joshua P, Kloos, Richard T, Cabanillas, Maria
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10554516/
http://dx.doi.org/10.1210/jendso/bvad114.2200
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author Iyer, Priyanka
Dadu, Ramona
Barque, Anna
Cleslei, Zanelli
Jiang, Huimin
Walsh, Sean
Zheng, Xingyu
Hao, Yangyang
Huang, Jing
Klopper, Joshua P
Kloos, Richard T
Cabanillas, Maria
author_facet Iyer, Priyanka
Dadu, Ramona
Barque, Anna
Cleslei, Zanelli
Jiang, Huimin
Walsh, Sean
Zheng, Xingyu
Hao, Yangyang
Huang, Jing
Klopper, Joshua P
Kloos, Richard T
Cabanillas, Maria
author_sort Iyer, Priyanka
collection PubMed
description Disclosure: P. Iyer: None. R. Dadu: Advisory Board Member; Self; Exelixis, Inc., Bayer, Inc. Research Investigator; Self; Merck, Exelixis, Inc., AstraZeneca. A. Barque: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. Z. Cleslei: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. H. Jiang: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. S. Walsh: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. X. Zheng: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. Y. Hao: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. J. Huang: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. J.P. Klopper: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. R.T. Kloos: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. M. Cabanillas: Advisory Board Member; Self; Exelixis, Inc., Bayer, Inc. Consulting Fee; Self; Lilly USA, LLC, Bayer, Inc. Research Investigator; Self; Genentech, Inc., Merck. TERT promoter mutations are relatively enriched in larger, aggressive thyroid cancers and these malignancies carry an independently decreased rate of disease free and disease specific survival when compared to thyroid cancers with wild type TERT promoter gene sequences. High quality analytical validation of a diagnostic test promotes confidence in the results which inform clinical decision making. TERT promoter mutation testing is now offered as a complement to Afirma Genomic Sequencing Classifier (GSC) testing in thyroid nodules that are not designated as molecularly benign. The Afirma TERT test was developed at Veracyte, Inc. based on the AmpliSeq-for-Illumina technology to detect TERT promoter variants C228T and C250T in genomic DNA (gDNA) extracted from patient thyroid fine needle aspirates (FNAs) sent for Afirma GSC testing, without any need for increased needle passes or sample volume. Herein, we describe the analytical performance of the Afirma TERT test. We examined the analytical sensitivity of the Afirma TERT test to varied input DNA amounts and the limit of detection (LOD) of variant allele frequency (VAF). We determined the negative percent agreement (NPA) and the positive percent agreement (PPA) of the Afirma TERT test against a reference primer pair. We also evaluated the analytical specificity from potential interfering substances such as RNA and blood gDNA. Further, the intra-run, inter-run and inter-laboratory reproducibility of the assay were examined. Analytical sensitivity analysis showed that the Afirma TERT test is tolerant to variation in DNA input amount (7-13 ng) and can detect expected positive TERT promoter variants down to 5% VAF LOD at 7ng DNA input with > 95% sensitivity. Both NPA and PPA were 100% against the reference primer pair. Analytical specificity studies demonstrated that the Afirma TERT test remains accurate in presence of 20% RNA or 80% blood gDNA for an average patient sample that typically has 30% VAF. The Afirma TERT test results are highly reproducible across different operators, runs, reagent lots, and laboratories. The Afirma TERT test also demonstrated a 100% confirmation rate when compared with an external NGS-based reference assay executed in a non-Veracyte laboratory. The analytical robustness and reproducibility of the Afirma TERT test support its routine clinical use among thyroid nodules with indeterminate cytology that are Afirma GSC suspicious or Bethesda V/VI nodules. Presentation: Saturday, June 17, 2023
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spelling pubmed-105545162023-10-06 SAT630 Analytical Validation Of A Telomerase Reverse Transcriptase (TERT) Promoter Mutation Assay Iyer, Priyanka Dadu, Ramona Barque, Anna Cleslei, Zanelli Jiang, Huimin Walsh, Sean Zheng, Xingyu Hao, Yangyang Huang, Jing Klopper, Joshua P Kloos, Richard T Cabanillas, Maria J Endocr Soc Tumor Biology Disclosure: P. Iyer: None. R. Dadu: Advisory Board Member; Self; Exelixis, Inc., Bayer, Inc. Research Investigator; Self; Merck, Exelixis, Inc., AstraZeneca. A. Barque: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. Z. Cleslei: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. H. Jiang: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. S. Walsh: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. X. Zheng: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. Y. Hao: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. J. Huang: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. J.P. Klopper: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. R.T. Kloos: Employee; Self; Veracyte, Inc. Stock Owner; Self; Veracyte, Inc. M. Cabanillas: Advisory Board Member; Self; Exelixis, Inc., Bayer, Inc. Consulting Fee; Self; Lilly USA, LLC, Bayer, Inc. Research Investigator; Self; Genentech, Inc., Merck. TERT promoter mutations are relatively enriched in larger, aggressive thyroid cancers and these malignancies carry an independently decreased rate of disease free and disease specific survival when compared to thyroid cancers with wild type TERT promoter gene sequences. High quality analytical validation of a diagnostic test promotes confidence in the results which inform clinical decision making. TERT promoter mutation testing is now offered as a complement to Afirma Genomic Sequencing Classifier (GSC) testing in thyroid nodules that are not designated as molecularly benign. The Afirma TERT test was developed at Veracyte, Inc. based on the AmpliSeq-for-Illumina technology to detect TERT promoter variants C228T and C250T in genomic DNA (gDNA) extracted from patient thyroid fine needle aspirates (FNAs) sent for Afirma GSC testing, without any need for increased needle passes or sample volume. Herein, we describe the analytical performance of the Afirma TERT test. We examined the analytical sensitivity of the Afirma TERT test to varied input DNA amounts and the limit of detection (LOD) of variant allele frequency (VAF). We determined the negative percent agreement (NPA) and the positive percent agreement (PPA) of the Afirma TERT test against a reference primer pair. We also evaluated the analytical specificity from potential interfering substances such as RNA and blood gDNA. Further, the intra-run, inter-run and inter-laboratory reproducibility of the assay were examined. Analytical sensitivity analysis showed that the Afirma TERT test is tolerant to variation in DNA input amount (7-13 ng) and can detect expected positive TERT promoter variants down to 5% VAF LOD at 7ng DNA input with > 95% sensitivity. Both NPA and PPA were 100% against the reference primer pair. Analytical specificity studies demonstrated that the Afirma TERT test remains accurate in presence of 20% RNA or 80% blood gDNA for an average patient sample that typically has 30% VAF. The Afirma TERT test results are highly reproducible across different operators, runs, reagent lots, and laboratories. The Afirma TERT test also demonstrated a 100% confirmation rate when compared with an external NGS-based reference assay executed in a non-Veracyte laboratory. The analytical robustness and reproducibility of the Afirma TERT test support its routine clinical use among thyroid nodules with indeterminate cytology that are Afirma GSC suspicious or Bethesda V/VI nodules. Presentation: Saturday, June 17, 2023 Oxford University Press 2023-10-05 /pmc/articles/PMC10554516/ http://dx.doi.org/10.1210/jendso/bvad114.2200 Text en © The Author(s) 2023. Published by Oxford University Press on behalf of the Endocrine Society. https://creativecommons.org/licenses/by-nc-nd/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs licence (https://creativecommons.org/licenses/by-nc-nd/4.0/), which permits non-commercial reproduction and distribution of the work, in any medium, provided the original work is not altered or transformed in any way, and that the work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Tumor Biology
Iyer, Priyanka
Dadu, Ramona
Barque, Anna
Cleslei, Zanelli
Jiang, Huimin
Walsh, Sean
Zheng, Xingyu
Hao, Yangyang
Huang, Jing
Klopper, Joshua P
Kloos, Richard T
Cabanillas, Maria
SAT630 Analytical Validation Of A Telomerase Reverse Transcriptase (TERT) Promoter Mutation Assay
title SAT630 Analytical Validation Of A Telomerase Reverse Transcriptase (TERT) Promoter Mutation Assay
title_full SAT630 Analytical Validation Of A Telomerase Reverse Transcriptase (TERT) Promoter Mutation Assay
title_fullStr SAT630 Analytical Validation Of A Telomerase Reverse Transcriptase (TERT) Promoter Mutation Assay
title_full_unstemmed SAT630 Analytical Validation Of A Telomerase Reverse Transcriptase (TERT) Promoter Mutation Assay
title_short SAT630 Analytical Validation Of A Telomerase Reverse Transcriptase (TERT) Promoter Mutation Assay
title_sort sat630 analytical validation of a telomerase reverse transcriptase (tert) promoter mutation assay
topic Tumor Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10554516/
http://dx.doi.org/10.1210/jendso/bvad114.2200
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