TY - JOUR
T1 - Performance of next-generation sequencing on small tumor specimens and/or low tumor content samples using a commercially available platform
AU - Morris, Scott
AU - Subramanian, Janakiraman
AU - Gel, Esma
AU - Runger, George
AU - Thompson, Eric
AU - Mallery, David
AU - Weiss, Glen
N1 - Funding Information:
Paradigm Diagnostics funded this work (SM, DM and ET are all employed by Paradigm Diagnostics; GW was a consultant for Paradigm Diagnostics). The specific roles of these authors are articulated in the’author contributions’ section. The funders had a role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. We are grateful to Felicia Craciunescu, Andrey Loskutov, and Micheal Ballado for assistance in specimen processing.
Publisher Copyright:
© 2018 Morris et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
PY - 2018/4
Y1 - 2018/4
N2 - Background Next generation sequencing tests (NGS) are usually performed on relatively small core biopsy or fine needle aspiration (FNA) samples. Data is limited on what amount of tumor by volume or minimum number of FNA passes are needed to yield sufficient material for running NGS. We sought to identify the amount of tumor for running the PCDx NGS platform. Methods 2,723 consecutive tumor tissues of all cancer types were queried and reviewed for inclusion. Information on tumor volume, success of performing NGS, and results of NGS were compiled. Assessment of sequence analysis, mutation calling and sensitivity, quality control, drug associations, and data aggregation and analysis were performed. Results 6.4% of samples were rejected from all testing due to insufficient tumor quantity. The number of genes with insufficient sensitivity make definitive mutation calls increased as the percentage of tumor decreased, reaching statistical significance below 5% tumor content. The number of drug associations also decreased with a lower percentage of tumor, but this difference only became significant between 1–3%. The number of drug associations did decrease with smaller tissue size as expected. Neither specimen size or percentage of tumor affected the ability to pass mRNA quality control. A tumor area of 10 mm2 provides a good margin of error for specimens to yield adequate drug association results. Conclusions Specimen suitability remains a major obstacle to clinical NGS testing. We determined that PCR-based library creation methods allow the use of smaller specimens, and those with a lower percentage of tumor cells to be run on the PCDx NGS platform.
AB - Background Next generation sequencing tests (NGS) are usually performed on relatively small core biopsy or fine needle aspiration (FNA) samples. Data is limited on what amount of tumor by volume or minimum number of FNA passes are needed to yield sufficient material for running NGS. We sought to identify the amount of tumor for running the PCDx NGS platform. Methods 2,723 consecutive tumor tissues of all cancer types were queried and reviewed for inclusion. Information on tumor volume, success of performing NGS, and results of NGS were compiled. Assessment of sequence analysis, mutation calling and sensitivity, quality control, drug associations, and data aggregation and analysis were performed. Results 6.4% of samples were rejected from all testing due to insufficient tumor quantity. The number of genes with insufficient sensitivity make definitive mutation calls increased as the percentage of tumor decreased, reaching statistical significance below 5% tumor content. The number of drug associations also decreased with a lower percentage of tumor, but this difference only became significant between 1–3%. The number of drug associations did decrease with smaller tissue size as expected. Neither specimen size or percentage of tumor affected the ability to pass mRNA quality control. A tumor area of 10 mm2 provides a good margin of error for specimens to yield adequate drug association results. Conclusions Specimen suitability remains a major obstacle to clinical NGS testing. We determined that PCR-based library creation methods allow the use of smaller specimens, and those with a lower percentage of tumor cells to be run on the PCDx NGS platform.
UR - http://www.scopus.com/inward/record.url?scp=85046095305&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85046095305&partnerID=8YFLogxK
U2 - 10.1371/journal.pone.0196556
DO - 10.1371/journal.pone.0196556
M3 - Article
C2 - 29702695
AN - SCOPUS:85046095305
SN - 1932-6203
VL - 13
JO - PloS one
JF - PloS one
IS - 4
M1 - e0196556
ER -