TY - JOUR
T1 - Droplet Digital PCR for Mutation Detection in Formalin-Fixed, Paraffin-Embedded Melanoma Tissues
T2 - A Comparison with Sanger Sequencing and Pyrosequencing
AU - McEvoy, Ashleigh C.
AU - Wood, Benjamin A.
AU - Ardakani, Nima M.
AU - Pereira, Michelle R.
AU - Pearce, Robert
AU - Cowell, Lester
AU - Robinson, Cleo
AU - Grieu-Iacopetta, Fabienne
AU - Spicer, Alexander J.
AU - Amanuel, Benhur
AU - Ziman, Melanie
AU - Gray, Elin S.
PY - 2018/3/1
Y1 - 2018/3/1
N2 - The identification of somatic mutations is crucial for guiding therapeutic decisions about personalized melanoma treatment. However, genetic analysis of tumors is usually performed on limited and often low-quality DNA from tumors with low tumor cellularity and high tumor heterogeneity. Different mutation-detection platforms exist, with varying analytical sensitivities. Here we evaluated the detection of common mutations in BRAF, NRAS, and TERT promoter in 40 melanoma FFPE tissues using Droplet Digital (dd)PCR, and compared the results to the detection rates obtained by Sanger sequencing and pyrosequencing. The cellularity of tumors analyzed ranged from 5% to 50% (n = 28) and 50% to 90% (n = 12). Overall, droplet digital (dd)PCR was more sensitive, detecting mutations in 12.5% and 23% of tumors deemed as wild-type by pyrosequencing and Sanger sequencing, respectively. The increased sensitivity of ddPCR was more apparent among tumors with <50% tumor cellularity. Implementation of ddPCR-based assays may facilitate analysis of early-stage tumors and support research into improving outcomes in melanoma patients.
AB - The identification of somatic mutations is crucial for guiding therapeutic decisions about personalized melanoma treatment. However, genetic analysis of tumors is usually performed on limited and often low-quality DNA from tumors with low tumor cellularity and high tumor heterogeneity. Different mutation-detection platforms exist, with varying analytical sensitivities. Here we evaluated the detection of common mutations in BRAF, NRAS, and TERT promoter in 40 melanoma FFPE tissues using Droplet Digital (dd)PCR, and compared the results to the detection rates obtained by Sanger sequencing and pyrosequencing. The cellularity of tumors analyzed ranged from 5% to 50% (n = 28) and 50% to 90% (n = 12). Overall, droplet digital (dd)PCR was more sensitive, detecting mutations in 12.5% and 23% of tumors deemed as wild-type by pyrosequencing and Sanger sequencing, respectively. The increased sensitivity of ddPCR was more apparent among tumors with <50% tumor cellularity. Implementation of ddPCR-based assays may facilitate analysis of early-stage tumors and support research into improving outcomes in melanoma patients.
UR - http://www.scopus.com/inward/record.url?scp=85042367399&partnerID=8YFLogxK
U2 - 10.1016/j.jmoldx.2017.11.009
DO - 10.1016/j.jmoldx.2017.11.009
M3 - Article
C2 - 29305225
AN - SCOPUS:85042367399
SN - 1525-1578
VL - 20
SP - 240
EP - 252
JO - The Journal of Molecular Diagnostics
JF - The Journal of Molecular Diagnostics
IS - 2
ER -