TY - JOUR
T1 - PIK3CA mutation enrichment and quantitation from blood and tissue
AU - Keraite, Ieva
AU - Alvarez-Garcia, Virginia
AU - Garcia-Murillas, Isaac
AU - Beaney, Matthew
AU - Turner, Nicholas C.
AU - Bartos, Clare
AU - Oikonomidou, Olga
AU - Kersaudy-Kerhoas, Maïwenn
AU - Leslie, Nicholas R.
PY - 2020/10/13
Y1 - 2020/10/13
N2 - PIK3CA is one of the two most frequently mutated genes in breast cancers, occurring in 30-40% of cases. Four frequent 'hotspot' PIK3CA mutations (E542K, E545K, H1047R and H1047L) account for 80-90% of all PIK3CA mutations in human malignancies and represent predictive biomarkers. Here we describe a PIK3CA mutation specific nuclease-based enrichment assay, which combined with a low-cost real-time qPCR detection method, enhances assay detection sensitivity from 5% for E542K and 10% for E545K to 0.6%, and from 5% for H1047R to 0.3%. Moreover, we present a novel flexible prediction method to calculate initial mutant allele frequency in tissue biopsy and blood samples with low mutant fraction. These advancements demonstrated a quick, accurate and simple detection and quantitation of PIK3CA mutations in two breast cancer cohorts (first cohort n = 22, second cohort n = 25). Hence this simple, versatile and informative workflow could be applicable for routine diagnostic testing where quantitative results are essential, e.g. disease monitoring subject to validation in a substantial future study.
AB - PIK3CA is one of the two most frequently mutated genes in breast cancers, occurring in 30-40% of cases. Four frequent 'hotspot' PIK3CA mutations (E542K, E545K, H1047R and H1047L) account for 80-90% of all PIK3CA mutations in human malignancies and represent predictive biomarkers. Here we describe a PIK3CA mutation specific nuclease-based enrichment assay, which combined with a low-cost real-time qPCR detection method, enhances assay detection sensitivity from 5% for E542K and 10% for E545K to 0.6%, and from 5% for H1047R to 0.3%. Moreover, we present a novel flexible prediction method to calculate initial mutant allele frequency in tissue biopsy and blood samples with low mutant fraction. These advancements demonstrated a quick, accurate and simple detection and quantitation of PIK3CA mutations in two breast cancer cohorts (first cohort n = 22, second cohort n = 25). Hence this simple, versatile and informative workflow could be applicable for routine diagnostic testing where quantitative results are essential, e.g. disease monitoring subject to validation in a substantial future study.
UR - http://www.scopus.com/inward/record.url?scp=85092500121&partnerID=8YFLogxK
U2 - 10.1038/s41598-020-74086-w
DO - 10.1038/s41598-020-74086-w
M3 - Article
C2 - 33051521
SN - 2045-2322
VL - 10
JO - Scientific Reports
JF - Scientific Reports
M1 - 17082
ER -
