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QPOLE : A Quick, Simple, and Cheap Alternative for POLE Sequencing in Endometrial Cancer by Multiplex Genotyping Quantitative Polymerase Chain Reaction.

Authors :
Van den Heerik ASVM
Ter Haar NT
Vermij L
Jobsen JJ
Brinkhuis M
Roothaan SM
Leon-Castillo A
Ortoft G
Hogdall E
Hogdall C
Van Wezel T
Lutgens LCHW
Haverkort MAD
Khattra J
McAlpine JN
Creutzberg CL
Smit VTHBM
Gilks CB
Horeweg N
Bosse T
Source :
JCO global oncology [JCO Glob Oncol] 2023 May; Vol. 9, pp. e2200384.
Publication Year :
2023

Abstract

Purpose: Detection of 11 pathogenic variants in the POLE gene in endometrial cancer (EC) is critically important to identify women with a good prognosis and reduce overtreatment. Currently, POLE status is determined by DNA sequencing, which can be expensive, relatively time-consuming, and unavailable in hospitals without specialized equipment and personnel. This may hamper the implementation of POLE -testing in clinical practice. To overcome this, we developed and validated a rapid, low-cost POLE hotspot test by a quantitative polymerase chain reaction (qPCR) assay, QPOLE .<br />Materials and Methods: Primer and fluorescence-labeled 5'-nuclease probe sequences of the 11 established pathogenic POLE mutations were designed. Three assays, QPOLE -frequent for the most common mutations and QPOLE -rare-1 and QPOLE-rare-2 for the rare variants, were developed and optimized using DNA extracted from formalin-fixed paraffin-embedded tumor tissues. The simplicity of the design enables POLE status assessment within 4-6 hours after DNA isolation. An interlaboratory external validation study was performed to determine the practical feasibility of this assay.<br />Results: Cutoffs for POLE wild-type, POLE -mutant, equivocal, and failed results were predefined on the basis of a subset of POLE mutants and POLE wild-types for the internal and external validation. For equivocal cases, additional DNA sequencing is recommended. Performance in 282 EC cases, of which 99 were POLE -mutated, demonstrated an overall accuracy of 98.6% (95% CI, 97.2 to 99.9), a sensitivity of 95.2% (95% CI, 90.7 to 99.8), and a specificity of 100%. After DNA sequencing of 8.8% equivocal cases, the final sensitivity and specificity were 96.0% (95% CI, 92.1 to 99.8) and 100%. External validation confirmed feasibility and accuracy.<br />Conclusion: QPOLE is a qPCR assay that is a quick, simple, and reliable alternative for DNA sequencing. QPOLE detects all pathogenic variants in the exonuclease domain of the POLE gene. QPOLE will make low-cost POLE -testing available for all women with EC around the globe.

Details

Language :
English
ISSN :
2687-8941
Volume :
9
Database :
MEDLINE
Journal :
JCO global oncology
Publication Type :
Academic Journal
Accession number :
37229628
Full Text :
https://doi.org/10.1200/GO.22.00384