Ultrasensitive measurement of hotspot mutations in tumor DNA in blood using error-suppressed multiplexed deep sequencing
Cette étude évalue la faisabilité d'une méthode à base de séquençage de prochaine génération pour quantifier la fraction de molécules d'ADN contenant des mutations spécifiques de tumeurs parmi l'ADN normal du plasma
Detection of cell-free tumor DNA in the blood has offered promise as a cancer biomarker, but practical clinical implementations have been impeded by the lack of a sensitive and accurate method for quantitation that is also simple, inexpensive, and readily scalable. Here we present an approach that uses next-generation sequencing to quantify the small fraction of DNA molecules that contain tumor-specific mutations within a background of normal DNA in plasma. Using layers of sequence redundancy designed to distinguish true mutations from sequencer misreads and PCR misincorporations, we achieved a detection sensitivity of approximately 1 variant in 5,000 molecules. In addition, the attachment of modular barcode tags to the DNA fragments to be sequenced facilitated the simultaneous analysis of more than a hundred patient samples. As proof of principle, we demonstrated the successful use of this method to follow treatment-associated changes in circulating tumor DNA levels in patients with non-small cell lung cancer. Our findings suggest that the deep sequencing approach described here may be applied to the development of a practical diagnostic test that measures tumor-derived DNA levels in blood.
Cancer Research , résumé, 2012