We have used Inivata's enhanced tagged-amplicon deep sequencing (TAm-Seq™) platform to harness the power of circulating tumor DNA (ctDNA) to accurately identify low level mutations in multiple cancers, monitor the progression of disease and track patient response to therapy.

We have developed an improved TAm-Seq platform, which enables detection of point mutations and indels in ctDNA with high levels of sensitivity and specificity. Our standardised TAm-Seq platform has a rapid turnaround time and is performed on plasma obtained from blood collected in a single 9mL standard EDTA tube. The TAm-Seq assay is able to analyze regions of interest (either hotspots or entire coding regions) in 35 distinct genes in the human genome with such sensitivity as to detect mutations at an allele frequency less than 1% with unpublished data demonstrating sensitivity as low as 0.1%.

We performed longitudinal analyses on a series of case studies in different types of cancer to determine molecular profiles and monitor treatment response. Patient A is a stage IV NSCLC patient with bone metastases. The patient was unresponsive to initial treatment, with a suspected T790M EGFR mutation. Patient B is a BRAF V600E-positive cerebral anaplastic xanthoastrocytoma patient with first-line treatment of surgery and radiotherapy, presenting at relapse with rare multiple extra-cranial (lymph node and bone) metastases.

An initial cohort of non-small cell lung cancer (NSCLC) patients, including the Patient A case study, had blood drawn at 2 time points (Day 1 and Day 21) which were analysed using TAm-Seq demonstrating clinical response correlation to reduction in mutant alleles between pre- and post-treatment. In addition to this, Patient A was confirmed by TAm-Seq as having the suspected T790M EGFR mutation at a 0.57% frequency. As tumor biopsy tissue was unavailable for testing resistant mutations in this instance, the detection of the T790M mutation by TAm-Seq enabled the patient to become eligible to receive AZD9291 off-label by special permission. The patient went on to show a complete metabolic response on PET scan. Patient B, previously presenting with a BRAF V600E mutant xanthoastrocytoma, was initiated off-label vemurafenib with ctDNA analysis detecting mutation frequency at 3.45% prior to treatment and 2.16% following 2 weeks of treatment. Follow-up samples have been collected at multiple time points for patient A and B with testing in progress. Additional case studies are in progress and all data will be presented.

This work demonstrates the potential utility of using ctDNA to identify resistance mutations and monitor treatment response of a tumor when a conventional biopsy is unobtainable. It highlights the importance of high sensitivity mutation detection and analyzing multiple genomic regions, as such a high proportion of ctDNA mutations in our data were present at an allele frequency of <1%, which lower sensitivity detection methods would have missed.

Citation Format: Sarah K. Smalley, Davina Gale, Andrew RJ Lawson, Jordi Remon-Masip, Esperanza Perez, Michelle Pugh, Karen Howarth, Mehdi Touat, Jean-Charles Soria, Antoine Hollebecque, Benjamin Besse, Tim Forshew, Vincent Plagnol, Ludovic Lacroix, Nitzan Rosenfeld. Assessing the clinical applications of ctDNA in patients with advanced stage metastatic cancer using our enhanced TAm-Seq platform. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B7.

©2015 American Association for Cancer Research.
2015