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High-sensitivity monitoring of ctDNA by patient-specific sequencing panels and integration of variant reads

View ORCID ProfileJonathan C. M. Wan, View ORCID ProfileKatrin Heider, Davina Gale, Suzanne Murphy, Eyal Fisher, James Morris, View ORCID ProfileFlorent Mouliere, Dineika Chandrananda, Andrea Marshall, Andrew B. Gill, Pui Ying Chan, Emily Barker, Gemma Young, View ORCID ProfileWendy N. Cooper, Irena Hudecova, Francesco Marass, Graham R. Bignell, Constantine Alifrangis, Mark R. Middleton, Ferdia A. Gallagher, Christine Parkinson, Amer Durrani, Ultan McDermott, View ORCID ProfileChristopher G. Smith, Charles Massie, Pippa G. Corrie, Nitzan Rosenfeld
doi: https://doi.org/10.1101/759399
Jonathan C. M. Wan
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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Katrin Heider
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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Davina Gale
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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Suzanne Murphy
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
3Cambridge University Hospitals NHS Foundation Trust, Cambridge CB2 0QQ, UK
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Eyal Fisher
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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James Morris
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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Florent Mouliere
4Amsterdam UMC, Vrije Universiteit Amsterdam, Department of Pathology, Cancer Center Amsterdam, de Boelelaan 1117, Amsterdam, The Netherlands
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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Dineika Chandrananda
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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Andrea Marshall
5Warwick Clinical Trials Unit, University of Warwick, Coventry, UK
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Andrew B. Gill
3Cambridge University Hospitals NHS Foundation Trust, Cambridge CB2 0QQ, UK
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Pui Ying Chan
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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Emily Barker
6Cambridge Clinical Trials Unit – Cancer Theme, Cambridge, UK
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Gemma Young
6Cambridge Clinical Trials Unit – Cancer Theme, Cambridge, UK
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Wendy N. Cooper
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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Irena Hudecova
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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Francesco Marass
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
#Department of Biosystems Science and Engineering, ETH Zurich, 4058 Basel, Switzerland
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Graham R. Bignell
7Wellcome Sanger Institute, Hinxton CB10 1SA, UK
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Constantine Alifrangis
7Wellcome Sanger Institute, Hinxton CB10 1SA, UK
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Mark R. Middleton
8National Institute for Health Research Biomedical Research Centre, Oxford, UK
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Ferdia A. Gallagher
3Cambridge University Hospitals NHS Foundation Trust, Cambridge CB2 0QQ, UK
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Christine Parkinson
3Cambridge University Hospitals NHS Foundation Trust, Cambridge CB2 0QQ, UK
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Amer Durrani
3Cambridge University Hospitals NHS Foundation Trust, Cambridge CB2 0QQ, UK
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Ultan McDermott
7Wellcome Sanger Institute, Hinxton CB10 1SA, UK
‡AstraZeneca, CRUK Cambridge Institute, Robinson Way, Cambridge, UK CB2 0RE
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Christopher G. Smith
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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Charles Massie
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
9Department of Oncology, University of Cambridge Hutchison–MRC Research Centre, Box 197, Cambridge Biomedical Campus, Cambridge CB2 0XZ, UK
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Pippa G. Corrie
3Cambridge University Hospitals NHS Foundation Trust, Cambridge CB2 0QQ, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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Nitzan Rosenfeld
1Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
2Cancer Research UK Major Centre – Cambridge, Cancer Research UK Cambridge Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK
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  • For correspondence: nitzan.rosenfeld@cruk.cam.ac.uk
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Abstract

Circulating tumor-derived DNA (ctDNA) can be used to monitor cancer dynamics noninvasively. Patients with small tumors have few copies of ctDNA in plasma, resulting in limited sensitivity to detect low-volume or residual disease. We show that sampling limitations can be overcome and sensitivity for ctDNA detection can be improved by massively parallel sequencing when hundreds to thousands of mutations are identified by tumor genotyping. We describe the INtegration of VAriant Reads (INVAR) analysis pipeline, which combines patient-specific mutation lists with both custom error-suppression methods and signal enrichment based on biological features of ctDNA. In this framework, the sensitivity can be estimated independently for each sample based on the number of informative reads, which is the product of the number of mutations analyzed and the average depth of unique sequencing reads. We applied INVAR to deep sequencing data generated by custom hybrid-capture panels, and showed that when ~106 informative reads were obtained INVAR allowed detection of tumor-derived DNA fractions to parts per million (ppm). In serial samples from patients with advanced melanoma on treatment, we detected ctDNA when imaging confirmed tumor volume of ~1cm3. In patients with resected early-stage melanoma, ctDNA was detected in 40% of patients who later relapsed, with higher rates of detection when more informative reads were obtained. We further demonstrated that INVAR can be generalized and allows improved detection of ctDNA from whole-exome and low-depth whole-genome sequencing data.

Footnotes

  • ↵† C.M., P.G.C. and N.R. jointly supervised this work

  • One Sentence Summary: Integrating tumor-derived sequences across large panels of patient-specific mutations offers enhanced sensitivity for ctDNA detection and monitoring from both high-depth and low-depth plasma sequencing data.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted September 11, 2019.
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High-sensitivity monitoring of ctDNA by patient-specific sequencing panels and integration of variant reads
Jonathan C. M. Wan, Katrin Heider, Davina Gale, Suzanne Murphy, Eyal Fisher, James Morris, Florent Mouliere, Dineika Chandrananda, Andrea Marshall, Andrew B. Gill, Pui Ying Chan, Emily Barker, Gemma Young, Wendy N. Cooper, Irena Hudecova, Francesco Marass, Graham R. Bignell, Constantine Alifrangis, Mark R. Middleton, Ferdia A. Gallagher, Christine Parkinson, Amer Durrani, Ultan McDermott, Christopher G. Smith, Charles Massie, Pippa G. Corrie, Nitzan Rosenfeld
bioRxiv 759399; doi: https://doi.org/10.1101/759399
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High-sensitivity monitoring of ctDNA by patient-specific sequencing panels and integration of variant reads
Jonathan C. M. Wan, Katrin Heider, Davina Gale, Suzanne Murphy, Eyal Fisher, James Morris, Florent Mouliere, Dineika Chandrananda, Andrea Marshall, Andrew B. Gill, Pui Ying Chan, Emily Barker, Gemma Young, Wendy N. Cooper, Irena Hudecova, Francesco Marass, Graham R. Bignell, Constantine Alifrangis, Mark R. Middleton, Ferdia A. Gallagher, Christine Parkinson, Amer Durrani, Ultan McDermott, Christopher G. Smith, Charles Massie, Pippa G. Corrie, Nitzan Rosenfeld
bioRxiv 759399; doi: https://doi.org/10.1101/759399

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