Abstract

Next generation sequencing has revolutionised genomic studies of cancer, having facilitated the development of precision oncology treatments based on a tumour’s molecular profile. We aimed to develop a targeted gene sequencing panel for application to disparate cancer types with particular focus on tumours of the head and neck, plus test for utility in liquid biopsy. The final panel designed through Roche/Nimblegen combined 451 cancer-associated genes (2.01 Mb target region). 136 patient DNA samples were collected for performance and application testing. Panel sensitivity and precision were measured using well-characterised DNA controls (n = 47), and specificity by Sanger sequencing of the Aryl Hydrocarbon Receptor Interacting Protein (AIP) gene in 89 patients. Assessment of liquid biopsy application employed a pool of synthetic circulating tumour DNA (ctDNA). Library preparation and sequencing were conducted on Illumina-based platforms prior to analysis with our accredited (ISO15189) bioinformatics pipeline. We achieved a mean coverage of 395x, with sensitivity and specificity of >99% and precision of >97%. Liquid biopsy revealed detection to 1.25% variant allele frequency. Application to head and neck tumours/cancers resulted in detection of mutations aligned to published databases. In conclusion, we have developed an analytically-validated panel for application to cancers of disparate types with utility in liquid biopsy.

Details

Title
Development and validation of a targeted gene sequencing panel for application to disparate cancers
Author
McCabe, Mark J 1 ; Gauthier, Marie-Emilie A 2 ; Chan, Chia-Ling 3 ; Thompson, Tanya J 4 ; MC De Sousa, Sunita 5 ; Puttick, Clare 3 ; Grady, John P 3 ; Gayevskiy, Velimir 3 ; Jiang, Tao 3 ; Ying, Kevin 3 ; Cipponi, Arcadi 6 ; Deng, Niantao 7 ; Swarbrick, Alex 6   VIAFID ORCID Logo  ; Thomas, Melissa L 8 ; Lord, Reginald V 9 ; Johns, Amber L 6 ; Kohonen-Corish, Maija 10 ; Sandra A O’Toole 11 ; Clark, Jonathan 12 ; Mueller, Simon A 13 ; Gupta, Ruta 12 ; McCormack, Ann I 14 ; Dinger, Marcel E 15 ; Cowley, Mark J 16 ; Aghmesheh, Morteza 17 ; Amor, David 18 ; Andrews, Lesley 19 ; Antill, Yoland 20 ; Armitage, Shane 21 ; Arnold, Leanne 21 ; Balleine, Rosemary 22 ; Bastick, Patti 23 ; Beesley, Jonathan 24 ; Beilby, John 25 ; Bennett, Ian 26 ; Blackburn, Anneke 27 ; Bogwitz, Michael 28 ; Botes, Leon 19 ; Brennan, Meagan 29 ; Brown, Melissa 30 ; Buckley, Michael 31 ; Burgess, Matthew 32 ; Burke, Jo 33 ; Butow, Phyllis 34 ; Caldon, Liz 35 ; Callen, David 36 ; Campbell, Ian 37 ; Chauhan, Deepa 38 ; Chauhan, Manisha 39 ; Chenevix-Trench, Georgia 24 ; Christian, Alice 40 ; Clarke, Christine 41 ; Cohen, Paul 42 ; Colley, Alison 43 ; Crook, Ashley 44 ; Cui, James 45 ; Culling, Bronwyn 46 ; Cummings, Margaret 47 ; Dawson, Sarah-Jane 48 ; deFazio, Anna 49 ; Delatycki, Martin 32 ; Dickson, Rebecca 50 ; Dixon, Joanne 40 ; Dobrovic, Alexander 51 ; Dudding, Tracy 52 ; Edkins, Ted 53 ; Edwards, Stacey 54 ; Eisenbruch, Maurice 55 ; Gelareh Farshid 56 ; Fellows, Andrew 51 ; Fenton, Georgina 57 ; Field, Michael 58 ; Flanagan, James 59 ; Fong, Peter 60 ; Forrest, Laura 61 ; Fox, Stephen 51 ; French, Juliet 62 ; Friedlander, Michael 63 ; Gaff, Clara 64 ; Davi Gallego Ortega 65 ; Gattas, Mike 66 ; George, Peter 67 ; Giles, Graham 68 ; Gill, Grantley 69 ; Greening, Sian 17 ; Haan, Eric 70 ; Harris, Marion 71 ; Hart, Stewart 72 ; Hayward, Nick 24 ; Heiniger, Louise 73 ; Hopper, John 74 ; Hunt, Clare 75 ; James, Paul 76 ; Jenkins, Mark 74 ; Kefford, Rick 77 ; Kidd, Alexa 40 ; Kirk, Judy 78 ; Koehler, Jessica 19 ; Kollias, James 79 ; Lakhani, Sunil 80 ; Lindeman, Geoff 81 ; Lipton, Lara 82 ; Lobb, Liz 73 ; Mann, Graham 83 ; Marsh, Deborah 84 ; McLachaln, Sue Ann 85 ; Meiser, Bettina 19 ; Milne, Roger 86 ; Nightingale, Sophie 87 ; Shona O’Connell 75 ; Sarah O’Sullivan 88 ; Pachter, Nick 89 ; Patterson, Briony 90 ; Phillips, Kelly 91 ; Pieper, Ellen 92 ; Rickard, Edwina 93 ; Robinson, Bridget 94 ; Saleh, Mona 95 ; Salisbury, Elizabeth 96 ; Sambrook, Joe 97 ; Saunders, Christobel 98 ; Saunus, Jodi 99 ; Scott, Elizabeth 100 ; Scott, Clare 101 ; Scott, Rodney 102 ; Sexton, Adrienne 28 ; Shelling, Andrew 103 ; Simpson, Peter 104 ; Southey, Melissa 105 ; Spurdle, Amanda 106 ; Taylor, Jessica 89 ; Thorne, Heather 107 ; Trainer, Alison 108 ; Tucker, Kathy 19 ; Visvader, Jane 109 ; Walker, Logan 110 ; Williams, Rachael 111 ; Winship, Ingrid 112 ; Young, Mary-Anne 113 

1  Kinghorn Centre for Clinical Genomics, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia; Hormones and Cancer Group, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia; St Vincent’s Clinical School, UNSW Australia, Sydney, NSW, Australia 
2  Kinghorn Centre for Clinical Genomics, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia; The Sydney Head and Neck Cancer Institute, Chris O’Brien Lifehouse, Sydney, Australia; Children’s Cancer Institute, Randwick, NSW, Australia 
3  Kinghorn Centre for Clinical Genomics, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia 
4  Hormones and Cancer Group, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia 
5  Endocrine and Metabolic Unit, Royal Adelaide Hospital, Adelaide, SA, Australia; Adult Genetics Unit, Royal Adelaide Hospital, Adelaide, SA, Australia; Department of Genetics and Molecular Pathology, Centre for Cancer Biology, an SA Pathology and University of South Australia alliance, Adelaide, SA, Australia; School of Medicine, University of Adelaide, Adelaide, SA, Australia 
6  The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia 
7  St Vincent’s Clinical School, UNSW Australia, Sydney, NSW, Australia; The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia 
8  St Vincent’s Centre for Applied Medical Research, Darlinghurst, NSW, Australia 
9  St Vincent’s Centre for Applied Medical Research, Darlinghurst, NSW, Australia; Notre Dame University School of Medicine, Sydney, NSW, Australia 
10  The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia; St George and Sutherland Clinical School, UNSW Australia, Sydney, NSW, Australia; Woolcock Institute of Medical Research, University of Sydney, Sydney, NSW, Australia 
11  Northern Clinical School, The University of Sydney, Royal North Shore Hospital, St Leonards, NSW, Australia; Sydney Medical School, The University of Sydney, Camperdown,, NSW, Australia; Western Sydney University Medical School, Campbelltown, NSW, Australia; Australian Clinical Labs, Bella Vista, NSW, Australia 
12  The Sydney Head and Neck Cancer Institute, Chris O’Brien Lifehouse, Sydney, Australia; Department of Tissue Pathology and Diagnostic Oncology, Royal Prince Alfred Hospital, Camperdown, NSW, Australia; Central Clinical School, The University of Sydney, Sydney, NSW, Australia 
13  Kinghorn Centre for Clinical Genomics, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia; The Sydney Head and Neck Cancer Institute, Chris O’Brien Lifehouse, Sydney, Australia; Department for Oto-Rhino-Laryngology, Head and Neck Surgery, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland 
14  Hormones and Cancer Group, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia; St Vincent’s Clinical School, UNSW Australia, Sydney, NSW, Australia; Department of Endocrinology, St Vincent’s Hospital, Darlinghurst, NSW, Australia 
15  Kinghorn Centre for Clinical Genomics, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia; St Vincent’s Clinical School, UNSW Australia, Sydney, NSW, Australia 
16  Kinghorn Centre for Clinical Genomics, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia; St Vincent’s Clinical School, UNSW Australia, Sydney, NSW, Australia; Children’s Cancer Institute, Randwick, NSW, Australia 
17  Illawarra Cancer Care Centre, Wollongong Hospital, Wollongong, NSW, Australia 
18  Genetic Health Services Victoria, Royal Children’s Hospital, Melbourne, VIC, Australia 
19  Hereditary Cancer Clinic, Prince of Wales Hospital, Randwick, NSW, Australia 
20  Department of Haem and Medical Oncology, Peter MacCallum Cancer Centre, East Melbourne, VIC, Australia 
21  Molecular Genetics Lab, Royal Brisbane and Women’s Hospital, Brisbane, QLD, Australia 
22  Department of Translational Oncology, Westmead Hospital, Westmead, NSW, Australia 
23  St George Hospital, Kogarah, NSW, Australia 
24  Queensland Institute of Medical Research, Herston, QLD, Australia 
25  Queen Elizabeth Medical Centre, Nedlands, WA, Australia 
26  Silverton Place, Brisbane, QLD, Australia 
27  John Curtin School of Medical Research, Australian National University, Canberra, ACT, Australia 
28  Familial Cancer Centre, The Royal Melbourne Hospital, Parkville, VIC, Australia 
29  NSW Breast Cancer Institute, Westmead, NSW, Australia 
30  Department of Biochemistry, University of Queensland, St Lucia, QLD, Australia 
31  Molecular and Cytogenetics Unit, Prince of Wales Hospital, Randwick, NSW, Australia 
32  Clinical Genetics Service, Austin Health, Heidelberg, VIC, Australia 
33  Royal Hobart Hospital, Hobart, TAS, Australia 
34  Medical Psychology Unit, Royal Prince Alfred Hospital, Camperdown, NSW, Australia 
35  Replication and Genome Stability Cancer Division, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia 
36  Dame Roma Mitchell Cancer Research Laboratories, University of Adelaide/Hanson Institute, Adelaide, SA, Australia 
37  Peter MaCallum Cancer Centre, East Melbourne, VIC, Australia 
38  School of Psychology, University of Sydney, Sydney, NSW, Australia 
39  St Vincent’s Hospital Cancer Genetics Clinic, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia 
40  Genetics Department, Central Region Genetics Service, Wellington Hospital, Wellington, New Zealand 
41  Westmead Institute for Cancer Research, University of Sydney, Westmead Hospital, Westmead, NSW, Australia 
42  St John of God Subiaco Hospital, Subiaco, WA, Australia 
43  Department of Clinical Genetics, Liverpool Health Service, Liverpool, NSW, Australia 
44  Department of Clinical Genetics, Royal North Shore Hospital, St Leonards, NSW, Australia 
45  Epidemiology and Preventive Medicine, Monash University, Prahran, VIC, Australia 
46  Molecular and Clinical Genetics, Royal Prince Alfred Hospital, Camperdown, NSW, Australia 
47  Department of Pathology, University of Queensland Medical School, Herston, QLD, Australia 
48  Molecular Genetics Department, Cambridge University, Cambridge, United Kingdom 
49  Department of Gynaecological Oncology, Westmead Institute for Cancer Research, Westmead Hospital, Westmead, NSW, Australia 
50  Associate Genetic Counsellor, Royal North Shore Hospital, St Leonards, NSW, Australia 
51  Department of Pathology, Peter MacCallum Cancer Centre, East Melbourne, VIC, Australia 
52  Hunter Genetics, Hunter Area Health Service, Waratah, NSW, Australia 
53  Clinical Chemistry, Prince Margaret Hospital for Children, Perth, WA, Australia 
54  Department of Biochemistry and Molecular Biology, University of Queensland, St Lucia, QLD, Australia 
55  Department of Multicultural Health, University of Sydney, Sydney, NSW, Australia 
56  Tissue Pathology, IMVS, Adelaide, SA, Australia 
57  South West Family Cancer Clinic, Liverpool Hospital, Liverpool, NSW, Australia 
58  Clinical Geneticist, Royal North Shore Hospital, St Leonards, NSW, Australia 
59  Epigenetics Unit, Department of Surgery and Oncology, Imperial College London, London, United Kingdom 
60  Medical Oncology Department, Regional Cancer and Blood Services, Auckland City Hospital, Auckland, New Zealand 
61  Psychosocial Cancer Genetics Research Group, Parkville Familial Cancer Centre, Melbourne, VIC, Australia 
62  School of Molecular and Microbial Sciences, University of Queensland, St Lucia, QLD, Australia 
63  Department of Medical Oncology, Prince of Wales Hospital, Randwick, NSW, Australia 
64  Victorian Clinical Genetics Service, Royal Melbourne Hospital, Parkville, VIC, Australia 
65  Tumour Development Group, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia 
66  Queensland Clinical Genetic Service, Royal Children’s Hospital, Herston, QLD, Australia 
67  Clinical Biochemistry Unit, Canterbury Health Labs, Christchurch, New Zealand 
68  Cancer Epidemiology Centre, Anti Cancer Council of Victoria, Carlton South, VIC, Australia 
69  Department of Surgery, Royal Adelaide Hospital, Adelaide, SA, Australia 
70  Department of Medical Genetics, Women’s and Children’s Hospital, North Adelaide, SA, Australia 
71  Familial Cancer Clinic, Peter MacCallum Cancer Centre, East Melbourne, VIC, Australia 
72  Breast and Ovarian Cancer Genetics, Monash Medical Centre, Bentleigh East, VIC, Australia 
73  Medical Psychology Research Unit, The University of Sydney, Camperdown, NSW, Australia 
74  Centre for M.E.G.A. Epidemiology, University of Melbourne, Carlton, VIC, Australia 
75  Southern Health Familial Cancer Centre, Monash Medical Centre, Clayton, VIC, Australia 
76  Clinical Geneticist, Genetic Health Services, Monash Medical Centre, Clayton, VIC, Australia 
77  Director Medical Oncology, Westmead Hospital, Westmead, NSW, Australia 
78  Familial Cancer Service, Department of Medicine, Westmead Hospital, Westmead, NSW, Australia 
79  Breast Endocrine and Surgical Unit, Royal Adelaide Hospital, Adelaide, SA, Australia 
80  University of Queensland Centre for Clinical Research, University of Queensland, The Royal Brisbane and Women’s Hospital, Herston, QLD, Australia 
81  Breast Cancer Laboratory, Walter and Eliza Hall Institute, Parkville, VIC, Australia 
82  Medcial Oncology and Clinical Haematology Unit, Western Hospital, Footscray, VIC, Australia 
83  Westmead Institute for Cancer Research, Westmead Millennium Institute, Westmead, NSW, Australia 
84  Kolling Institute of Medical Research, Royal North Shore Hospital, St Leonards, NSW, Australia 
85  Department of Oncology, St Vincent’s Hospital, Fitzroy, VIC, Australia 
86  Centro Nacional de Investigaciones Oncologicas, Madrid, Spain 
87  Western Health and Peter MacCallum Cancer Centre, East Melbourne, VIC, Australia 
88  Genetic Services of Western Australia, Subiaco, WA, Australia 
89  Familial Cancer and Clinical Genetics, Royal Melbourne Hospital, Parkville, VIC, Australia 
90  Tasmanian Clinical Genetics Service, Royal Hobart Hospital, Hobart, TAS, Australia 
91  Department of Medical Oncology, Peter MacCallum Cancer Centre, East Melbourne, VIC, Australia 
92  Parkville Familial Cancer Centre and Genomic Medicine, Melbourne, VIC, Australia 
93  Familial Cancer Centre, Westmead Hospital, Westmead, NSW, Australia 
94  Oncology Service, Christchurch Hospital, Christchurch, New Zealand 
95  Centre for Genetic Education, Prince of Wales Hospital, Randwick, NSW, Australia 
96  Anatomical Pathology, Prince of Wales Hospital, Randwick, NSW, Australia 
97  Peter MacCallum Cancer Centre, East Melbourne, VIC, Australia 
98  School of Surgery and Pathology, Queen Elizabeth Medical Centre, Nedlands, WA, Australia 
99  Breast Pathology, University of Queensland Centre for Clinical Research, Royal Brisbane and Women’s Hospital, Herston, QLD, Australia 
100  South View Clinic, Kogarah, NSW, Australia 
101  Research Department, Walter and Eliza Hall Institute, Melbourne, VIC, Australia 
102  Hunter Area Pathology Service, John Hunter Hospital, New Lambton Heights, NSW, Australia 
103  Obstetrics and Gynaecology, University of Auckland, Auckland, New Zealand 
104  University of Queensland, Herston, QLD, Australia 
105  Genetic Epidemiology Laboratory, Department of Pathology, University of Melbourne, Melbourne, VIC, Australia 
106  Cancer Unit, Queensland Institute of Medical Research, Brisbane, QLD, Australia 
107  Research Department, Peter MacCallum Cancer Centre, East Melbourne, VIC, Australia 
108  University of NSW, Prince of Wales Hospital, Randwick, NSW, Australia 
109  The Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia 
110  Molecular Cancer Epidemiology Laboratory, Queensland Institute of Medical Research, Herston, QLD, Australia 
111  Family Cancer Clinic, St Vincent’s Hospital, Darlinghurst, NSW, Australia 
112  Department of Genetics, Royal Melbourne Hospital, Parkville, VIC, Australia 
113  Genome.One, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia 
Pages
1-16
Publication year
2019
Publication date
Nov 2019
Publisher
Nature Publishing Group
e-ISSN
20452322
Source type
Scholarly Journal
Language of publication
English
DOI
https://doi.org/10.1038/s41598-019-52000-3
ProQuest document ID
2315966111
Copyright
© 2019. This work is published under http://creativecommons.org/licenses/by/4.0/ (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.