Achieving high-sensitivity for clinical applications using augmented exome sequencing.
BACKGROUND
Whole exome sequencing is increasingly used for the clinical evaluation of genetic disease, yet the variation of coverage and sensitivity over medically relevant parts of the genome remains poorly understood. Several sequencing-based assays continue to provide coverage that is inadequate for clinical assessment.


METHODS
Using sequence data obtained from the NA12878 reference sample and pre-defined lists of medically-relevant protein-coding and noncoding sequences, we compared the breadth and depth of coverage obtained among four commercial exome capture platforms and whole genome sequencing. In addition, we evaluated the performance of an augmented exome strategy, ACE, that extends coverage in medically relevant regions and enhances coverage in areas that are challenging to sequence. Leveraging reference call-sets, we also examined the effects of improved coverage on variant detection sensitivity.


RESULTS
We observed coverage shortfalls with each of the conventional exome-capture and whole-genome platforms across several medically interpretable genes. These gaps included areas of the genome required for reporting recently established secondary findings (ACMG) and known disease-associated loci. The augmented exome strategy recovered many of these gaps, resulting in improved coverage in these areas. At clinically-relevant coverage levels (100 % bases covered at ≥20×), ACE improved coverage among genes in the medically interpretable genome (>90 % covered relative to 10-78 % with other platforms), the set of ACMG secondary finding genes (91 % covered relative to 4-75 % with other platforms) and a subset of variants known to be associated with human disease (99 % covered relative to 52-95 % with other platforms). Improved coverage translated into improvements in sensitivity, with ACE variant detection sensitivities (>97.5 % SNVs, >92.5 % InDels) exceeding that observed with conventional whole-exome and whole-genome platforms.


CONCLUSIONS
Clinicians should consider analytical performance when making clinical assessments, given that even a few missed variants can lead to reporting false negative results. An augmented exome strategy provides a level of coverage not achievable with other platforms, thus addressing concerns regarding the lack of sensitivity in clinically important regions. In clinical applications where comprehensive coverage of medically interpretable areas of the genome requires higher localized sequencing depth, an augmented exome approach offers both cost and performance advantages over other sequencing-based tests.
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Anil
A
Patwardhan
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Jason
J
Harris
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Nan
N
Leng
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Gabor
G
Bartha
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Deanna M
DM
Church
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Shujun
S
Luo
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Christian
C
Haudenschild
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Mark
M
Pratt
Biosystems and Biomaterials Division, National Institute of Standards and Technology, Gaithersburg, Maryland USA.
Justin
J
Zook
Biosystems and Biomaterials Division, National Institute of Standards and Technology, Gaithersburg, Maryland USA.
Marc
M
Salit
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Jeanie
J
Tirch
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Massimo
M
Morra
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Stephen
S
Chervitz
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Ming
M
Li
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Michael
M
Clark
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Sarah
S
Garcia
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Gemma
G
Chandratillake
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Scott
S
Kirk
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA ; Center for Inherited Cardiovascular Disease, Stanford University School of Medicine, Stanford, California 94305 USA.
Euan
E
Ashley
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA ; Department of Genetics, Stanford University School of Medicine, Stanford, California 94305 USA.
Michael
M
Snyder
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA ; Departments of Bioengineering & Genetics, Stanford University, Stanford, California 94305 USA.
Russ
R
Altman
Department of Genetics, Stanford University School of Medicine, Stanford, California 94305 USA.
Carlos
C
Bustamante
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA ; Institute for Computational Health Sciences, University of California, San Francisco, California 94158 USA.
Atul J
AJ
Butte
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
John
J
West
Personalis, Inc, 1330 O'Brien Drive, Menlo Park, California 94025 USA.
Richard
R
Chen
D059472:Exome; D015894:Genome, Human; D006801:Humans; D017422:Sequence Analysis, DNA / Q000379:methods*