Sequencing-based microsatellite instability testing using as few as six markers for high-throughput clinical diagnostics.
Hum Mutat
; 41(1): 332-341, 2020 01.
Article
em En
| MEDLINE
| ID: mdl-31471937
ABSTRACT
Microsatellite instability (MSI) testing of colorectal cancers (CRCs) is used to screen for Lynch syndrome (LS), a hereditary cancer-predisposition, and can be used to predict response to immunotherapy. Here, we present a single-molecule molecular inversion probe and sequencing-based MSI assay and demonstrate its clinical validity according to existing guidelines. We amplified 24 microsatellites in multiplex and trained a classifier using 98 CRCs, which accommodates marker specific sensitivities to MSI. Sample classification achieved 100% concordance with the MSI Analysis System v1.2 (Promega) in three independent cohorts, totaling 220 CRCs. Backward-forward stepwise selection was used to identify a 6-marker subset of equal accuracy to the 24-marker panel. Assessment of assay detection limits showed that the 24-marker panel is marginally more robust to sample variables than the 6-marker subset, detecting as little as 3% high levels of MSI DNA in sample mixtures, and requiring a minimum of 10 template molecules to be sequenced per marker for >95% accuracy. BRAF c.1799 mutation analysis was also included to streamline LS testing, with all c.1799T>A variants being correctly identified. The assay, therefore, provides a cheap, robust, automatable, and scalable MSI test with internal quality controls, suitable for clinical cancer diagnostics.
Palavras-chave
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Marcadores Genéticos
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Testes Genéticos
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Repetições de Microssatélites
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Predisposição Genética para Doença
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Instabilidade de Microssatélites
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Ensaios de Triagem em Larga Escala
Tipo de estudo:
Diagnostic_studies
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Guideline
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Prognostic_studies
Limite:
Humans
Idioma:
En
Ano de publicação:
2020
Tipo de documento:
Article