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Targeted deep sequencing of urothelial bladder cancers and associated urinary DNA : a 23‐gene panel with utility for non‐invasive diagnosis and risk stratification

2019; Wiley; Volume: 124; Issue: 3 Linguagem: Inglês

10.1111/bju.14808

1464-410X

Douglas G. Ward, Naheema S. Gordon, Rebecca H. Boucher, Sarah Pirrie, Laura Baxter, Sascha Ott, Lee Silcock, Celina Whalley, Joanne Stockton, Andrew D. Beggs, Mike Griffiths, Ben Abbotts, Hanieh Ijakipour, Fathimath N. Latheef, Robert A. Robinson, Andrew J. P. White, Nicholas D. James, Maurice P. Zeegers, Kar Keung Cheng, Richard T. Bryan,

Cancer Genomics and Diagnostics

Objectives To develop a focused panel of somatic mutations ( SM s) present in the majority of urothelial bladder cancers ( UBC s), to investigate the diagnostic and prognostic utility of this panel, and to compare the identification of SM s in urinary cell‐pellet (cp) DNA and cell‐free (cf) DNA as part of the development of a non‐invasive clinical assay. Patients and Methods A panel of SM s was validated by targeted deep‐sequencing of tumour DNA from 956 patients with UBC . In addition, amplicon and capture‐based targeted sequencing measured mutant allele frequencies ( MAF s) of SM s in 314 urine cp DNA s and 153 urine cf DNA s. The association of SM s with grade, stage, and clinical outcomes was investigated by univariate and multivariate Cox models. Concordance between SM s detected in tumour tissue and cp DNA and cf DNA was assessed. Results The panel comprised SM s in 23 genes: TERT (promoter), FGFR 3 , PIK 3 CA , TP 53 , ERCC 2 , RHOB , ERBB 2 , HRAS , RXRA , ELF 3 , CDKN 1A , KRAS , KDM 6A , AKT 1 , FBXW 7 , ERBB 3 , SF 3B1 , CTNNB 1 , BRAF , C3orf70 , CREBBP , CDKN 2A , and NRAS ; 93.5–98.3% of UBC s of all grades and stages harboured ≥1 SM (mean: 2.5 SM s/tumour). RAS mutations were associated with better overall survival ( P = 0.04). Mutations in RXRA , RHOB and TERT (promoter) were associated with shorter time to recurrence ( P < 0.05). MAF s in urinary cf DNA and cp DNA were highly correlated; using a capture‐based approach, >94% of tumour SM s were detected in both cpDNA and cf DNA . Conclusions SM s are reliably detected in urinary cp DNA and cf DNA . The technical capability to identify very low MAF s is essential to reliably detect UBC , regardless of the use of cp DNA or cf DNA . This 23‐gene panel shows promise for the non‐invasive diagnosis and risk stratification of UBC .