RESUMEN
OBJECTIVE: Conventional genetic tests (quantitative fluorescent-PCR [QF-PCR] and single nucleotide polymorphism-array) only diagnose ~40% of fetuses showing ultrasound abnormalities. Rapid exome sequencing (rES) may improve this diagnostic yield, but includes challenges such as uncertainties in fetal phenotyping, variant interpretation, incidental unsolicited findings, and rapid turnaround times. In this study, we implemented rES in prenatal care to increase diagnostic yield. METHODS: We prospectively studied 55 fetuses. Inclusion criteria were: (a) two or more independent major fetal anomalies, (b) hydrops fetalis or bilateral renal cysts alone, or (c) one major fetal anomaly and a first-degree relative with the same anomaly. In addition to conventional genetic tests, we performed trio rES analysis using a custom virtual gene panel of ~3850 Online Mendelian Inheritance in Man (OMIM) genes. RESULTS: We established a genetic rES-based diagnosis in 8 out of 23 fetuses (35%) without QF-PCR or array abnormalities. Diagnoses included MIRAGE (SAMD9), Zellweger (PEX1), Walker-Warburg (POMGNT1), Noonan (PTNP11), Kabuki (KMT2D), and CHARGE (CHD7) syndrome and two cases of Osteogenesis Imperfecta type 2 (COL1A1). In six cases, rES diagnosis aided perinatal management. The median turnaround time was 14 (range 8-20) days. CONCLUSION: Implementing rES as a routine test in the prenatal setting is challenging but technically feasible, with a promising diagnostic yield and significant clinical relevance.
Asunto(s)
Anomalías Múltiples/diagnóstico , Secuenciación del Exoma , Diagnóstico Prenatal/métodos , Anomalías Múltiples/epidemiología , Anomalías Múltiples/genética , Adulto , Pruebas Diagnósticas de Rutina/estadística & datos numéricos , Estudios de Factibilidad , Femenino , Feto/diagnóstico por imagen , Pruebas Genéticas/métodos , Pruebas Genéticas/estadística & datos numéricos , Humanos , Recién Nacido , Masculino , Países Bajos/epidemiología , Reacción en Cadena de la Polimerasa/métodos , Polimorfismo de Nucleótido Simple , Valor Predictivo de las Pruebas , Embarazo , Resultado del Embarazo/epidemiología , Diagnóstico Prenatal/estadística & datos numéricos , Estudios Prospectivos , Ultrasonografía PrenatalRESUMEN
Leukemias are genetically heterogeneous and diagnostics therefore includes various standard-of-care (SOC) techniques, including karyotyping, SNP-array and FISH. Optical genome mapping (OGM) may replace these as it detects different types of structural aberrations simultaneously and additionally detects much smaller aberrations (500 bp vs 5-10 Mb with karyotyping). However, its resolution may still be too low to define clinical relevance of aberrations when they are located between two OGM labels or when labels are not distinct enough. Here, we test the potential of Cas9-directed long-read sequencing (LRS) as an additional technique to resolve such potentially relevant new findings. From an internal Bionano implementation study we selected ten OGM calls that could not be validated with SOC methods. Per variant we designed crRNAs for Cas9 enrichment, prepared libraries and sequenced them on a MinION/GridION device. We could confirm all aberrations and, importantly, the actual breakpoints of the OGM calls were located between 0.2 and 5.5 kb of the OGM-estimated breakpoints, confirming the high reliability of OGM. Furthermore, we show examples of redefinition of aberrations between labels that enable judgment of clinical relevance. Our results suggest that Cas9-directed LRS can be a relevant and flexible secondary technique in diagnostic workflows including OGM.