Bead-Based Isolation of Circulating Tumor DNA from Pancreatic Cancer Patients Enables High Fidelity Next Generation Sequencing.

INTRODUCTION: Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive cancers and poses a challenge to the treating clinician. With the emergence of genomic profiling technologies, circulating tumor DNA (ctDNA) is increasingly recognized as a versatile biomarker for risk stratification and disease monitoring. We aimed to compare two commercially available NGS panels in a cohort of patients with advanced PDAC undergoing palliative chemotherapy. METHODS: CtDNA was isolated with a magnetic bead-based protocol from two consecutive blood samples before and during chemotherapy in 21 patients with PDAC. Mutations were assessed by using a panel covering 15 (GP15) or 50 (GP50) cancer-associated genes. Results were compared to tumor tissue (GP15), if available. RESULTS: Isolation of ctDNA resulted in a high mean value of 1.9 ng/µL (total volume of ~40 µL). Although the same number of patients were positive for at least one mutation (76%), the most commonly mutated oncogene in PDAC, KRAS, was detectable in an additional 25% of all patients with the GP15 panel due to a higher coverage. The genomic concordance rate between tissue DNA and ctDNA analyses was 65.22%. DISCUSSION: Our study demonstrates the feasibility of an NGS-based approach for ctDNA analysis and underlines the importance of using a disease-specific panel with a sufficiently high coverage.

Genetic diagnosis and clinical evaluation of severe fetal akinesia syndrome.

OBJECTIVE: In this retrospective study, we describe the clinical course, ultrasound findings and genetic investigations of fetuses affected by fetal akinesia. MATERIALS AND METHODS: We enrolled 22 eukaryotic fetuses of 18 families, diagnosed with fetal akinesia between 2008 and 2016 at the Department of Obstetrics and Feto-Maternal Medicine at the Medical University of Vienna. Routine genetic evaluation included karyotyping and chromosomal microarray analysis. Retrospectively, exome sequencing was performed in the index case of 11 families, if stored DNA was available. Confirmation analyses and genetic diagnosis of siblings were performed by using Sanger sequencing. RESULTS: Whole exome sequencing identified pathogenic variants of CNTN1, RYR1, NEB, GLDN, HRAS and TNNT3 in six cases of 11 families. In three of these families, the variants were confirmed in the respective sibling. CONCLUSIONS: The present study demonstrates a high diagnostic yield of exome sequencing in fetuses affected by akinesia syndrome, especially if family history is positive. Still, in a large part the underlying genetic cause remained unknown, whereas precise clinical evaluation in combination with exome sequencing shows to be the best tool to find the disease causing variants.

Genetic association in female stress urinary incontinence based on proteomic findings: a case-control study.

INTRODUCTION AND HYPOTHESIS: Previous studies have indicated a hereditary component of stress urinary incontinence; however, evidence on candidate genes or single-nucleotide polymorphisms (SNPs) is scarce. We hypothesize a genetic association of female stress urinary incontinence based on significant differences of the urinary and serum proteomic pattern in the identical study population. METHODS: Case-control study of 19 patients and 19 controls. We searched for known SNPs of SUI candidate genes (COL1A1, MMP1, SERPINA5, UMOD) in the database of short genetic variations and PubMed. Genomic DNA was isolated using QIAamp DNA Blood Midi Kit (Qiagen). We performed Sanger sequencing of selected exons and introns. RESULTS: The rs885786 SNP of the SERPINA5 gene was identified in 15 cases and 10 controls (p = 0.09). The rs6113 SNP of the SERPINA5 gene was present in 4 controls compared to 0 cases (p = 0.105). The rs4293393, rs13333226 and rs13335818 SNPs of the UMOD gene were identified in five cases and two controls (p = 0.20), the rs1800012 SNP of the COL1A1 gene in five cases versus four controls (p = 0.24) and the homozygous rs1799750 SNP of the MMP1 gene in eight cases versus five controls (p = 0.18). The combination of the rs885786 SNP of the SERPINA5 gene and rs179970 SNP of the MMP1 gene was detected in ten cases versus five controls (p = 0.072). CONCLUSIONS: We found nonsignificant trends toward associations of SNPs on the SERPINA5, UMOD and MMP1 gene and SUI.