Dataset Information

Harmonization of Next-Generation Sequencing Procedure in Italian Laboratories: A Multi-Institutional Evaluation of the SiRe® Panel.

ABSTRACT: Background: Next-generation sequencing (NGS) needs to be validated and standardized to ensure that cancer patients are reliably selected for target treatments. In Italy, NGS is performed in several institutions and harmonization of wet and dry procedures is needed. To this end, a consortium of five different laboratories, covering the most part of the Italian peninsula, was constituted. A narrow gene panel (SiRe®) covering 568 clinically relevant mutations in six different genes (EGFR, KRAS, NRAS, BRAF, cKIT, and PDGFR?) with a predictive role for therapy selection in non-small cell lung cancer (NSCLC), gastrointestinal stromal tumor, colorectal carcinoma (CRC), and melanoma was evaluated in each participating laboratory. Methods: To assess the NGS inter-laboratory concordance, the SiRe® panel, with a related kit and protocol for library preparation, was used in each center to analyze a common set of 20 NSCLC and CRC routine samples. Concordance rate, in terms of mutation detected and relative allelic frequencies, was assessed. Then, each institution prospectively analyzed an additional set of 40 routine samples (for a total of 160 specimens) to assess the reproducibility of the NGS run parameters in each institution. Results: An inter-laboratory agreement of 100% was reached in analyzing the data obtained from the 20 common sample sets; the concordance rate of allelic frequencies distribution was 0.989. The prospective analysis of the run metric parameters obtained by each center locally showed that the analytical performance of the SiRe® panel in the different institutions was highly reproducible. Conclusions: The SiRe® panel represents a robust diagnostic tool to harmonize the NGS procedure in different Italian laboratories.

SUBMITTER: Malapelle U

PROVIDER: S-EPMC7078327 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Publications

Harmonization of Next-Generation Sequencing Procedure in Italian Laboratories: A Multi-Institutional Evaluation of the SiRe® Panel.

Malapelle Umberto U Pepe Francesco F Pisapia Pasquale P Sgariglia Roberta R Nacchio Mariantonia M De Luca Caterina C Lacalamita Rosanna R Tommasi Stefania S Pinto Rosamaria R Palomba Grazia G Palmieri Giuseppe G Vacirca Davide D Barberis Massimo M Bottillo Irene I Grammatico Paola P Grillo Lucia Rosalba LR Costa Valerio V Smeraglio Riccardo R Bruzzese Dario D Troncone Giancarlo G

Frontiers in oncology 20200311

<b>Background:</b> Next-generation sequencing (NGS) needs to be validated and standardized to ensure that cancer patients are reliably selected for target treatments. In Italy, NGS is performed in several institutions and harmonization of wet and dry procedures is needed. To this end, a consortium of five different laboratories, covering the most part of the Italian peninsula, was constituted. A narrow gene panel (SiRe®) covering 568 clinically relevant mutations in six different genes (<i>EGFR, ...[more]

PMID: 32219061

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Project description:BackgroundHigh-throughput Next-Generation Sequencing (NGS) techniques are advancing genomics and molecular biology research. This technology generates substantially large data which puts up a major challenge to the scientists for an efficient, cost and time effective solution to analyse such data. Further, for the different types of NGS data, there are certain common challenging steps involved in analysing those data. Spliced alignment is one such fundamental step in NGS data analysis which is extremely computational intensive as well as time consuming. There exists serious problem even with the most widely used spliced alignment tools. TopHat is one such widely used spliced alignment tools which although supports multithreading, does not efficiently utilize computational resources in terms of CPU utilization and memory. Here we have introduced PVT (Pipelined Version of TopHat) where we take up a modular approach by breaking TopHat's serial execution into a pipeline of multiple stages, thereby increasing the degree of parallelization and computational resource utilization. Thus we address the discrepancies in TopHat so as to analyze large NGS data efficiently.ResultsWe analysed the SRA dataset (SRX026839 and SRX026838) consisting of single end reads and SRA data SRR1027730 consisting of paired-end reads. We used TopHat v2.0.8 to analyse these datasets and noted the CPU usage, memory footprint and execution time during spliced alignment. With this basic information, we designed PVT, a pipelined version of TopHat that removes the redundant computational steps during 'spliced alignment' and breaks the job into a pipeline of multiple stages (each comprising of different step(s)) to improve its resource utilization, thus reducing the execution time.ConclusionsPVT provides an improvement over TopHat for spliced alignment of NGS data analysis. PVT thus resulted in the reduction of the execution time to ~23% for the single end read dataset. Further, PVT designed for paired end reads showed an improved performance of ~41% over TopHat (for the chosen data) with respect to execution time. Moreover we propose PVT-Cloud which implements PVT pipeline in cloud computing system.

Dataset Information

Harmonization of Next-Generation Sequencing Procedure in Italian Laboratories: A Multi-Institutional Evaluation of the SiRe® Panel.

Publications

Harmonization of Next-Generation Sequencing Procedure in Italian Laboratories: A Multi-Institutional Evaluation of the SiRe® Panel.

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