Implementation of next generation sequencing technology for somatic mutation detection in routine laboratory practice

Giardina, Tindaro; Robinson, Cleo; Grieu-Iacopetta, Fabienne; Millward, Michael; Iacopetta, Barry; Spagnolo, Dominic V.; Amanuel, Benhur

doi:10.1016/j.pathol.2018.01.005

Cited by 32 publications

(19 citation statements)

References 20 publications

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“…On the other hand, the cancer tissue was almost exhausted after the H&E and IHC staining of the specimens in cases of rebiopsy, so there was no tissue left for further EGFR T790M testing, and ctDNA had to be used as a surrogate for determination of EGFR status. Unfortunately, because every liquid biopsy technique had its own limitations which lead to a certain percentage of false negative results, cancer tissue remains the preferred alternative for EGFR T790M testing by expert consensus so far, although the intratumor heterogeneity may affect the accuracy of the final results and is also not perfect for biopsy tissue . A maximum of three sections of tumor tissue were sufficient for BaseScope T790M assay, leading to a decrease in the quantity of tissue slides required for gene mutation testing compare with the classical techniques.…”

Section: Discussionmentioning

confidence: 99%

EGFR T790M detection in formalin‐fixed paraffin‐embedded tissues of patients with lung cancer using RNA‐based in situ hybridization: A preliminary feasibility study

Shi

et al. 2019

Thoracic Cancer

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Background Following drug resistance in patients with lung cancer treated by EGFR TKIs, a biopsy is required to obtain sufficient cancer tissue for T790M detection in order to select potential beneficiaries suitable for third‐generation EGFR TKIs, such as osimertinib. The purpose of this study was to explore the feasibility of using a new in situ analysis technique based on RNA target sequences to detect EGFR T790M in lung cancer. Methods A total of 28 formalin‐fixed paraffin‐embedded (FFPE) samples from 24 lung adenocarcinoma patients archived in Peking Union Medical College Hospital from 2015 to 2016 were collected. The BaseScope T790M detection technique by in situ hybridization on FFPE slides was used to analyze the mutation of EGFR T790M, and then the results were compared with the data acquired by Scorpions ARMS assay, which is the so‐called gold standard for EGFR gene mutation testing. The sensitivity and specificity of the BaseScope T790M detection technique were preliminarily evaluated. Results Of the 28 FFPE specimens, the average proportion of T790M‐positive cells was 35.78% ± 17.68% in 18 samples with EGFR T790M, confirmed by Scorpions ARMS assay, Compared with real‐time PCR assay, the sensitivity and specificity of BaseScope T790M were all 100% in our cohort. Conclusion BaseScope T790M assay could be completed on only one FFPE slide and the visualized molecular result overplayed with histomorphological information perfectly, so it may be the alternative method for EGFR T790M evaluation. BaseScope assay has potential clinical utility, and it will be necessary to carry out validation with a large number of cases.

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Section: Discussionmentioning

confidence: 99%

EGFR T790M detection in formalin‐fixed paraffin‐embedded tissues of patients with lung cancer using RNA‐based in situ hybridization: A preliminary feasibility study

Shi

et al. 2019

Thoracic Cancer

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“…This agrees with most recent study results that reveal that the TST 26 panel can detect somatic alterations with VAF ‡3%. 35 However, Giardina et al 36 demonstrated a detectable VAF less than 3% with this gene panel. Several studies focused on the comparison of pre-and postsurgical ctDNA reported decreased frequency of mutated variants after tumor resection in various cancers.…”

Section: Discussionmentioning

confidence: 78%

Comparison of Somatic Mutation Profiles Between Formalin-Fixed Paraffin Embedded Tissues and Plasma Cell-Free DNA from Ovarian Cancer Patients Before and After Surgery

Jagelková

Zelinová

Laučeková

et al. 2020

BioResearch Open Access

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Ovarian carcinogenesis can be induced by a large number of somatic gene mutations. Circulating tumor DNA (ctDNA) released into peripheral blood can provide insights into the genomic landscape of cancer cells and monitor their dynamics. Our aim was to detect and compare the genetic profiles in tumor tissue and plasma before and after tumor resection in ovarian cancer patients. All three samples were collected from each patient. In this study, we used a commercial cancer panel to identify somatic mutations in 26 genes in seven selected patients through next-generation sequencing on the Illumina platform. Overall, 16 variants with pathogenic effect were identified in the TP53, PIK3CA, PTEN, APC, NRAS, KRAS, GNAS, and MET genes involved in important signaling pathways. The genetic alterations found in the presurgical plasma in six of seven ovarian cancer patients were no longer present in the plasma after tumor surgical removal. Identical variants in formalin-fixed paraffin embedded (FFPE) tissues and preoperative plasma specimens were observed in only two cases. These findings suggest that the detected presurgical pathogenic variants absent in postsurgery plasma are associated with the primary ovarian tumor. Finally, the low-identified concordance between FFPE and plasma can be due to various factors, but most likely to high tumor heterogeneity and low ctDNA level.

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“…In contrast, another investigation has directly focused on a concrete cancer type such as in nonsmall cell lung cancer (NSCLC) (Legras et al, 2018). Although several studies validated NGSbased panels on both solid tumors and hematologic malignancies (Cottrell et al, 2014;Garcia et al, 2017), few reports have aimed to demonstrate that the TsT26 panel is a validated method to implement in the clinical practice in a considerable number of varied tumor tissues (Fisher et al, 2016;Giardina et al, 2018). In addition, this panel has also been used to validate another molecular testing platform in 90 NSCLC tumor samples (Quinn et al, 2015).…”

Section: Discussionmentioning

confidence: 99%

Peer Review #1 of "Validation and clinical application of a targeted next-generation sequencing gene panel for solid and hematologic malignancies (v0.1)"

Wallace¹

2020

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Background: Next-generation sequencing (NGS) is a high-throughput technology that has become widely integrated in molecular diagnostics laboratories. Among the large diversity of NGS-based panels, the Trusight Tumor 26 (TsT26) enables the detection of lowfrequency variants across 26 genes using the MiSeq platform. Methods: We describe the inter-laboratory validation and subsequent clinical application of the panel in 399 patients presenting a range of tumor types, including gastrointestinal (GI, 29%), hematologic (18%), lung (13%), gynecological and breast (8% each), among others. Results: The panel is highly accurate with a test sensitivity of 92%, and demonstrated high specificity and positive predictive values (95% and 96%, respectively). Sequencing testing was successful in two-thirds of patients, while the remaining third failed due to unsuccessful qualitycontrol filtering. Most detected variants were observed in the TP53 (28%), KRAS (16%), APC (10%) and PIK3CA (8%) genes. Overall, 372 variants were identified, primarily distributed as missense (81%), stop gain (9%) and frameshift (7%) altered sequences and mostly reported as pathogenic (78%) and variants of uncertain significance (19%). Only 14% of patients received targeted treatment based on the variant determined by the panel. The variants most frequently observed in GI and lung tumors were: KRASc.35G>A (p.G12D), c.35G>T (p.G12V) and c.34G>T (p.G12C). Conclusions: Prior panel validation allowed its use in the laboratory daily practice by providing several relevant and potentially targetable variants across multiple tumors. However, this study is limited by high sample inadequacy rate, raising doubts as to continuity in the clinical setting.

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Implementation of next generation sequencing technology for somatic mutation detection in routine laboratory practice

Cited by 32 publications

References 20 publications

EGFR T790M detection in formalin‐fixed paraffin‐embedded tissues of patients with lung cancer using RNA‐based in situ hybridization: A preliminary feasibility study

EGFR T790M detection in formalin‐fixed paraffin‐embedded tissues of patients with lung cancer using RNA‐based in situ hybridization: A preliminary feasibility study

Comparison of Somatic Mutation Profiles Between Formalin-Fixed Paraffin Embedded Tissues and Plasma Cell-Free DNA from Ovarian Cancer Patients Before and After Surgery

Peer Review #1 of "Validation and clinical application of a targeted next-generation sequencing gene panel for solid and hematologic malignancies (v0.1)"

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