Bruno da Silveira Corrêa , Fernanda De-Paris , Guilherme Danielski Viola , Tiago Finger Andreis , Clévia Rosset , Fernanda Sales Luiz Vianna , Luis Fernando da Rosa Rivero , Francine Hehn de Oliveira , Patricia Ashton-Prolla , Gabriel de Souza Macedo
{"title":"下一代测序在福尔马林固定石蜡包埋的非小细胞肺癌肿瘤样本中的有效性所面临的挑战","authors":"Bruno da Silveira Corrêa , Fernanda De-Paris , Guilherme Danielski Viola , Tiago Finger Andreis , Clévia Rosset , Fernanda Sales Luiz Vianna , Luis Fernando da Rosa Rivero , Francine Hehn de Oliveira , Patricia Ashton-Prolla , Gabriel de Souza Macedo","doi":"10.1016/j.anndiagpath.2023.152249","DOIUrl":null,"url":null,"abstract":"<div><h3>Introduction</h3><p>Next-generation sequencing (NGS) of Formalin-Fixed and Paraffin-Embedded (FFPE) specimens is routine in precision oncology practice. However, results are not always conclusive, and it is important to identify which factors may influence FFPE tumor sequencing success.</p></div><div><h3>Materials and methods</h3><p>Here, we evaluated the influence of pre-analytical factors on 705 samples of non-small cell lung cancer specimens that underwent NGS testing. Factors such as tumor site, tumor cell percentage, fragment size, primary tumor or metastasis, presence of necrosis, DNA purity, DNA concentration, sample origin and year of testing.</p></div><div><h3>Results</h3><p>The overall NGS success rate was 84.9 % (<em>n</em> = 599). Bone site specimens had a very low success rate (42.1 %), differing from lung samples (79.8 %) (<em>P</em> < 0.05). Samples with tumor percentages <5 % (success rate of 44.4 %) represented 14.1 % of failed sequencings. Moreover, samples with tumor percentages >10 %–20 % (82 %) did not differ from those with >30 % (88.9 %) on sequencing outcomes (<em>P</em> = 0.086). Specimens that provided DNA concentrations >2.0 ng/uL, 1.0–2.0 ng/uL, 0.5–1.0 ng/uL and <0.5 ng/uL had success rates of 92 %, 77.1 %, 61.3 % and 20.4 %, respectively. Small fragments (≤0.2 cm<sup>2</sup>) had a success rate of 74.7 % and were more prevalent in the unsuccessful group (<em>P</em> < 0.05).</p></div><div><h3>Conclusions</h3><p>Our results suggest that tumor percentage, fragment size, decalcified bone specimens, and DNA concentration are potential modifiers of NGS success rates. Interestingly, specimens with tumor percentages between 10 % and 20 % have the same sequencing outcome than specimens with >30 %. These results can strengthen the understanding of factors that lead to NGS success variability.</p></div>","PeriodicalId":50768,"journal":{"name":"Annals of Diagnostic Pathology","volume":null,"pages":null},"PeriodicalIF":1.5000,"publicationDate":"2023-12-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Challenges to the effectiveness of next-generation sequencing in formalin-fixed paraffin-embedded tumor samples for non-small cell lung cancer\",\"authors\":\"Bruno da Silveira Corrêa , Fernanda De-Paris , Guilherme Danielski Viola , Tiago Finger Andreis , Clévia Rosset , Fernanda Sales Luiz Vianna , Luis Fernando da Rosa Rivero , Francine Hehn de Oliveira , Patricia Ashton-Prolla , Gabriel de Souza Macedo\",\"doi\":\"10.1016/j.anndiagpath.2023.152249\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><h3>Introduction</h3><p>Next-generation sequencing (NGS) of Formalin-Fixed and Paraffin-Embedded (FFPE) specimens is routine in precision oncology practice. However, results are not always conclusive, and it is important to identify which factors may influence FFPE tumor sequencing success.</p></div><div><h3>Materials and methods</h3><p>Here, we evaluated the influence of pre-analytical factors on 705 samples of non-small cell lung cancer specimens that underwent NGS testing. Factors such as tumor site, tumor cell percentage, fragment size, primary tumor or metastasis, presence of necrosis, DNA purity, DNA concentration, sample origin and year of testing.</p></div><div><h3>Results</h3><p>The overall NGS success rate was 84.9 % (<em>n</em> = 599). Bone site specimens had a very low success rate (42.1 %), differing from lung samples (79.8 %) (<em>P</em> < 0.05). Samples with tumor percentages <5 % (success rate of 44.4 %) represented 14.1 % of failed sequencings. Moreover, samples with tumor percentages >10 %–20 % (82 %) did not differ from those with >30 % (88.9 %) on sequencing outcomes (<em>P</em> = 0.086). Specimens that provided DNA concentrations >2.0 ng/uL, 1.0–2.0 ng/uL, 0.5–1.0 ng/uL and <0.5 ng/uL had success rates of 92 %, 77.1 %, 61.3 % and 20.4 %, respectively. Small fragments (≤0.2 cm<sup>2</sup>) had a success rate of 74.7 % and were more prevalent in the unsuccessful group (<em>P</em> < 0.05).</p></div><div><h3>Conclusions</h3><p>Our results suggest that tumor percentage, fragment size, decalcified bone specimens, and DNA concentration are potential modifiers of NGS success rates. Interestingly, specimens with tumor percentages between 10 % and 20 % have the same sequencing outcome than specimens with >30 %. These results can strengthen the understanding of factors that lead to NGS success variability.</p></div>\",\"PeriodicalId\":50768,\"journal\":{\"name\":\"Annals of Diagnostic Pathology\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":1.5000,\"publicationDate\":\"2023-12-21\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Annals of Diagnostic Pathology\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S1092913423001478\",\"RegionNum\":4,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q3\",\"JCRName\":\"PATHOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Annals of Diagnostic Pathology","FirstCategoryId":"3","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S1092913423001478","RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"PATHOLOGY","Score":null,"Total":0}
Challenges to the effectiveness of next-generation sequencing in formalin-fixed paraffin-embedded tumor samples for non-small cell lung cancer
Introduction
Next-generation sequencing (NGS) of Formalin-Fixed and Paraffin-Embedded (FFPE) specimens is routine in precision oncology practice. However, results are not always conclusive, and it is important to identify which factors may influence FFPE tumor sequencing success.
Materials and methods
Here, we evaluated the influence of pre-analytical factors on 705 samples of non-small cell lung cancer specimens that underwent NGS testing. Factors such as tumor site, tumor cell percentage, fragment size, primary tumor or metastasis, presence of necrosis, DNA purity, DNA concentration, sample origin and year of testing.
Results
The overall NGS success rate was 84.9 % (n = 599). Bone site specimens had a very low success rate (42.1 %), differing from lung samples (79.8 %) (P < 0.05). Samples with tumor percentages <5 % (success rate of 44.4 %) represented 14.1 % of failed sequencings. Moreover, samples with tumor percentages >10 %–20 % (82 %) did not differ from those with >30 % (88.9 %) on sequencing outcomes (P = 0.086). Specimens that provided DNA concentrations >2.0 ng/uL, 1.0–2.0 ng/uL, 0.5–1.0 ng/uL and <0.5 ng/uL had success rates of 92 %, 77.1 %, 61.3 % and 20.4 %, respectively. Small fragments (≤0.2 cm2) had a success rate of 74.7 % and were more prevalent in the unsuccessful group (P < 0.05).
Conclusions
Our results suggest that tumor percentage, fragment size, decalcified bone specimens, and DNA concentration are potential modifiers of NGS success rates. Interestingly, specimens with tumor percentages between 10 % and 20 % have the same sequencing outcome than specimens with >30 %. These results can strengthen the understanding of factors that lead to NGS success variability.
期刊介绍:
A peer-reviewed journal devoted to the publication of articles dealing with traditional morphologic studies using standard diagnostic techniques and stressing clinicopathological correlations and scientific observation of relevance to the daily practice of pathology. Special features include pathologic-radiologic correlations and pathologic-cytologic correlations.