Development of a semi-conductor sequencing-based panel for genotyping of colon and lung cancer by the Onconetwork consortium

Bastiaan B J Tops; Nicola Normanno; Henriette Kurth; Eliana Amato; Andrea Mafficini; Nora Rieber; Delphine Le Corre; Anna Maria Rachiglio; Anne Reiman; Orla Sheils; Christoph Noppen; Ludovic Lacroix; Ian A Cree; Aldo Scarpa; Marjolijn J L Ligtenberg; Pierre Laurent-Puig

doi:10.1186/s12885-015-1015-5

Development of a semi-conductor sequencing-based panel for genotyping of colon and lung cancer by the Onconetwork consortium

Bastiaan B J Tops, Nicola Normanno, Henriette Kurth, Eliana Amato, Andrea Mafficini, Nora Rieber, Delphine Le Corre, Anna Maria Rachiglio, Anne Reiman, Orla Sheils, Christoph Noppen, Ludovic Lacroix, Ian A Cree, Aldo Scarpa, Marjolijn J L Ligtenberg, Pierre Laurent-Puig

Research output: Contribution to journal › Article › peer-review

45 Citations (Scopus)

199 Downloads (Pure)

Abstract

BACKGROUND: The number of predictive biomarkers that will be necessary to assess in clinical practice will increase with the availability of drugs that target specific molecular alterations. Therefore, diagnostic laboratories are confronted with new challenges: costs, turn-around-time and the amount of material required for testing will increase with the number of tests performed on a sample. Our consortium of European clinical research laboratories set out to test if semi-conductor sequencing provides a solution for these challenges.

METHODS: We designed a multiplex PCR targeting 87 hotspot regions in 22 genes that are of clinical interest for lung and/or colorectal cancer. The gene-panel was tested by 7 different labs in their own clinical setting using ion-semiconductor sequencing.

RESULTS: We analyzed 155 samples containing 112 previously identified mutations in the KRAS, EGFR en BRAF genes. Only 1 sample failed analysis due to poor quality of the DNA. All other samples were correctly genotyped for the known mutations, even as low as 2%, but also revealed other mutations. Optimization of the primers used in the multiplex PCR resulted in a uniform coverage distribution over the amplicons that allows for efficient pooling of samples in a sequencing run.

CONCLUSIONS: We show that a semi-conductor based sequencing approach to stratify colon and lung cancer patients is feasible in a clinical setting.

Original language	English
Pages (from-to)	26
Journal	BMC Cancer
Volume	15
DOIs	https://doi.org/10.1186/s12885-015-1015-5
Publication status	Published - 31 Jan 2015
Externally published	Yes

Keywords

Colonic Neoplasms
DNA Mutational Analysis
Genotyping Techniques
High-Throughput Nucleotide Sequencing
Humans
Lung Neoplasms
Multiplex Polymerase Chain Reaction
Mutation
Mutation Rate
Reproducibility of Results
Journal Article
Research Support, Non-U.S. Gov't

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

Access to Document

10.1186/s12885-015-1015-5Licence: CC BY

DevelopmentFinal published version, 273 KBLicence: CC BY

Anne Reiman
- CEES School of Science - Assistant Professor Academic
Person: Teaching and Research

Cite this

Tops, B. B. J., Normanno, N., Kurth, H., Amato, E., Mafficini, A., Rieber, N., Le Corre, D., Rachiglio, A. M., Reiman, A., Sheils, O., Noppen, C., Lacroix, L., Cree, I. A., Scarpa, A., Ligtenberg, M. J. L., & Laurent-Puig, P. (2015). Development of a semi-conductor sequencing-based panel for genotyping of colon and lung cancer by the Onconetwork consortium. BMC Cancer, 15, 26. https://doi.org/10.1186/s12885-015-1015-5

Tops, BBJ, Normanno, N, Kurth, H, Amato, E, Mafficini, A, Rieber, N, Le Corre, D, Rachiglio, AM, Reiman, A, Sheils, O, Noppen, C, Lacroix, L, Cree, IA, Scarpa, A, Ligtenberg, MJL & Laurent-Puig, P 2015, 'Development of a semi-conductor sequencing-based panel for genotyping of colon and lung cancer by the Onconetwork consortium', BMC Cancer, vol. 15, pp. 26. https://doi.org/10.1186/s12885-015-1015-5

@article{6d2ccb262a0d4e5286d2d0434fee7012,

title = "Development of a semi-conductor sequencing-based panel for genotyping of colon and lung cancer by the Onconetwork consortium",

abstract = "BACKGROUND: The number of predictive biomarkers that will be necessary to assess in clinical practice will increase with the availability of drugs that target specific molecular alterations. Therefore, diagnostic laboratories are confronted with new challenges: costs, turn-around-time and the amount of material required for testing will increase with the number of tests performed on a sample. Our consortium of European clinical research laboratories set out to test if semi-conductor sequencing provides a solution for these challenges.METHODS: We designed a multiplex PCR targeting 87 hotspot regions in 22 genes that are of clinical interest for lung and/or colorectal cancer. The gene-panel was tested by 7 different labs in their own clinical setting using ion-semiconductor sequencing.RESULTS: We analyzed 155 samples containing 112 previously identified mutations in the KRAS, EGFR en BRAF genes. Only 1 sample failed analysis due to poor quality of the DNA. All other samples were correctly genotyped for the known mutations, even as low as 2%, but also revealed other mutations. Optimization of the primers used in the multiplex PCR resulted in a uniform coverage distribution over the amplicons that allows for efficient pooling of samples in a sequencing run.CONCLUSIONS: We show that a semi-conductor based sequencing approach to stratify colon and lung cancer patients is feasible in a clinical setting.",

keywords = "Colonic Neoplasms, DNA Mutational Analysis, Genotyping Techniques, High-Throughput Nucleotide Sequencing, Humans, Lung Neoplasms, Multiplex Polymerase Chain Reaction, Mutation, Mutation Rate, Reproducibility of Results, Journal Article, Research Support, Non-U.S. Gov't",

author = "Tops, {Bastiaan B J} and Nicola Normanno and Henriette Kurth and Eliana Amato and Andrea Mafficini and Nora Rieber and {Le Corre}, Delphine and Rachiglio, {Anna Maria} and Anne Reiman and Orla Sheils and Christoph Noppen and Ludovic Lacroix and Cree, {Ian A} and Aldo Scarpa and Ligtenberg, {Marjolijn J L} and Pierre Laurent-Puig",

year = "2015",

month = jan,

day = "31",

doi = "10.1186/s12885-015-1015-5",

language = "English",

volume = "15",

pages = "26",

journal = "BMC Cancer",

issn = "1471-2407",

publisher = "BioMed Central",

}

TY - JOUR

T1 - Development of a semi-conductor sequencing-based panel for genotyping of colon and lung cancer by the Onconetwork consortium

AU - Tops, Bastiaan B J

AU - Normanno, Nicola

AU - Kurth, Henriette

AU - Amato, Eliana

AU - Mafficini, Andrea

AU - Rieber, Nora

AU - Le Corre, Delphine

AU - Rachiglio, Anna Maria

AU - Reiman, Anne

AU - Sheils, Orla

AU - Noppen, Christoph

AU - Lacroix, Ludovic

AU - Cree, Ian A

AU - Scarpa, Aldo

AU - Ligtenberg, Marjolijn J L

AU - Laurent-Puig, Pierre

PY - 2015/1/31

Y1 - 2015/1/31

N2 - BACKGROUND: The number of predictive biomarkers that will be necessary to assess in clinical practice will increase with the availability of drugs that target specific molecular alterations. Therefore, diagnostic laboratories are confronted with new challenges: costs, turn-around-time and the amount of material required for testing will increase with the number of tests performed on a sample. Our consortium of European clinical research laboratories set out to test if semi-conductor sequencing provides a solution for these challenges.METHODS: We designed a multiplex PCR targeting 87 hotspot regions in 22 genes that are of clinical interest for lung and/or colorectal cancer. The gene-panel was tested by 7 different labs in their own clinical setting using ion-semiconductor sequencing.RESULTS: We analyzed 155 samples containing 112 previously identified mutations in the KRAS, EGFR en BRAF genes. Only 1 sample failed analysis due to poor quality of the DNA. All other samples were correctly genotyped for the known mutations, even as low as 2%, but also revealed other mutations. Optimization of the primers used in the multiplex PCR resulted in a uniform coverage distribution over the amplicons that allows for efficient pooling of samples in a sequencing run.CONCLUSIONS: We show that a semi-conductor based sequencing approach to stratify colon and lung cancer patients is feasible in a clinical setting.

AB - BACKGROUND: The number of predictive biomarkers that will be necessary to assess in clinical practice will increase with the availability of drugs that target specific molecular alterations. Therefore, diagnostic laboratories are confronted with new challenges: costs, turn-around-time and the amount of material required for testing will increase with the number of tests performed on a sample. Our consortium of European clinical research laboratories set out to test if semi-conductor sequencing provides a solution for these challenges.METHODS: We designed a multiplex PCR targeting 87 hotspot regions in 22 genes that are of clinical interest for lung and/or colorectal cancer. The gene-panel was tested by 7 different labs in their own clinical setting using ion-semiconductor sequencing.RESULTS: We analyzed 155 samples containing 112 previously identified mutations in the KRAS, EGFR en BRAF genes. Only 1 sample failed analysis due to poor quality of the DNA. All other samples were correctly genotyped for the known mutations, even as low as 2%, but also revealed other mutations. Optimization of the primers used in the multiplex PCR resulted in a uniform coverage distribution over the amplicons that allows for efficient pooling of samples in a sequencing run.CONCLUSIONS: We show that a semi-conductor based sequencing approach to stratify colon and lung cancer patients is feasible in a clinical setting.

KW - Colonic Neoplasms

KW - DNA Mutational Analysis

KW - Genotyping Techniques

KW - High-Throughput Nucleotide Sequencing

KW - Humans

KW - Lung Neoplasms

KW - Multiplex Polymerase Chain Reaction

KW - Mutation

KW - Mutation Rate

KW - Reproducibility of Results

KW - Journal Article

KW - Research Support, Non-U.S. Gov't

U2 - 10.1186/s12885-015-1015-5

DO - 10.1186/s12885-015-1015-5

M3 - Article

C2 - 25637035

SN - 1471-2407

VL - 15

SP - 26

JO - BMC Cancer

JF - BMC Cancer

ER -

Development of a semi-conductor sequencing-based panel for genotyping of colon and lung cancer by the Onconetwork consortium

Abstract

Keywords

UN SDGs

Access to Document

Fingerprint

Profiles

Anne Reiman

Cite this