Quantitative reverse transcription-polymerase chain reaction measurement of HASH1 (ASCL1), a marker for small cell lung carcinomas with neuroendocrine features

Bart A Westerman, Sari Neijenhuis, Ankie Poutsma, Renske D M Steenbergen, Roderick H J Breuer, Monique Egging, Inge J van Wijk, Cees B M Oudejans

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

PURPOSE: The Human Achaete-Scute homologue 1 (HASH1, ASCL1), a lineage-specific basic helix-loop-helix member of the achaete-scute family, is essential for the generation of pulmonary neuroendocrine (NE) cells during lung development. In small cell lung cancer (SCLC), the most lethal form of lung cancer, the gene is highly expressed and the expression of HASH1 correlates with NE features found in SCLCs. Here we describe a highly sensitive reverse transcription-PCR method for quantifying HASH1 mRNA in clinical samples, using real-time fluorescence resonance energy transfer technology (LightCycler).

EXPERIMENTAL DESIGN: The HASH1-positive NE cell line NCI-H187 was compared with the non-NE cell line NCI-N417 by quantitative reverse transcription-PCR. Signals were normalized using the housekeeping gene PBGD, which is pseudogene free. Subsequently, HASH1 expression in RNA isolated from biopsies from SCLC patients (n = 4) was compared with biopsies from non-SCLC (NSCLC) patients (n = 2) or normal bronchus (n = 2).

RESULTS: The HASH1-positive NE cell line NCI-H187 showed 50,000-fold higher normalized expression of HASH1 than did the non-NE cell line NCI-N417, indicating that the method is applicable over a wide dynamic range. Normalized average mRNA expression levels in SCLC clinical samples were 1,000-fold higher than in the NSCLC samples. Expression in normal bronchus was comparable to the expression levels in the NSCLC.

CONCLUSIONS: These results show that marked and measurable differences exist between SCLCs and other lung tissues (either NSCLC or normal bronchus). We show that the method is applicable to small biopsy samples and can discriminate SCLC from NSCLC. This method could contribute to diagnosis based on molecular profiling of tumors.

Original languageEnglish
Pages (from-to)1082-6
Number of pages5
JournalClinical Cancer Research
Volume8
Issue number4
Publication statusPublished - Apr 2002

Cite this

Westerman, Bart A ; Neijenhuis, Sari ; Poutsma, Ankie ; Steenbergen, Renske D M ; Breuer, Roderick H J ; Egging, Monique ; van Wijk, Inge J ; Oudejans, Cees B M. / Quantitative reverse transcription-polymerase chain reaction measurement of HASH1 (ASCL1), a marker for small cell lung carcinomas with neuroendocrine features. In: Clinical Cancer Research. 2002 ; Vol. 8, No. 4. pp. 1082-6.
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title = "Quantitative reverse transcription-polymerase chain reaction measurement of HASH1 (ASCL1), a marker for small cell lung carcinomas with neuroendocrine features",
abstract = "PURPOSE: The Human Achaete-Scute homologue 1 (HASH1, ASCL1), a lineage-specific basic helix-loop-helix member of the achaete-scute family, is essential for the generation of pulmonary neuroendocrine (NE) cells during lung development. In small cell lung cancer (SCLC), the most lethal form of lung cancer, the gene is highly expressed and the expression of HASH1 correlates with NE features found in SCLCs. Here we describe a highly sensitive reverse transcription-PCR method for quantifying HASH1 mRNA in clinical samples, using real-time fluorescence resonance energy transfer technology (LightCycler).EXPERIMENTAL DESIGN: The HASH1-positive NE cell line NCI-H187 was compared with the non-NE cell line NCI-N417 by quantitative reverse transcription-PCR. Signals were normalized using the housekeeping gene PBGD, which is pseudogene free. Subsequently, HASH1 expression in RNA isolated from biopsies from SCLC patients (n = 4) was compared with biopsies from non-SCLC (NSCLC) patients (n = 2) or normal bronchus (n = 2).RESULTS: The HASH1-positive NE cell line NCI-H187 showed 50,000-fold higher normalized expression of HASH1 than did the non-NE cell line NCI-N417, indicating that the method is applicable over a wide dynamic range. Normalized average mRNA expression levels in SCLC clinical samples were 1,000-fold higher than in the NSCLC samples. Expression in normal bronchus was comparable to the expression levels in the NSCLC.CONCLUSIONS: These results show that marked and measurable differences exist between SCLCs and other lung tissues (either NSCLC or normal bronchus). We show that the method is applicable to small biopsy samples and can discriminate SCLC from NSCLC. This method could contribute to diagnosis based on molecular profiling of tumors.",
keywords = "Basic Helix-Loop-Helix Transcription Factors, Biomarkers, Tumor/genetics, Carcinoma, Neuroendocrine/genetics, Carcinoma, Non-Small-Cell Lung/genetics, Carcinoma, Small Cell/genetics, DNA-Binding Proteins/genetics, Gene Expression Regulation, Neoplastic, Humans, Hydroxymethylbilane Synthase/genetics, Lung Neoplasms/genetics, RNA, Complementary/genetics, RNA, Messenger/genetics, RNA, Neoplasm/genetics, Reverse Transcriptase Polymerase Chain Reaction, Transcription Factors/genetics, Tumor Cells, Cultured, Up-Regulation",
author = "Westerman, {Bart A} and Sari Neijenhuis and Ankie Poutsma and Steenbergen, {Renske D M} and Breuer, {Roderick H J} and Monique Egging and {van Wijk}, {Inge J} and Oudejans, {Cees B M}",
year = "2002",
month = "4",
language = "English",
volume = "8",
pages = "1082--6",
journal = "Clinical Cancer Research",
issn = "1078-0432",
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Quantitative reverse transcription-polymerase chain reaction measurement of HASH1 (ASCL1), a marker for small cell lung carcinomas with neuroendocrine features. / Westerman, Bart A; Neijenhuis, Sari; Poutsma, Ankie; Steenbergen, Renske D M; Breuer, Roderick H J; Egging, Monique; van Wijk, Inge J; Oudejans, Cees B M.

In: Clinical Cancer Research, Vol. 8, No. 4, 04.2002, p. 1082-6.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - Quantitative reverse transcription-polymerase chain reaction measurement of HASH1 (ASCL1), a marker for small cell lung carcinomas with neuroendocrine features

AU - Westerman, Bart A

AU - Neijenhuis, Sari

AU - Poutsma, Ankie

AU - Steenbergen, Renske D M

AU - Breuer, Roderick H J

AU - Egging, Monique

AU - van Wijk, Inge J

AU - Oudejans, Cees B M

PY - 2002/4

Y1 - 2002/4

N2 - PURPOSE: The Human Achaete-Scute homologue 1 (HASH1, ASCL1), a lineage-specific basic helix-loop-helix member of the achaete-scute family, is essential for the generation of pulmonary neuroendocrine (NE) cells during lung development. In small cell lung cancer (SCLC), the most lethal form of lung cancer, the gene is highly expressed and the expression of HASH1 correlates with NE features found in SCLCs. Here we describe a highly sensitive reverse transcription-PCR method for quantifying HASH1 mRNA in clinical samples, using real-time fluorescence resonance energy transfer technology (LightCycler).EXPERIMENTAL DESIGN: The HASH1-positive NE cell line NCI-H187 was compared with the non-NE cell line NCI-N417 by quantitative reverse transcription-PCR. Signals were normalized using the housekeeping gene PBGD, which is pseudogene free. Subsequently, HASH1 expression in RNA isolated from biopsies from SCLC patients (n = 4) was compared with biopsies from non-SCLC (NSCLC) patients (n = 2) or normal bronchus (n = 2).RESULTS: The HASH1-positive NE cell line NCI-H187 showed 50,000-fold higher normalized expression of HASH1 than did the non-NE cell line NCI-N417, indicating that the method is applicable over a wide dynamic range. Normalized average mRNA expression levels in SCLC clinical samples were 1,000-fold higher than in the NSCLC samples. Expression in normal bronchus was comparable to the expression levels in the NSCLC.CONCLUSIONS: These results show that marked and measurable differences exist between SCLCs and other lung tissues (either NSCLC or normal bronchus). We show that the method is applicable to small biopsy samples and can discriminate SCLC from NSCLC. This method could contribute to diagnosis based on molecular profiling of tumors.

AB - PURPOSE: The Human Achaete-Scute homologue 1 (HASH1, ASCL1), a lineage-specific basic helix-loop-helix member of the achaete-scute family, is essential for the generation of pulmonary neuroendocrine (NE) cells during lung development. In small cell lung cancer (SCLC), the most lethal form of lung cancer, the gene is highly expressed and the expression of HASH1 correlates with NE features found in SCLCs. Here we describe a highly sensitive reverse transcription-PCR method for quantifying HASH1 mRNA in clinical samples, using real-time fluorescence resonance energy transfer technology (LightCycler).EXPERIMENTAL DESIGN: The HASH1-positive NE cell line NCI-H187 was compared with the non-NE cell line NCI-N417 by quantitative reverse transcription-PCR. Signals were normalized using the housekeeping gene PBGD, which is pseudogene free. Subsequently, HASH1 expression in RNA isolated from biopsies from SCLC patients (n = 4) was compared with biopsies from non-SCLC (NSCLC) patients (n = 2) or normal bronchus (n = 2).RESULTS: The HASH1-positive NE cell line NCI-H187 showed 50,000-fold higher normalized expression of HASH1 than did the non-NE cell line NCI-N417, indicating that the method is applicable over a wide dynamic range. Normalized average mRNA expression levels in SCLC clinical samples were 1,000-fold higher than in the NSCLC samples. Expression in normal bronchus was comparable to the expression levels in the NSCLC.CONCLUSIONS: These results show that marked and measurable differences exist between SCLCs and other lung tissues (either NSCLC or normal bronchus). We show that the method is applicable to small biopsy samples and can discriminate SCLC from NSCLC. This method could contribute to diagnosis based on molecular profiling of tumors.

KW - Basic Helix-Loop-Helix Transcription Factors

KW - Biomarkers, Tumor/genetics

KW - Carcinoma, Neuroendocrine/genetics

KW - Carcinoma, Non-Small-Cell Lung/genetics

KW - Carcinoma, Small Cell/genetics

KW - DNA-Binding Proteins/genetics

KW - Gene Expression Regulation, Neoplastic

KW - Humans

KW - Hydroxymethylbilane Synthase/genetics

KW - Lung Neoplasms/genetics

KW - RNA, Complementary/genetics

KW - RNA, Messenger/genetics

KW - RNA, Neoplasm/genetics

KW - Reverse Transcriptase Polymerase Chain Reaction

KW - Transcription Factors/genetics

KW - Tumor Cells, Cultured

KW - Up-Regulation

M3 - Article

VL - 8

SP - 1082

EP - 1086

JO - Clinical Cancer Research

JF - Clinical Cancer Research

SN - 1078-0432

IS - 4

ER -