Krüppel-Like factor 6, is frequently down-regulated and induces apoptosis in non-small cell lung cancer cells

Genshi Ito, Mika Uchiyama, Masashi Kondo, Shoichi Mori, Noriyasu Usami, Osamu Maeda, Tsutomu Kawabe, Yoshinori Hasegawa, Kaoru Shimokata, Yoshitaka Sekido

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Abstract

Krüppel-like factor 6 (KLF6) is a ubiquitously expressed zinc finger transcriptional factor, which has been suggested to be a candidate tumor suppressor gene in prostate cancer and astrocytic glioma. Because KLF6 is located at chromosome 10p15, where non-small cell lung cancers (NSCLCs) also exhibit frequent allelic loss, we hypothesized that the inactivation of KLF6 is also involved in the development of NSCLC. To determine this, we performed mutational analysis for 105 NSCLCs, including 9 cell lines and 96 primary tumors, and Northern blot analysis for 74 NSCLCs, including the 9 cell lines and 65 primary tumors. Although somatic mutations were not detected in the coding sequence of KLF6, expression of KLF6 mRNA was down-regulated in the 9 cell lines and in 55 (85%) of the 65 primary tumors compared with normal lung tissue. Treatment of two cell lines expressing KLF6 at low levels with 5-azacytidine did not induce KLF6 expression, suggesting that KLF6 down-regulation is not due to promoter hypermethylation. We also performed loss of heterozygosity (LOH) analysis using the laser capture microdissection technique, and found that 21 of 62 (34%) informative samples had LOH in the KLF6 gene locus. Comparing the LOH status with mRNA expression of KLF6, we found that 14 of the 14 (100%) samples with LOH showed KLF6 down-regulation, and that even 23 of 31 (74%) samples without LOH also showed this down-regulation. We also studied the expression of the WAF1 gene, a possible downstream gene of KLF6, and detected simultaneous down-regulation of WAF1 and KLF6 mRNA in 6 of 9 (67%) cell lines and 48 of the 55 (87%) primary tumors, although there was not a significant association between loss of KLF6 and WAF1 expression. Furthermore, colony formation assay of two NSCLC cell lines (NCI-H1299 and NCI-H2009) induced a markedly reduced colony formation by KLF6 transfection, and Annexin V staining and terminal deoxynucleotidyl transferase-mediated nick end labeling assays revealed that KLF6 induced apoptosis. Our present studies demonstrated that KLF6 is frequently down-regulated in NSCLC and suppresses tumor growth via induction of apoptosis in NSCLC, which may suggest that KLF6 is a tumor suppressor for NSCLC.

Original languageEnglish
Pages (from-to)3838-3843
Number of pages6
JournalCancer Research
Volume64
Issue number11
DOIs
Publication statusPublished - 01-06-2004

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Non-Small Cell Lung Carcinoma
Loss of Heterozygosity
Apoptosis
Cell Line
Down-Regulation
Neoplasms
Messenger RNA
Laser Capture Microdissection
Azacitidine
DNA Nucleotidylexotransferase
Annexin A5
Zinc Fingers
Astrocytoma
Tumor Suppressor Genes
Northern Blotting
Genes
Transfection
Prostatic Neoplasms
Chromosomes
Staining and Labeling

All Science Journal Classification (ASJC) codes

  • Oncology
  • Cancer Research

Cite this

Ito, Genshi ; Uchiyama, Mika ; Kondo, Masashi ; Mori, Shoichi ; Usami, Noriyasu ; Maeda, Osamu ; Kawabe, Tsutomu ; Hasegawa, Yoshinori ; Shimokata, Kaoru ; Sekido, Yoshitaka. / Krüppel-Like factor 6, is frequently down-regulated and induces apoptosis in non-small cell lung cancer cells. In: Cancer Research. 2004 ; Vol. 64, No. 11. pp. 3838-3843.
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abstract = "Kr{\"u}ppel-like factor 6 (KLF6) is a ubiquitously expressed zinc finger transcriptional factor, which has been suggested to be a candidate tumor suppressor gene in prostate cancer and astrocytic glioma. Because KLF6 is located at chromosome 10p15, where non-small cell lung cancers (NSCLCs) also exhibit frequent allelic loss, we hypothesized that the inactivation of KLF6 is also involved in the development of NSCLC. To determine this, we performed mutational analysis for 105 NSCLCs, including 9 cell lines and 96 primary tumors, and Northern blot analysis for 74 NSCLCs, including the 9 cell lines and 65 primary tumors. Although somatic mutations were not detected in the coding sequence of KLF6, expression of KLF6 mRNA was down-regulated in the 9 cell lines and in 55 (85{\%}) of the 65 primary tumors compared with normal lung tissue. Treatment of two cell lines expressing KLF6 at low levels with 5-azacytidine did not induce KLF6 expression, suggesting that KLF6 down-regulation is not due to promoter hypermethylation. We also performed loss of heterozygosity (LOH) analysis using the laser capture microdissection technique, and found that 21 of 62 (34{\%}) informative samples had LOH in the KLF6 gene locus. Comparing the LOH status with mRNA expression of KLF6, we found that 14 of the 14 (100{\%}) samples with LOH showed KLF6 down-regulation, and that even 23 of 31 (74{\%}) samples without LOH also showed this down-regulation. We also studied the expression of the WAF1 gene, a possible downstream gene of KLF6, and detected simultaneous down-regulation of WAF1 and KLF6 mRNA in 6 of 9 (67{\%}) cell lines and 48 of the 55 (87{\%}) primary tumors, although there was not a significant association between loss of KLF6 and WAF1 expression. Furthermore, colony formation assay of two NSCLC cell lines (NCI-H1299 and NCI-H2009) induced a markedly reduced colony formation by KLF6 transfection, and Annexin V staining and terminal deoxynucleotidyl transferase-mediated nick end labeling assays revealed that KLF6 induced apoptosis. Our present studies demonstrated that KLF6 is frequently down-regulated in NSCLC and suppresses tumor growth via induction of apoptosis in NSCLC, which may suggest that KLF6 is a tumor suppressor for NSCLC.",
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Ito, G, Uchiyama, M, Kondo, M, Mori, S, Usami, N, Maeda, O, Kawabe, T, Hasegawa, Y, Shimokata, K & Sekido, Y 2004, 'Krüppel-Like factor 6, is frequently down-regulated and induces apoptosis in non-small cell lung cancer cells', Cancer Research, vol. 64, no. 11, pp. 3838-3843. https://doi.org/10.1158/0008-5472.CAN-04-0185

Krüppel-Like factor 6, is frequently down-regulated and induces apoptosis in non-small cell lung cancer cells. / Ito, Genshi; Uchiyama, Mika; Kondo, Masashi; Mori, Shoichi; Usami, Noriyasu; Maeda, Osamu; Kawabe, Tsutomu; Hasegawa, Yoshinori; Shimokata, Kaoru; Sekido, Yoshitaka.

In: Cancer Research, Vol. 64, No. 11, 01.06.2004, p. 3838-3843.

Research output: Contribution to journalArticle

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T1 - Krüppel-Like factor 6, is frequently down-regulated and induces apoptosis in non-small cell lung cancer cells

AU - Ito, Genshi

AU - Uchiyama, Mika

AU - Kondo, Masashi

AU - Mori, Shoichi

AU - Usami, Noriyasu

AU - Maeda, Osamu

AU - Kawabe, Tsutomu

AU - Hasegawa, Yoshinori

AU - Shimokata, Kaoru

AU - Sekido, Yoshitaka

PY - 2004/6/1

Y1 - 2004/6/1

N2 - Krüppel-like factor 6 (KLF6) is a ubiquitously expressed zinc finger transcriptional factor, which has been suggested to be a candidate tumor suppressor gene in prostate cancer and astrocytic glioma. Because KLF6 is located at chromosome 10p15, where non-small cell lung cancers (NSCLCs) also exhibit frequent allelic loss, we hypothesized that the inactivation of KLF6 is also involved in the development of NSCLC. To determine this, we performed mutational analysis for 105 NSCLCs, including 9 cell lines and 96 primary tumors, and Northern blot analysis for 74 NSCLCs, including the 9 cell lines and 65 primary tumors. Although somatic mutations were not detected in the coding sequence of KLF6, expression of KLF6 mRNA was down-regulated in the 9 cell lines and in 55 (85%) of the 65 primary tumors compared with normal lung tissue. Treatment of two cell lines expressing KLF6 at low levels with 5-azacytidine did not induce KLF6 expression, suggesting that KLF6 down-regulation is not due to promoter hypermethylation. We also performed loss of heterozygosity (LOH) analysis using the laser capture microdissection technique, and found that 21 of 62 (34%) informative samples had LOH in the KLF6 gene locus. Comparing the LOH status with mRNA expression of KLF6, we found that 14 of the 14 (100%) samples with LOH showed KLF6 down-regulation, and that even 23 of 31 (74%) samples without LOH also showed this down-regulation. We also studied the expression of the WAF1 gene, a possible downstream gene of KLF6, and detected simultaneous down-regulation of WAF1 and KLF6 mRNA in 6 of 9 (67%) cell lines and 48 of the 55 (87%) primary tumors, although there was not a significant association between loss of KLF6 and WAF1 expression. Furthermore, colony formation assay of two NSCLC cell lines (NCI-H1299 and NCI-H2009) induced a markedly reduced colony formation by KLF6 transfection, and Annexin V staining and terminal deoxynucleotidyl transferase-mediated nick end labeling assays revealed that KLF6 induced apoptosis. Our present studies demonstrated that KLF6 is frequently down-regulated in NSCLC and suppresses tumor growth via induction of apoptosis in NSCLC, which may suggest that KLF6 is a tumor suppressor for NSCLC.

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