Effects of the demethylating agent, 5-azacytidine, on expression of the kallikrein-kinin genes in carcinoma cells of the lung and pleura

Our plain-English summary of Effects of the demethylating agent, 5-azacytidine, on expression of the kallikrein-kinin genes in carcinoma cells of the lung and pleura, by J. Wong, Y.Y. Sia, N.L. Misso, et al (Pathology Research International, 2011).

The study

• Authors: J. Wong, Y.Y. Sia, N.L. Misso, S. Aggarwal, A. Ng, K.D. Bhoola
• Journal: Pathology Research International
• Published: 2011;2011:167046
• Cited by: 12 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Tissue kallikrein (KLK1) and plasma kallikrein (KLKB1) may regulate the growth and proliferation of tumours of the lung and pleura, through the generation of kinin peptides that signal through the kinin B(1) (BDKRB1) and B(2) (BDKRB2) receptors. The development and progression of cancer results from genetic mutations, as well as epigenetic changes that include methylation of DNA at CpG islands. The aim of this study was to assess whether expression of the kallikrein-kinin genes in lung cancer and mesothelioma cells is regulated by DNA methylation. Quantitative reverse transcriptase-PCR and immunocytochemistry showed differences in the basal expression of the kallikrein-kinin genes and proteins in lung carcinoma and mesothelioma cells, compared with non-malignant lung epithelial and mesothelial cells, respectively. Following treatment with the demethylating agent, 5-azacytidine (5-AZA), KLKB1 mRNA expression was consistently increased in both lung carcinoma and mesothelioma cells, whereas KLK1, BDKRB1 and BDKRB2 mRNA expression was decreased or unchanged. Increased expression of KLKB1 after 5-AZA treatment suggests it may function as a tumour suppressor gene in cancers of the lung and pleura. Studies on DNA methylation of the kallikrein-kinin genes will enhance understanding of their role in carcinogenesis and provide insights into the importance of kallikreins as tumour biomarkers.

In plain English

This was a laboratory study of cells grown in dishes. It did not study people, and it did not measure asbestos exposure. The authors, working in Perth, Australia, asked whether DNA methylation, a chemical tag that can switch genes off, controls a group of genes called the kallikrein-kinin genes in lung cancer cells and malignant pleural mesothelioma cells. They used two lung cancer cell lines, three mesothelioma cell lines grown from the pleural fluid of three patients, and two non-malignant comparison cell lines. The cells were treated for 24 hours with 5-azacytidine, a drug that removes methylation, at 6.25 or 25 micromolar. Before treatment, gene activity in the cancer cells differed from the comparison cells. For example, KLK1 was more than 50-fold higher in NO36 mesothelioma cells than in Met-5A cells. After treatment, the KLKB1 (plasma kallikrein) gene rose to some extent in every cancer cell line except LO68, while the other genes fell or changed variably. The authors concluded that KLKB1 may act as a tumour suppressor gene. They noted that the data came from two to three experiments, so meaningful statistical analysis was not possible.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) cell-culture experiment, with computer mapping of CpG islands in the genes
• Who was studied: Human cell lines: non-malignant bronchial epithelial cells (BEAS-2B), lung adenocarcinoma (H2126), lung squamous cell carcinoma (H520), a transformed mesothelial cell line (Met-5A), and three malignant mesothelioma cell lines (LO68, NO36, JU77) derived from the pleural effusions of three patients with malignant pleural mesothelioma; Perth, Australia
• Exposure: The demethylating drug 5-azacytidine at 6.25 or 25 μM for 24 hours, compared with acetic acid (vehicle) alone. No asbestos exposure was studied; asbestos is mentioned only as background.
• Main results:
  • Before treatment, H2126 lung adenocarcinoma cells had greater mRNA expression than non-malignant BEAS-2B cells for KLK1 (21-fold), BDKRB1 (77-fold), BDKRB2 (300-fold) and KNG1 (44-fold)
  • Before treatment, H520 lung squamous carcinoma cells had much lower KLK1, BDKRB1 and BDKRB2 mRNA than BEAS-2B cells (100-, 590- and 60-fold lower, respectively)
  • Before treatment, KLK1 mRNA was greater in the mesothelioma cell lines than in Met-5A cells: NO36 more than 50-fold, LO68 more than 30-fold, JU77 more than 2-fold
  • Before treatment, KLKB1 mRNA was greater in the mesothelioma cell lines than in Met-5A cells: JU77 more than 45-fold, NO36 more than 20-fold, LO68 10-fold
  • In H520 cells, 25 μM 5-azacytidine produced a more than 40-fold increase in BDKRB2 expression
  • In NO36 and LO68 mesothelioma cells, 25 μM 5-azacytidine decreased KLK1, BDKRB1, BDKRB2 and KNG1 mRNA by 10- to 70-fold
  • In NO36 cells, KLKB1 mRNA increased more than 5-fold with 6.25 μM 5-azacytidine
  • 5-azacytidine increased KLKB1 expression to some extent in all lung cancer and mesothelioma cell lines except LO68
  • At 6.25 and 12.5 μM, 5-azacytidine had no significant effect on the growth of any of the cell types (data not shown)
• The authors' conclusion: The authors concluded that, because 5-azacytidine generally suppressed the kallikrein-kinin genes other than KLKB1, those genes are more likely to function as oncogenes. In contrast, the general increase in KLKB1 suggests it may be a tumour suppressor gene in cancers of the lung and pleura.
• Limitations the authors state:
  • Results were means of only two to three separate experiments, so meaningful statistical analysis was not possible; a 2-fold or greater change was treated as biologically significant
  • The KLKB1, BDKRB1 and KNG1 genes did not appear to have CpG islands in their putative promoter regions, which leaves open how methylation might affect these genes
  • The authors state that further studies using bisulphite sequencing are needed to detect CpG sites that are methylated differently in cancerous and noncancerous cells

Topics

Medical terms in this study, explained in our Lexicon: Carcinoma, Pleura, Lung Cancer, Mesothelioma, Mesothelial Cells, Carcinogenesis, Biomarker.

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