ZIC1 is silenced and has tumor suppressor function in malignant pleural mesothelioma

Our plain-English summary of ZIC1 is silenced and has tumor suppressor function in malignant pleural mesothelioma, by Y.Y. Cheng, M.B. Kirschner, N.C. Cheng, et al (Journal of Thoracic Oncology, 2013).

The study

• Authors: Y.Y. Cheng, M.B. Kirschner, N.C. Cheng, S. Gattani, S. Klebe, J.J. Edelman, M.P. Vallely, B.C. McCaughan, H.C. Jin, N. van Zandwijk, G. Reid
• Journal: Journal of Thoracic Oncology
• Published: 2013;8(10):1317-28
• Cited by: 29 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Introduction: Epigenetic inactivation of tumor suppressor genes is involved in the development of malignant pleural mesothelioma (MPM). ZIC1, a potential tumor suppressor gene involved in regulating cell growth and apoptosis, was investigated in MPM cell lines and tumors.

Methods: ZIC1 expression and promoter methylation were evaluated in MPM cell lines and tumor samples by quantitative polymerase chain reaction (PCR), Combined Bisulfite Restriction Analysis, and methylation-specific PCR. ZIC1 was reexpressed in cell lines and functional effects were assessed. miRNA expression was quantified by microarray and reverse transcription quantitative PCR. ZIC1 knockdown and miRNA inhibitors were used to study the relationship between ZIC1 and miRNA expression and confirmed by chromatin immunoprecipitation PCR.

Results: ZIC1 expression was low in MPM cells, and was correlated with ZIC1 promoter methylation and reversed upon decitabine treatment. ZIC1 reexpression inhibited proliferation and invasion in MPM cells whereas knockdown enhanced the growth of MeT-5A. In MPM tumor samples ZIC1 expression was either low or undetectable, with promoter methylation observed in 16 of 24 cases. The overexpression of miR-23a and miR-27a was reduced by ZIC1 reexpression, with inhibitors of miR-23a or miR-27a reducing colony formation. miR-23a overexpression was also associated with shorter survival of MPM patients.

Conclusion: ZIC1 is down-regulated in MPM through promoter methylation and acts as a tumor suppressor through down-regulation of its direct targets miR-23a and miR-27a.

In plain English

This laboratory study examined a gene called ZIC1 in malignant pleural mesothelioma (MPM). The authors used mesothelioma cell lines grown in the laboratory, normal mesothelial cells, and archived tumor samples from MPM patients who had extrapleural pneumonectomy surgery in Sydney, Australia, between 1994 and 2009. The study did not measure asbestos exposure. ZIC1 expression was low or undetectable in most mesothelioma cell lines. In those lines, the gene's control region carried chemical marks (methylation) that can switch genes off, and a demethylating drug, decitabine, increased ZIC1 expression. In tumor samples from 24 patients, ZIC1 was very low or undetectable, with methylation found in 16 of 24. Restoring ZIC1 in mesothelioma cells reduced their growth, invasion and colony formation. It also lowered two microRNAs, miR-23a and miR-27a. Among patients with the epithelioid subtype, miR-23a was 2-fold higher in the 8 shorter survivors than in the 8 longer survivors (p = 0.018). For miR-27a, the increase (1.4-fold, p = 0.114) did not meet the study's p less than 0.05 threshold for statistical significance. The authors concluded that ZIC1 acts as a tumor suppressor gene in MPM.

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 study using mesothelioma cell lines and archived tumor samples (gene expression, methylation and cell-function experiments)
• Who was studied: Five MPM cell lines from ATCC (H2052, H2452, H28, H226, MSTO), the immortalized mesothelial line MeT-5A, the primary lines MM05 and Ren, and normal human mesothelial cells from pericardial fluid. Also archived tumor blocks from 24 MPM patients who underwent extrapleural pneumonectomy at Royal Prince Alfred Hospital or Strathfield Private Hospital, Sydney, between 1994 and 2009.
• Exposure: Asbestos exposure was not studied or measured. The experimental treatments were the demethylating drug decitabine (2 µM for 5 days) and reexpression or knockdown of ZIC1 in cells.
• Main results:
  • In cell lines, ZIC1 was not detected in three of five ATCC MPM lines and was low in the two primary lines (MM05 and Ren). MeT-5A cells showed high levels.
  • No ZIC1 methylation was found in normal mesothelial cells; hypermethylation was found in all MPM lines. Decitabine increased ZIC1 expression in all cell lines.
  • In tumor samples, ZIC1 promoter methylation was found in 16 of 24 MPM samples (67%).
  • Restoring ZIC1 in MPM cell lines inhibited proliferation. Invasion was impaired in H2052, H28, REN and MM05 cells, and these cells also formed fewer colonies.
  • Reducing ZIC1 in MeT-5A cells increased colony formation and growth in soft agar.
  • ZIC1 overexpression in H28 cells reduced miR-23a, miR-27a, miR-30a and miR-30b. ChIP-PCR showed ZIC1 binding directly to the miR-23a/27a promoter. Inhibitors of miR-23a or miR-27a reduced colony formation in MSTO and H28 cells.
  • The survival analysis was limited to epithelioid tumors, with 8 patients per group. Patients with shorter survival (under 15 months) had a 2-fold higher miR-23a level (p = 0.018) than patients with longer survival (over 30 months).
  • For miR-27a, shorter survivors showed a 1.4-fold increase (p = 0.114). This is above the study's p less than 0.05 significance threshold.
• The authors' conclusion: The authors concluded that ZIC1 acts as a tumor suppressor gene in MPM, silenced through promoter hypermethylation. They concluded that it controls miR-23a and miR-27a, which were higher in patients with shorter survival, and suggested this as a potential avenue for therapy.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Lung cancer, Mesothelioma.

Medical terms in this study, explained in our Lexicon: Pleural Mesothelioma, Tumor Suppressor Gene, Apoptosis.

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