Epigenetic Inactivation of the Tumor Suppressor IRX1 Occurs Frequently in Lung Adenocarcinoma and Its Silencing Is Associated with Impaired Prognosis

Marburg Lung Center (UGMLC) and Translational Lung Research Center (TLRC) Heidelberg, German Center for Lung Research (DZL), Universities of Giessen, 35392 Giessen, Germany

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Department of Surgery, Thoraxklinik at Heidelberg University Hospital, 69126 Heidelberg, Germany

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Department of Pathology, Institute of Pathology, University Hospital Heidelberg, 69120 Heidelberg, Germany

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Department of Lung Development and Remodeling, Max-Planck-Institute for Heart and Lung Research, 61231 Bad Nauheim, Germany

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Institute of Molecular Oncology, Member of the German Center for Lung Research (DZL), Philipps-University, 35032 Marburg, Germany

Author to whom correspondence should be addressed.

Cancers 2020, 12(12), 3528; https://doi.org/10.3390/cancers12123528

Received: 16 October 2020 / Revised: 18 November 2020 / Accepted: 24 November 2020 / Published: 26 November 2020

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Lung cancer is one of the most commonly diagnosed cancers worldwide and the most common cause of cancer-related deaths. During lung carcinogenesis, epigenetic alteration of tumor-related genes is a frequent event and especially silencing of tumor suppressor genes is often found. In our work, we identified Iroquois homeobox 1 (IRX1) from the lung cancer susceptibility locus 5p15.33, as an epigenetically silenced target gene. We report frequent epigenetic inactivation of IRX1 in primary lung adenocarcinoma. Moreover, reduced expression and hypermethylation of IRX1 was correlated with an impaired prognosis of patients with lung adenocarcinoma. Functionally, IRX1 overexpression induced signs of apoptosis including fragmented nuclei and expression of a proapoptotic regulator. Loss of IRX1 expression by its promoter hypermethylation can serve as a diagnostic and prognostic lung cancer biomarker.

Abstract

Iroquois homeobox (IRX) encodes members of homeodomain containing genes which are involved in development and differentiation. Since it has been reported that the IRX1 gene is localized in a lung cancer susceptibility locus, the epigenetic regulation and function of IRX1 was investigated in lung carcinogenesis. We observed frequent hypermethylation of the IRX1 promoter in non-small cell lung cancer (NSCLC) compared to small cell lung cancer (SCLC). Aberrant IRX1 methylation was significantly correlated with reduced IRX1 expression. In normal lung samples, the IRX1 promoter showed lower median DNA methylation levels (<10%) compared to primary adenocarcinoma (ADC, 22%) and squamous cell carcinoma (SQCC, 14%). A significant hypermethylation and downregulation of IRX1 was detected in ADC and SQCC compared to matching normal lung samples (p < 0.0001). Low IRX1 expression was significantly correlated with impaired prognosis of ADC patients (p = 0.001). Reduced survival probability was also associated with higher IRX1 promoter methylation (p = 0.02). Inhibition of DNA methyltransferase (DNMT) activity reactivated IRX1 expression in human lung cancer cell lines. Induced DNMT3A and EZH2 expression was correlated with downregulation of IRX1. On the cellular level, IRX1 exhibits nuclear localization and expression of IRX1 induced fragmented nuclei in cancer cells. Localization of IRX1 and induction of aberrant nuclei were dependent on the presence of the homeobox of IRX1. By data mining, we showed that IRX1 is negatively correlated with oncogenic pathways and IRX1 expression induces the proapoptotic regulator BAX. In conclusion, we report that IRX1 expression is significantly associated with improved survival probability of ADC patients. IRX1 hypermethylation may serve as molecular biomarker for ADC diagnosis and prognosis. Our data suggest that IRX1 acts as an epigenetically regulated tumor suppressor in the pathogenesis of lung cancer. View Full-Text

Keywords: IRX; DNA methylation; epigenetic; lung cancer; adenocarcinoma; apoptosis; tumor suppressor; homeobox

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MDPI and ACS Style

Küster, M.M.; Schneider, M.A.; Richter, A.M.; Richtmann, S.; Winter, H.; Kriegsmann, M.; Pullamsetti, S.S.; Stiewe, T.; Savai, R.; Muley, T.; Dammann, R.H. Epigenetic Inactivation of the Tumor Suppressor IRX1 Occurs Frequently in Lung Adenocarcinoma and Its Silencing Is Associated with Impaired Prognosis. Cancers 2020, 12, 3528. https://doi.org/10.3390/cancers12123528

AMA Style

Küster MM, Schneider MA, Richter AM, Richtmann S, Winter H, Kriegsmann M, Pullamsetti SS, Stiewe T, Savai R, Muley T, Dammann RH. Epigenetic Inactivation of the Tumor Suppressor IRX1 Occurs Frequently in Lung Adenocarcinoma and Its Silencing Is Associated with Impaired Prognosis. Cancers. 2020; 12(12):3528. https://doi.org/10.3390/cancers12123528

Chicago/Turabian Style

Küster, Miriam M., Marc A. Schneider, Antje M. Richter, Sarah Richtmann, Hauke Winter, Mark Kriegsmann, Soni S. Pullamsetti, Thorsten Stiewe, Rajkumar Savai, Thomas Muley, and Reinhard H. Dammann. 2020. "Epigenetic Inactivation of the Tumor Suppressor IRX1 Occurs Frequently in Lung Adenocarcinoma and Its Silencing Is Associated with Impaired Prognosis" Cancers 12, no. 12: 3528. https://doi.org/10.3390/cancers12123528

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Epigenetic Inactivation of the Tumor Suppressor IRX1 Occurs Frequently in Lung Adenocarcinoma and Its Silencing Is Associated with Impaired Prognosis

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