Recurrent TERT promoter mutations identified in a large-scale study of multiple tumour types are associated with increased TERT expression and telomerase activation

doi:10.1016/j.ejca.2015.03.010

European Journal of Cancer

Volume 51, Issue 8, May 2015, Pages 969-976

https://doi.org/10.1016/j.ejca.2015.03.010 Get rights and content

Abstract

Background

Several somatic mutation hotspots were recently identified in the telomerase reverse transcriptase (TERT) promoter region in human cancers. Large scale studies of these mutations in multiple tumour types are limited, in particular in Asian populations. This study aimed to: analyse TERT promoter mutations in multiple tumour types in a large Chinese patient cohort, investigate novel tumour types and assess the functional significance of the mutations.

Methods

TERT promoter mutation status was assessed by Sanger sequencing for 13 different tumour types and 799 tumour tissues from Chinese cancer patients. Thymic epithelial tumours, gastrointestinal leiomyoma, and gastric schwannoma were included, for which the TERT promoter has not been previously sequenced. Functional studies included TERT expression by reverse-transcriptase quantitative polymerase chain reaction (RT-qPCR), telomerase activity by the telomeric repeat amplification protocol (TRAP) assay and promoter activity by the luciferase reporter assay.

Results

TERT promoter mutations were highly frequent in glioblastoma (83.9%), urothelial carcinoma (64.5%), oligodendroglioma (70.0%), medulloblastoma (33.3%) and hepatocellular carcinoma (31.4%). C228T and C250T were the most common mutations. In urothelial carcinoma, several novel rare mutations were identified. TERT promoter mutations were absent in gastrointestinal stromal tumour (GIST), thymic epithelial tumours, gastrointestinal leiomyoma, gastric schwannoma, cholangiocarcinoma, gastric and pancreatic cancer. TERT promoter mutations highly correlated with upregulated TERT mRNA expression and telomerase activity in adult gliomas. These mutations differentially enhanced the transcriptional activity of the TERT core promoter.

Conclusions

TERT promoter mutations are frequent in multiple tumour types and have similar distributions in Chinese cancer patients. The functional significance of these mutations reflect the importance to telomere maintenance and hence tumourigenesis, making them potential therapeutic targets.

Introduction

Telomeres are essential structural elements that seal and protect the ends of chromosomes from recombination and end-to-end fusion. Maintenance of the telomeres requires the ribonucleoprotein complex known as telomerase. Telomerase consists of a multiple proteins, including telomerase reverse transcriptase (TERT) and its integral RNA subunit. In normal somatic cells, telomeres gradually shorten after successive rounds of cell division, resulting in senescence. Most tumour cells overcome this limitation by re-activating telomerase; however, the mechanisms of this reactivation process remain unknown [1].

Recently, two somatic mutations in the promoter region of TERT were reported in melanomas [2], [3]. The two most common mutations occurred at −124 and −146 base pairs upstream of the TERT translation start site (hereafter referred to as C228T and C250T, respectively). These mutations occurred in nearly 70% of melanoma tumours and cell lines. Although the exact mechanism is unclear, each mutation independently generates a novel E-twenty-six (ETS) transcription factor binding site (GGAA/T) and has been shown to increase the transcriptional activity of the TERT promoter [3], [4]. Interestingly, TERT promoter mutations are not restricted to melanoma and occur frequently in several tumour types, including gliomas [4], [5], liposarcomas [4], urothelial carcinomas [4], [6], [7], [8], [9] and hepatocellular carcinomas [4], [10]. We sought to expand this work to search for additional TERT promoter mutations by investigating the TERT promoter mutation status of a large subset of cancers in a Chinese population, as most previous large-scale reports have been limited to Western populations. Furthermore, we assessed the TERT promoter mutation status of a number of tumour types that have not been investigated previously, including thymic epithelial tumours, gastrointestinal leiomyoma and gastric schwannoma. Finally, we assessed the functional consequence and relevance of the most frequent mutations identified in this study.

Section snippets

Tissue samples

A total of 799 tumour tissues from different tumour types were obtained from the archives of the Zhejiang Provincial People’s Hospital, between January 2007 and October 2013. All tissue samples were collected during surgical procedures with patients’ consent. Formalin-fixed, paraffin-embedded (FFPE) tissue samples were fixed in formalin and later embedded in paraffin. The xenografts and snap-frozen primary tumour tissues used for reverse-transcriptase quantitative polymerase chain reaction

Sequencing analysis of the TERT promoter mutations in Chinese cancer patients

We evaluated TERT promoter mutations successfully in 799 tumour specimens and identified 268 samples (33.5%) with TERT promoter mutations. Overall, the hotspot mutations C228T (64.6%) and C250T (23.9%) were the most common and were mutually exclusive. These TERT promoter mutations occurred in many tumour types, including glioblastoma, medulloblastoma, urothelial carcinoma, hepatocellular carcinoma and gallbladder carcinoma. TERT promoter mutations were absent in many types of tumours, including

Discussion

Since the first discovery of the TERT promoter mutations in melanoma [2], [3], many studies have been performed to establish their role as a novel genetic mechanism in human tumourigenesis. Subsequent research has revealed high frequencies of TERT promoter mutations in glioblastoma [4], [5], bladder cancer [6], hepatocellular carcinoma [10] and other human cancers [4]. However, most large-scale reports surveying multiple tumours are based on Western populations, with limited studies of Asian,

Conflict of interest statement

Under agreements between Duke University and Blueprint Medicines H.Y. is entitled to a share of the royalties received by the University on sales of products related to genes described in this manuscript. H.Y. receives royalties from Agios Pharmaceuticals, Sanofi-Aventis and Personal Genome Diagnostics. S.W. and H.Y. are co-founders and own stocks of Beijing Pangenomics Technology, Co. Ltd.

Acknowledgements

This work was supported by the National Natural Science Foundation of China (No. 81372598, No. 81071991, No. 81000016), Zhejiang Provincial Program for the Cultivation of High-level Innovative Health Talents, Major projects of Science and Technology Department of Zhejiang Province (Nos. 2011C13036-1, 2013T301-13). the V Foundation, the Accelerate Brain Cancer Cure Foundation, the Slomo and Cindy Silvian Foundation, the Voices Against Brain Cancer Foundation, the Pediatric Brain Tumor Foundation

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TERT promoter mutation is one of the most common genomic alterations in urothelial carcinoma (UC). Its prognostic role on patients' outcomes is still not clear.
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Despite the clear role of zone 2 hepatocytes in liver regeneration, little is known about their potential contribution to HCC formation. TERT is one of the most frequently mutated genes in HCC development [54]. TERT mutations lead to an overexpression of telomerase, the enzyme responsible for the maintenance of telomere length [20].
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¹: Dong-Sheng Huang, Zhaohui Wang, Xu-Jun He and Bill H. Diplas contributed equally to this work.

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Original ResearchRecurrent TERT promoter mutations identified in a large-scale study of multiple tumour types are associated with increased TERT expression and telomerase activation

Abstract

Background

Methods

Results

Conclusions

Introduction

Section snippets

Tissue samples

Sequencing analysis of the TERT promoter mutations in Chinese cancer patients

Discussion

Conflict of interest statement

Acknowledgements

Clin Neurol Neurosurg

Cancer Cell

Lancet Oncol

Telomere structure and telomerase in health and disease (review)

Int J Oncol

TERT promoter mutations in familial and sporadic melanoma

Science

Highly recurrent TERT promoter mutations in human melanoma

Science

TERT promoter mutations occur frequently in gliomas and a subset of tumors derived from cells with low rates of self-renewal

Proc Natl Acad Sci U S A

Upregulating mutations in the TERT promoter commonly occur in adult malignant gliomas and are strongly associated with total 1p19q loss

Acta Neuropathol

Comprehensive mutation analysis of the TERT promoter in bladder cancer and detection of mutations in voided urine

Eur Urol

TERT promoter mutations occur early in urothelial neoplasia and are biomarkers of early disease and disease recurrence in urine

Cancer Res

TERT promoter mutations in bladder cancer affect patient survival and disease recurrence through modification by a common polymorphism

Proc Natl Acad Sci USA

Frequency of TERT promoter mutations in human cancers

Nat Commun

Original Research
Recurrent TERT promoter mutations identified in a large-scale study of multiple tumour types are associated with increased TERT expression and telomerase activation