DNA methylation of the TERT promoter and its impact on human cancer

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Telomere maintenance is a hallmark of human cancer that enables replicative immortality. Most cancer cells acquire telomere maintenance by telomerase activation through expression of telomerase reverse transcriptase (TERT), a rate-limiting component of the telomerase holoenzyme. Although multiple cancer-specific genetic alterations such as gain of TERT copy number and recurrent TERT promoter mutations (TPM) have been identified, the majority of cancers still express TERT via unknown mechanisms. In the last decade, DNA methylation of the TERT promoter emerged as a putative epigenetic regulatory mechanism of telomerase activation in cancer. Here, we comparatively discuss studies that investigated the DNA methylation landscape of the TERT promoter. We further review the biological and clinical impacts of TERT promoter hypermethylation in cancer and provide insight into future applications of this phenomenon.

Introduction

Replicative immortality is a hallmark of cancer characterized by the ability of cancer cells to bypass senescence and achieve unlimited proliferative capacity [1]. This characteristic is largely dependent on the activation of a telomere maintenance mechanism (although not always; see Chapter X) and allows cancer cells to form macroscopic tumors as well as relapse or recur after initial treatment.

Telomeres are end-chromosomal DNA–protein complexes consisting primarily of (TTAGGG)n tandem DNA repeats protected by a 6-member protein complex known as shelterin [2]. Telomeres protect chromosomal ends from being recognized by the DNA repair machinery, thus preventing chromosomal end-to-end fusions. Progressive shortening of telomeric sequences occurs after each cell division as a result of the end-replication problem, driving normal somatic cells towards senescence once telomeres reach a critically shortened length [3]. In the absence of cell-cycle checkpoints, cells will continue to proliferate without entering senescence until they enter crisis, a chaotic state of chromosomal end-fusions and mitotic catastrophe. While the majority of cells will be irreversibly committed to apoptosis at this stage, a small population will spontaneously emerge and acquire unlimited replicative potential by actively maintaining their telomere lengths [3,4]. This process is known as cellular immortalization, a crucial step in carcinogenesis and cancer progression [5].

In humans, most somatic cells do not display a telomere maintenance mechanism. However, certain cell types including normal stem cells, germ cells and activated memory lymphocytes maintain telomere length through the activity of the telomerase enzymatic complex [6]. Research in the past few decades has revealed that the telomerase holoenzyme is tightly regulated in normal somatic cells, most importantly by repressing its rate-limiting component, telomerase reverse transcriptase (TERT). To overcome this intrinsic defensive mechanism against uncontrolled proliferation and malignant transformation, most cancer cells acquire alterations in order to aberrantly upregulate TERT expression and eventually enable telomere maintenance.

Genetic alterations that result in aberrant upregulation of TERT expression include TERT amplification [7], TERT rearrangements [8,9], and TERT promoter mutations (TPM) [10,11], which have been identified in several human cancer types. TPMs have been associated with upregulation of TERT expression and poorer clinical outcomes in several cancers including brain [12], thyroid [13], bladder [14••], and skin [15]. However, other common cancers (i.e. breast, prostate, lung, and colon) harbor a low frequency of TPM and/or other known genetic alterations, and the mechanism of telomerase activation in these tumors remains largely unknown.

Some of the understudied aspects in TERT transcriptional regulation are the epigenetic alterations at the TERT promoter. Although the interplay between histone modifications and DNA methylation has not yet been fully explored, numerous studies have provided clinical and mechanistic insights on the role of TERT promoter DNA methylation in cancer [16, 17, 18, 19]. In most cases, promoter DNA methylation of human genes results in downregulated expression or complete gene silencing [20]. Interestingly, the human TERT promoter that is highly enriched for CpG dinucleotides and thus is subject to DNA methylation does not behave in such a simplistic manner. In this review, we discuss recent progress in understanding the role of DNA methylation at the TERT promoter as a telomere maintenance activating mechanism in the context of human cancer, from both biological and clinical perspectives. Furthermore, we review how this knowledge can be implicated in potential telomere-based anti-cancer strategies.

Section snippets

Understanding the DNA methylation landscape of the TERT promoter in human cancer

Initial studies using human cell lines to assess DNA methylation as a form of epigenetic regulation at the TERT promoter have generated inconsistent results. One seminal study reported that non-TERT expressing normal cell lines had unmethylated/hypomethylated promoters, while two-thirds of TERT expressing cancer cell lines exhibited partial or total methylation at the promoter [21]. Contradictory to this, other groups have reported TERT expressing cancer cell lines with hypomethylated TERT

Biological role of cancer-specific DNA methylation within the TERT promoter

Although promoter DNA hypermethylation is generally associated with gene silencing [20], a genome-wide study in prostate cancer demonstrated that a large number of promoter-associated CpG islands that were hypermethylated surprisingly showed transcriptional activation [29]. This is consistent with the observation that TERT promoter (THOR) hypermethylation is associated with upregulation of TERT expression in cancers (Figure 1a). Functional analyses using luciferase reporter plasmid vectors have

Clinical investigation of cancer-specific DNA methylation within the TERT promoter

The prevalence of TERT promoter (THOR) hypermethylation was recently examined and reported to be ≥45% in over 1300 human cancer samples across 11 tumor types screened [27••]. This methylation signature exhibited striking specificity, as >90% of the tumors exceeded the median THOR methylation level of normal samples. Interestingly, THOR hypermethylation is more prevalent (>70%) in cancer types that lack TPMs, such as lung, breast, prostate, and colon cancers, compared to cancer types that

Clinical implications of THOR hypermethylation signature

Since the introduction of UTSS methylation status as a potential biomarker in pediatric brain tumors [19], several clinical studies published in the recent years have focused their analyses on this smaller region within THOR (Figure 2). The mapping of THOR, a larger region consisting of 52 CpG sites that display cancer-specific hypermethylation [27••], may lead to the identification of novel cancer-specific DNA methylation signatures that would serve as predictive and prognostic biomarkers in

Conclusion and future directions

In this review, we highlight the emerging role of epigenetic regulation through DNA methylation of the TERT promoter as a telomere maintenance activating mechanism in cancer. This phenomenon is an attractive target – both biologically and clinically – as it is commonly observed across most malignant human cancer types. Comprehensive understanding of the timing and mechanisms that govern cancer-specific DNA methylation of the TERT promoter during malignant transformation will be crucial in

Conflicts of interest statement

Nothing declared.

References and recommended reading

Papers of particular interest, published within the period of review, have been highlighted as:

  • • of special interest

  • •• of outstanding interest

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