Telomerase Reverse Transcriptase mRNA Expression Research Articles

The TERT promoter SNP rs2853669 decreases E2F1 transcription factor binding and increases mortality and recurrence risks in liver cancer.

A common single-nucleotide polymorphism in the telomerase reverse transcriptase (TERT) promoter, rs2853669 influences patient survival rates and the risk of developing cancer. Recently, several lines of evidence suggest that the rs2853669 suppresses TERT promoter mutation-mediated TERT expression levels and cancer mortality as well as recurrence rates. However, no reports are available on the impact of rs2853669 on TERT expression in hepatocellular carcinoma (HCC) and its association with patient survival. Here, we found that HCC-related overall and recurrence-free survival rates were not associated with TERT promoter mutation individually, but rs2853669 and the TERT promoter mutation in combination were associated with poor survival rates. TERT mRNA expression and telomere fluorescence levels were greater in patients with HCC who had both the combination. The combination caused TERT promoter methylation through regulating the binding of DNA methyltransferase 1 and histone deacetylase 1 to the TERT promoter in HCC cell lines. The TERT expression level was significantly higher in HCC tumor with a methylated promoter than in that with an unmethylated promoter. In conclusion, we demonstrate a substantial role for the rs2853669 in HCC with TERT promoter mutation, which suggests that the combination of the rs2853669 and the mutation indicate poor prognoses in liver cancer.

Thymocytes maintain immune activity through telomere elongation in rats under hypoxic conditions.

The main purpose of the present study was to investigate the change in thymocyte telomere length of rats exposed to different hypoxic conditions for different periods of time, as well as its effect on the immune system. A total of 110 male Wistar rats were randomly assigned to one of the three following groups: i) Sea level (SL) group, in which 10 rats were maintained at an altitude of 10 m; ii) moderate altitude (MA) group, in which 50 rats were maintained at an altitude of 2,260 m and then randomly sacrificed on days 1, 3, 7, 15 and 30 (n=10 each); and iii) simulated high altitude (SHA) group, in which 50 rats were maintained at a simulated altitude of 5,000 m, and then randomly sacrificed on days 1, 3, 7, 15 and 30 (n=10 each). The morphological changes of the thymus were observed, while the telomere length, the mRNA and protein expression levels of telomerase reverse transcriptase (TERT), and the peripheral blood lymphocyte count were measured. The results indicated that hypoxia induced morphological changes and apoptosis in thymocytes, as well as atrophy of the thymus tissue, and resulted in a significant increase in telomere length and TERT mRNA and protein expression levels. This effect appeared to be more pronounced in the SHA group compared with that in the MA group; however, no statistically significant changes were observed in the peripheral blood lymphocyte count. Based on these findings, the hypoxia-associated loss of thymic function appears to be only quantitative and not qualitative, and the thymus may be able to maintain its immune function even under hypoxic conditions.

Anti-aging effect of estrogen on telomerase activity in ovariectomised rats – animal model for menopause

Objective: The aim of this study was to investigate the anti-aging effects of exogenous estrogen on telomerase activity in ovariectomized female Sprague-Dawley rats.Methods: Thirty-three 12-week-old female rats were divided into three groups: the ovariectomized-Treated group (Treated, n = 11), the ovariectomized control group (OVX, n = 11) and the Sham-operated group (Sham, n = 11). The rats in the Treated group were given 0.21 mg/kg estradiol valerate intragastric administration while other two groups were given the amount of physiological saline daily. All of the animals were euthanized 12 weeks after treatment, and abdominal aortic blood samples were taken to assess the level of estradiol (E2), follicle stimulating hormone (FSH). Telomerase activity and telomerase reverse transcriptase (TERT) mRNA expression in the heart, liver, brain tissues of all rats were measured by reverse transcriptional polymerase chain reaction (RT-PCR) and competitive enzyme-linked immunosorbent assay (ELISA).Results: Compared to the OVX and Sham group, telomerase activity and TERT mRNA levels were significantly increased in the heart, liver and brain tissues of rats in the Treated group (p < 0.05). The telomerase expression was significantly higher in the heart than in liver and brain tissues in the Treated group (p < 0.05).Conclusion: Based on the findings in the ovariectomized female rats model, exogenous estrogen can significantly up-regulate telomerase activity and TERT mRNA expression to exert the effects of anti-aging.

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

BackgroundSeveral 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. MethodsTERT 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. ResultsTERT 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. ConclusionsTERT 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.

1037 Serum Human Telomerase Reverse Transcriptase Messenger RNA As a Novel Marker for Pancreatic Ductal Adenocarcinoma

1037 Serum Human Telomerase Reverse Transcriptase Messenger RNA As a Novel Marker for Pancreatic Ductal Adenocarcinoma

Telomerase redux: ready for prime time?

By maintaining genome integrity, controlling cell proliferation, and regulating tissue homeostasis, telomerase plays a critical role in the pathology of aging and cancer. Telomerase is composed of telomerase RNA, or telomerase RNA component (TERC), which serves as a template for telomeric DNA synthesis, and a catalytic subunit, telomerase reverse transcriptase (TERT). The canonical function of TERT is the synthesis of telomeric DNA repeats and the maintenance of telomere length. Recent studies suggest that adeno-associated virus (AAV)-expressed TERT in adult mice can increase life span and health span. Because TERT physically interacts with proteins that regulate gene expression, and because TERT's expression at high levels is associated with oncogenic transformation, its synthesis needs to be carefully regulated. Due to safety concerns, transient expression of TERT mRNA may be preferred for tissue engineering and adoptive stem cell therapy.

Prognostic quality of activating TERT promoter mutations in glioblastoma: interaction with the rs2853669 polymorphism and patient age at diagnosis.

Expression of the telomerase reverse transcriptase (TERT) might be altered by activating mutations of the rs2853669 polymorphism within the promoter region. Here we investigate the impact of these genomic alterations on telomerase activation and dissect their prognostic potential in glioblastoma (GBM). The respective TERT promoter region was sequenced in 126 GBM tissues and compared with clinical parameters and glioma biomarkers MGMT promoter methylation and IDH1 mutation. TERT mRNA expression, telomerase activity, and telomere lengths were determined by reverse transcriptase PCR, TRAP assay, and real-time PCR, respectively. Seventy-three percent of GBM patients harbored TERT promoter mutations associated with enhanced telomerase activity and TERT mRNA expression but reduced telomere lengths (P < .001 for all). Patients with mutated tumors exhibited significantly shorter overall survival in the entire cohort (11.5 vs 23.1 months; P < .0001) and in the primary GBM patient subgroup lacking IDH1 mutations (n = 120; P = .0084). This prognostic impact was confined to younger patients (aged <65 years), while the negative prognostic power of enhanced age at diagnosis was limited to those patients lacking TERT promoter mutations. Presence of the common single nucleotide polymorphism rs2853669, disrupting an endogenous Ets2 transcription factor-binding site, was associated with improved survival exclusively in patients with a wild-type TERT promoter. On the contrary, the shortest mean overall survival was detected in those patients harboring both an activating TERT promoter mutation and homozygous rs2853669 alleles. In summary, TERT promoter mutations are powerful prognosticators for worse course of disease in human GBM patients but their prognostic value is influenced by the rs2853669 polymorphism and age at diagnosis.

Cancer. TERT promoter mutations and telomerase reactivation in urothelial cancer.

Reactivation of telomerase, the chromosome end-replicating enzyme, drives human cell immortality and cancer. Point mutations in the telomerase reverse transcriptase (TERT) gene promoter occur at high frequency in multiple cancers, including urothelial cancer (UC), but their effect on telomerase function has been unclear. In a study of 23 human UC cell lines, we show that these promoter mutations correlate with higher levels of TERT messenger RNA (mRNA), TERT protein, telomerase enzymatic activity, and telomere length. Although previous studies found no relation between TERT promoter mutations and UC patient outcome, we find that elevated TERT mRNA expression strongly correlates with reduced disease-specific survival in two independent UC patient cohorts (n = 35; n = 87). These results suggest that high telomerase activity may be a better marker of aggressive UC tumors than TERT promoter mutations alone.

Down regulation of miR-202 modulates Mxd1 and Sin3A repressor complexes to induce apoptosis of pancreatic cancer cells

Aberrant regulation of microRNA expression in pancreatic cancers has been shown to play an important role in its inherent poor prognosis and malignant potential. MicroRNAs have also been shown to inhibit translation of genes by targeting the 3′-untranslated region (3-UTR) of mRNAs resulting in the inhibition of translation and often destruction of the mRNA. In the present study we investigated the role of the microRNA miR-202 in the apoptotic pathways of pancreatic cancer cells. The adamantyl-related molecule, 3-Cl-AHPC down-regulated expression of miR-202 and miR-578 resulting in the increased expression of mRNA and protein expression of their target genes, Max dimerization protein 1 (Mxd1/Mad1) and the Sin3A associated protein 18 (SAP18). Overexpression of pre-miR-202 led to diminished levels of Mxd1 and blocked the 3-Cl-AHPC-mediated increase in Mxd1 mRNA expression. The addition of the microRNA inhibitor 2′-O-methylated miR-202 enhanced the 3-Cl-AHPC-mediated increase of Mxd1 mRNA levels as well as 3-CI-AHPC-mediated apoptosis. We found increased Mxd1 bound to the Sin3A repressor protein complex through its increased binding with HDAC-2 and subsequently enhanced transcriptional repression in cells as evidenced by increased HDAC activity. Mxd1 also repressed human telomerase reverse transcriptase (hTERT) mRNA expression through its increased binding to the hTERT promoter site and resulted in decreased telomerase activity in cells. Our results demonstrate that down regulation of miR-202 increased the expression of its target Mxd1, followed by Mxd1 recruitment to the Sin3A repressor complex and through its dimerization with Max, and increased repression of Myc-Max target proteins.

Influence of the hTERT rs2736100 polymorphism on telomere length in gastric cancer.

To investigate the functional consequences of rs2736100 polymorphism in telomere length and examine its link to gastric cancer risk. Telomere length and human telomerase reverse transcriptase (hTERT) mRNA expression were measured in 35 gastric cancer tissues and 5 cell lines and correlated to rs2736100 polymorphism. The relationship between rs2736100 polymorphism and the risk of gastric cancer were examined in 243 gastric cancer patients and 246 healthy individuals. The rs2736100 A allele carrier is closely associated with reduced hTERT mRNA expression and shortened telomere length in gastric cancer tissue and cell lines. When gastric cancers were stratified by histological subtype, telomere length and hTERT mRNA levels were significantly increased in those with the C/C genotype in intestinal-type gastric cancer, but not in diffuse-type gastric cancer. Interestingly, there was no significant difference in the genotype and allele frequencies of the rs2736100 polymorphism between the patients with gastric cancer and healthy controls. The rs2736100 polymorphism of the hTERT gene is involved in the regulation of hTERT expression and telomere length, but not in the risk of gastric cancer.

Vitamin D suppresses leptin stimulation of cancer growth through microRNA.

Obesity is a pandemic and major risk factor for cancers. The reduction of obesity would have been an effective strategy for cancer prevention, but the reality is that worldwide obesity has kept increasing for decades, remaining a major avoidable cancer risk secondary only to smoke. The present studies suggest that vitamin D may be an effective agent to reduce obesity-associated cancer risks in women. Molecular analyses showed that leptin increased human telomerase reverse transcriptase (hTERT) mRNA expression and cell growth through estrogen receptor-α (ERα) activation in ovarian cancer cells, which was suppressed by 1α,25-dihydroxyvitamin D3 [1,25(OH)2D3]. The suppression was compromised when miR-498 induction by the hormone was depleted with microRNA (miRNA) sponges. In mice, high-fat diet (HFD) stimulation of ovarian tumor growth was remarkably suppressed by 1,25(OH)2D3 analogue EB1089, which was also compromised by miR-498 sponges. EB1089 did not alter HFD-induced increase in serum leptin levels but increased miR-498 and decreased the diet-induced hTERT expression in tumors. Quantitative RT-PCR analyses revealed an inverse correlation between hTERT mRNA and miR-498 in response to 1,25(OH)2D3 in estrogen-sensitive ovarian, endometrial, and breast cancers. The studies suggest that miR-498-mediated hTERT downregulation is a key event mediating the anti-leptin activity of 1,25(OH)2D3 in estrogen-sensitive tumors in women.

Effects of epigallocatechin gallate on the proliferation and apoptosis of the nasopharyngeal carcinoma cell line CNE2.

The present study explored the effects of epigallocatechin gallate (EGCG) on the cell cycle, proliferation and apoptosis of the nasopharyngeal carcinoma cell line CNE2 in vitro. The proliferation of CNE2 cells was detected using the cell counting kit-8 method. Cell cycle distribution and apoptosis were detected using flow cytometry. The human telomerase reverse transcriptase (hTERT) mRNA expression was determined using reverse transcription polymerase chain reactions. The protein expression of hTERT and Myc proto-oncogene protein (c-Myc) was observed using western blot analysis. EGCG inhibited the proliferation of CNE2 cells in a concentration-dependent manner (P<0.05) and blocked the cell cycle progression of the cells. In the low concentration (100 μg/ml) group, the cell cycle arrest showed a time-dependent manner. However, as the concentration increased and action time was prolonged, this time dependency became less marked. EGCG promoted the apoptosis of CNE2 cells in a time-dependent manner. In addition, EGCG downregulated the mRNA and protein expression of hTERT and downregulated the expression of c-Myc protein. Downregulation of the expression of hTERT and c-Myc was more evident in the high-dose group (200 μg/mL). In conclusion, EGCG has proliferation-inhibiting, cell cycle-blocking and apoptosis-promoting effects on CNE2 cells. EGCG may be developed into an auxiliary therapeutic agent for the treatment of nasopharyngeal carcinoma.

TERT promoter mutations in gliomas, genetic associations and clinico-pathological correlations.

Background:The role of telomerase reverse transcriptase (TERT) in gliomagenesis has been recently further strengthened by the frequent occurrence of TERT promoter mutations (TERTp-mut) in gliomas and evidence that the TERT SNP genetic rs2736100 influences glioma risk. TERTp-mut creates a binding site for Ets/TCF transcription factors, whereas the common rs2853669 polymorphism disrupts another Ets/TCF site on TERT promoter.Methods:We sequenced for TERTp-mut in 807 glioma DNAs and in 235 blood DNAs and analysed TERT expression by RT-PCR in 151 samples. TERTp-mut status and TERTp polymorphism rs2853669 were correlated with histology, genomic profile, TERT mRNA expression, clinical outcome and rs2736100 genotype.Results:TERTp-mut identified in 60.8% of gliomas (491 out of 807) was globally associated with poorer outcome (Hazard ratio (HR)=1.50). We defined, based on TERTp-mut and IDH mutation status, four prognostic groups: (1) TERTp-mut and IDH-mut associated with 1p19q codeletion, overall survival (OS)>17 years; (2) TERTp-wt and IDH-mut, associated with TP53 mutation, OS=97.5 months; (3) TERTp-wt and IDH-wt, with no specific association, OS=31.6 months; (4) TERTp-mut and IDH-wt, associated with EGFR amplification, OS=15.4 months. TERTp-mut was associated with higher TERT mRNA expression, whereas the rs2853669 variant was associated with lower TERT mRNA expression. The mutation of CIC (a repressor of ETV1-5 belonging to the Ets/TCF family) was also associated with TERT mRNA upregulation.Conclusions:In addition to IDH mutation status, defining the TERTp-mut status of glial tumours should afford enhanced prognostic stratification of patients with glioma. We also show that TERTp-mut, rs2853669 variant and CIC mutation influence Tert expression. This effect could be mediated by Ets/TCF transcription factors.

Silencing of TERT decreases levels of miR-1, miR-21, miR-29a and miR-208a in cardiomyocytes.

To test the hypothesis that telomerase reverse transcriptase (TERT) as an RNA-dependent RNA polymerase could be involved in the amplification of microRNA (miRNA), we have determined the levels of immature and mature miRNA in cultured neonatal rat cardiomyocytes, during the silencing of TERT by siRNA. The silencing of the TERT gene led to the reduction of both telomerase activity and the TERT mRNA expression when compared with scrambled RNA. TERT gene silencing resulted in the decrement of three studied mature miRNAs levels: miRNA-21, miRNA-29a and miRNA-208a when compared with scrambled RNA; but miRNA-1, it was not changed significantly. At the same time, levels of immature miRNA-1 and miRNA-208a were not changed, although the levels of immature miRNA-29a and pri-miRNA-1 were decreased. The data obtained allow us to permit that TERT is a genome-independent source of mature miRNA, and the changes in telomerase activity can significantly influence the level of miRNA in cardiomyocytes.

TERT Promoter Mutations Lead to High Transcriptional Activity under Hypoxia and Temozolomide Treatment and Predict Poor Prognosis in Gliomas

ObjectiveThis study explored the effects of telomerase reverse transcriptase (TERT) promoter mutations on transcriptional activity of the TERT gene under hypoxic and temozolomide (TMZ) treatment conditions, and investigated the status and prognostic value of these mutations in gliomas.MethodsThe effect of TERT promoter mutations on the transcriptional activity of the TERT gene under hypoxic and TMZ treatment conditions was investigated in glioma cells using the luciferase assay. TERT promoter mutations were detected in 101 glioma samples (grades I–IV) and 49 other brain tumors by sequencing. TERT mRNA expression in gliomas was examined by real-time PCR. Hazard ratios from survival analysis of glioma patients were determined relative to the presence of TERT promoter mutations.ResultsMutations in the TERT promoter enhanced gene transcription even under hypoxic and TMZ treatment conditions, inducing upregulation of TERT mRNA expression. Mutations were detected in gliomas, but not in meningiomas, pituitary adenomas, cavernomas, intracranial metastases, normal brain tissues, or peripheral blood of glioma patients. Patients with TERT promoter mutations had lower survival rates, even after adjusting for other known or potential risk factors, and the incidence of mutation was correlated with patient age.Conclusion TERT promoter mutations were specific to gliomas. TERT promoter mutations maintained its ability of inducing high transcriptional activity even under hypoxic and TMZ treatment conditions, and the presence of mutations was associated with poor prognosis in glioma patients. These findings demonstrate that TERT promoter mutations are novel prognostic markers for gliomas that can inform prospective therapeutic strategies.

Transforming growth factor-β1 regulates the telomerase reverse transcriptase in rat hepatic stellate cells.

To determine the effect of transforming growth factor-β1 (TGF-β1) on the expression of telomerase in hepatic stellate cells (HSCs) in rats and the role of TGF-β1 in the development of liver fibrosis. Primary HSCs were isolated from normal rats by density gradient separation and divided into 2 groups for culturing. The morphology of HSCs was identified by the inverted fluorescence microscope. The purity of HSCs was identified by immunohistological expression and fluorescence analysis. One group of HSCs was treated with different concentrations (0, 0.1, 1, and 10 ng/mL) of TGF-β1 for 24 h, while the other group was treated with 1 ng/mL TGF-β1 and cultured for 3, 6, and 9 days. The mRNA expression of telomerase reverse transcriptase (TERT) was assessed and compared by polymerase chain reaction. Cell morphology showed that TGF-β1 triggered the differentiation of HSCs from a quiescent phenotype into highly activated myofibroblasts. TERT mRNA expression in the primary HSCs showed slight increase with the culture time, though with no statistical difference between the results at various time points (P>0.05). TGF-β1 at 0.1 ng/mL did not significantly affect the TERT mRNA level compared with the 0 ng/mL group, while 1 ng/mL and 10 ng/mL TGF-β1 significantly decreased the level of TERT mRNA (P<0.05). TGF-β1 at 1 ng/mL had only weak effect on TERT mRNA expression after the 3 day treatment compared with the 0 ng/mL group (P>0.05). TGF-β1 at 1 ng/mL significantly inhibited TERT mRNA expression 6 days after the treatment (P<0.05). TGF-β1 inhibited the expression of TERT mRNA level in the HSCs in both dose- and time-dependent manner. TGF-β1 may contribute to the transdifferentiation of HSCs by reducing TERT levels to develop hepatic fibrosis.

Expression of human telomerase reverse transcriptase mRNA in the peripheral blood of gallbladder carcinoma patients

Objective To study the human telomerase reverse transcriptase (hTERT) mRNA in peripheral blood of patients with gallbladder carcinoma and its clinical significance.Methods Fifty cases of gallbladder cancer (GBC group),50 cases of adenomatous polyps (AMP group) and 50 normal volunteers (control group) were selected.The expression of hTERT mRNA in peripheral blood was detected by using real-time fluorescent quantitative polymerase chain reaction (FQ-PCR).Results The positive rate of hTERT mRNA in GBC group (66％) was significantly higher than in AMP group (14％) (P ＜ 0.05),and that in AMP group was higher than in control group (P ＜ 0.05).There was significant difference in the hTERT mRNA expression between T3 + T4 and T1 + T2 or between negative and positive lymph nodes (P ＜ 0.05).Conclusion The hTERT mRNA expression in peripheral blood of GBC was significantly increased,and associated with depth of invasion and lymph node metastasis. Key words: Gallbladder; Human telomerase reverse transcriptase; Polymerase chain reaction

Frequent somatic mutations of the telomerase reverse transcriptase promoter in ovarian clear cell carcinoma but not in other major types of gynaecological malignancy

Up-regulated expression of telomerase reverse transcriptase (TERT) and subsequent maintenance of telomere length are essential in tumour development. Recent studies have implicated somatic gain-of-function mutations at the TERT promoter as one of the mechanisms that promote transcriptional activation of TERT; however, it remains unclear whether this genetic abnormality is prevalent in gynaecological neoplasms. We performed mutational analysis in a total of 525 gynaecological cancers, and correlated TERT promoter mutations with clinicopathological features. With the exception of ovarian clear cell carcinomas, in which mutations were found in 37 (15.9%) of 233 cases, the majority of gynaecological malignancies were wild-type. TERT promoter mutation does not appear to be an early event during oncogenesis, as it was not detected in the contiguous endometriosis associated with ovarian clear cell carcinoma. Ovarian clear cell carcinoma cell lines with TERT promoter mutations exhibited higher TERT mRNA expression than those with wild-type sequences (p = 0.0238). TERT promoter mutation tended to be mutually exclusive with loss of ARID1A protein expression (p = 4.4 × 10(-9) ) and PIK3CA mutation (p = 0.0019) in ovarian clear cell carcinomas. No associations with disease-specific survival were observed for ovarian clear cell carcinoma. The above results, in conjunction with our previous report showing longer telomeres in ovarian clear cell carcinomas relative to other types of ovarian cancer, suggests that aberrations in telomere biology may play an important role in the pathogenesis of ovarian clear cell carcinoma.

Telomerase Reverse Transcriptase Promoter Mutations in Bladder Cancer: High Frequency Across Stages, Detection in Urine, and Lack of Association with Outcome

BackgroundHotspot mutations in the promoter of the gene coding for telomerase reverse transcriptase (TERT) have been described and proposed to activate gene expression. ObjectivesTo investigate TERT mutation frequency, spectrum, association with expression and clinical outcome, and potential for detection of recurrences in urine in patients with urothelial bladder cancer (UBC). Design, setting, and participantsA set of 111 UBCs of different stages was used to assess TERT promoter mutations by Sanger sequencing and TERT messenger RNA (mRNA) expression by reverse transcription-quantitative polymerase chain reaction. The two most frequent mutations were investigated, using a SNaPshot assay, in an independent set of 184 non–muscle-invasive and 173 muscle-invasive UBC (median follow-up: 53 mo and 21 mo, respectively). Voided urine from patients with suspicion of incident UBC (n=174), or under surveillance after diagnosis of non–muscle-invasive UBC (n=194), was tested using a SNaPshot assay. Outcome measurements and statistical analysisAssociation of mutation status with age, sex, tobacco, stage, grade, fibroblast growth factor receptor 3 (FGFR3) mutation, progression-free survival, disease-specific survival, and overall survival. Results and limitationsIn the two series, 78 of 111 (70%) and 283 of 357 (79%) tumors harbored TERT mutations, C228T being the most frequent substitution (83% for both series). TERT mutations were not associated with clinical or pathologic parameters, but were more frequent among FGFR3 mutant tumors (p=0.0002). There was no association between TERT mutations and mRNA expression (p=0.3). Mutations were not associated with clinical outcome. In urine, TERT mutations had 90% specificity in subjects with hematuria but no bladder tumor, and 73% in recurrence-free UBC patients. The sensitivity was 62% in incident and 42% in recurrent UBC. A limitation of the study is its retrospective nature. ConclusionsSomatic TERT promoter mutations are an early, highly prevalent genetic event in UBC and are not associated with TERT mRNA levels or disease outcomes. A SNaPshot assay in urine may help to detect UBC recurrences.

Overexpression and promoter mutation of the TERT gene in malignant pleural mesothelioma

Malignant pleural mesothelioma (MPM) is a very aggressive tumor with no known curative treatment. Better knowledge of the molecular mechanisms of mesothelial carcinogenesis is required to develop new therapeutic strategies. MPM, like all cancer cells, needs to maintain telomere length to prevent senescence. Previous studies suggested that the telomere lengthening mechanism in MPM is based mainly on telomerase activity. For this reason, we focused on the key catalytic enzyme, TERT (telomerase reverse transcriptase), by analyzing its gene expression in MPM and by studying the mechanism underlying its upregulation. We used our large collection of MPM composed of 61 MPM in culture and 71 frozen MPM tumor samples. Evaluation of TERT mRNA expression by quantitative RT-PCR showed overexpression in MPM in culture compared with normal mesothelial cells, and in MPM tumor samples compared with normal pleura. We identified a 'hot spot' of mutations in the TERT gene core promoter in both MPM in culture and in MPM tumor samples with an overall frequency of 15%. Furthermore, data clearly identified mutation in the TERT promoter as a mechanism of TERT mRNA upregulation in MPM. In contrast, gene copy number amplification was not associated with TERT overexpression. Then, we analyzed the clinicopathological, etiological and genetic characteristics of MPM with mutations in the TERT promoter. TERT promoter mutations were more frequent in MPM with sarcomatoid histologic subtype (P<0.01), and they were frequently associated with CDKN2A gene inactivation (P=0.03). In conclusion, a subgroup of MPM presents TERT promoter mutations, which lead to TERT mRNA upregulation. This is the first recurrent gain-of-function oncogenic mutations identified in MPM.