Limitless self-renewal is one of the hallmarks of cancer and is attained by telomere maintenance, essentially through telomerase (hTERT) activation. Transcriptional regulation of hTERT is believed to play a major role in telomerase activation in human cancers. The dominant interest in telomerase results from its role in cancer. The role of telomeres and telomere maintenance mechanisms is well established as a major driving force in generating chromosomal and genomic instability. Cancer cells have acquired the ability to overcome their fate of senescence via telomere length maintenance mechanisms, mainly by telomerase activation. hTERT expression is up-regulated in tumors via multiple genetic and epigenetic mechanisms including hTERT amplifications, hTERT structural variants, hTERT promoter mutations and epigenetic modifications through hTERT promoter methylation. Genetic (hTERT promoter mutations) and epigenetic (hTERT promoter methylation and miRNAs) events were shown to have clinical implications in cancers that depend on hTERT activation. Knowing that telomeres are crucial for cellular self-renewal, the mechanisms responsible for telomere maintenance have a crucial role in cancer diseases and might be important oncological biomarkers. Thus, rather than quantifying TERT expression and its correlation with telomerase activation, the discovery and the assessment of the mechanisms responsible for TERT upregulation offers important information that may be used for diagnosis, prognosis, and treatment monitoring in oncology. Furthermore, a better understanding of these mechanisms may promote their translation into effective targeted cancer therapies. Herein, we reviewed the underlying mechanisms of hTERT regulation, their role in oncogenesis, and the potential clinical applications in telomerase-dependent cancers.

Replicative immortality is a hallmark of cancer governed by telomere maintenance. About 90% of human cancers maintain their telomeres by activating telomerase, driven by transcriptional upregulation of telomerase reverse transcriptase (TERT). Although TERT promoter mutations (TPMs) are a major cancer-associated genetic mechanism of TERT upregulation, many cancers exhibit TERT upregulation without TPMs. In this study, we described TERT Hypermethylated Oncological Region (THOR), a 433-bp genomic region encompassing 52 CpG sites located immediately upstream of the TERT core promoter, as a cancer-associated epigenetic mechanism of TERT upregulation. Unmethylated THOR repressed TERT promoter activity regardless of TPMs status, and hypermethylation of THOR counteracted this repressive function. THOR methylation analysis in 1,352 human tumors revealed frequent (>45%) cancer-associated DNA hypermethylation in 9 of 11 (82%) tumor types screened. Additionally, THOR hypermethylation — either independently or along with TPMs — accounted for how approximately 90% of human cancers can aberrantly activate telomerase. Thus, we propose THOR hypermethylation as a prevalent telomerase activating mechanism in cancer that can act independently or in conjunction with TPMs, further supporting the utility of THOR hypermethylation as a prognostic biomarker.

Retroperitoneal lymph node dissection is recommended for residual masses greater than 1 cm after chemotherapy of nonseminomatous germ cell tumors. Currently there is no reliable predictor of post-chemotherapy retroperitoneal lymph node dissection histology. Up to 50% of patients harbor necrosis/fibrosis only so that a potentially morbid surgery has limited therapeutic value. In this study we evaluated the ability of defined serum miRNAs to predict residual viable nonseminomatous germ cell tumors after chemotherapy. Levels of serum miRNA, including miR-371a-3p, miR-373-3p and miR-367-3p, were measured using the ampTSmiR (amplification targeted serum miRNA) test in 82 patients, including 39 in cohort 1 and 43 in cohort 2, who were treated with orchiectomy, chemotherapy and post-chemotherapy retroperitoneal lymph node dissection. miRNA levels were compared to clinical characteristics and serum tumor markers, and correlated with the presence of viable germ cell tumor vs fibrosis/necrosis and teratoma. ROC analysis was done to determine miRNA discriminative capacity. miRNA levels were significantly associated with disease extent at chemotherapy and they decreased significantly after chemotherapy. Conventional serum tumor marker levels were uninformative after chemotherapy. However, after chemotherapy miRNA levels remained elevated in patients harboring viable germ cell tumor in post-chemotherapy retroperitoneal lymph node dissection specimens. miR-371a-3p demonstrated the highest discriminative capacity for viable germ cell tumors (AUC 0.874, 95% CI 0.774-0.974, p <0.0001). Using an adapted hypothetical cutoff of 3 cm or less for surgical intervention miR-371a-3p correctly stratified all patients with viable residual retroperitoneal germ cell tumors with 100% sensitivity (p = 0.02). Our study demonstrates for the first time the potential value of miR-371a-3p to predict viable germ cell tumors in residual masses after chemotherapy. Prospective studies are required to confirm clinical usefulness.

In urothelial bladder cancer (UBC), risk stratification remains an important unmet need. Limitless self‐renewal, governed by TERT expression and telomerase activation, is crucial for cancer progression. Thus, telomerase activation through the interplay of mutations (TERTp^Mut) and epigenetic alterations in the TERT promoter may provide further insight into UBC behavior. Here, we investigated the combined effect of TERTp^Mut and the TERT Hypermethylated Oncological Region (THOR) status on telomerase activation and patient outcome in a UBC international cohort (n = 237). We verified that TERTp^Mut were frequent (76.8%) and present in all stages and grades of UBC. Hypermethylation of THOR was associated with higher TERT expression and higher‐risk disease in nonmuscle invasive bladder cancers (NMIBC). TERTp^Mut alone predicted disease recurrence (HR: 3.18, 95%CI 1.84 to 5.51, p < 0.0001) but not progression in NMIBC. Combined THOR^high/TERTp^Mut increased the risk of disease recurrence (HR 5.12, p < 0.0001) and progression (HR 3.92, p = 0.025). Increased THOR hypermethylation doubled the risk of stage progression of both TERTp^wt and TERTp^Mut NMIBC. These results highlight that both mechanisms are common and coexist in bladder cancer and while TERTp^Mut is an early event in bladder carcinogenesis THOR hypermethylation is a dynamic process that contributes to disease progression. While the absence of alterations comprises an extremely indolent phenotype, the combined genetic and epigenetic alterations of TERT bring additional prognostic value in NMIBC and provide a novel insight into telomere biology in cancer.

Prostate cancer (PCa) is the most frequently diagnosed cancer in men and the second most common cause of cancer-related mortality among men in the developed world. Conventional anti-PCa therapies include surgery, radiation, hormonal ablation, and chemotherapy. Despite increasing efforts, these therapies are not effective for patients with advanced and/or metastatic disease. In most cases, cancer therapies fail due to an incomplete depletion of tumor cells, resulting in tumor relapse. The cancer stem cell (CSC) hypothesis is an emerging model that explains many of the molecular characteristics of oncological disease as well as the tendency of cancers to relapse, metastasize, and develop resistance to conventional therapies. CSCs are a reservoir of cancer cells that exhibit properties of self-renewal and the ability to reestablish the heterogeneous tumor cell population. The existence of PCa stem cells offers a theoretical explanation for many uncertainties regarding PCa and also for treatment resistance and disease progression once clinical cure is achieved. Therapies targeting CSCs might therefore lead to more effective cancer treatments, divergent from a traditional anti-proliferative approach, based on tumor bulk reduction accompanied by CSC-specific inhibition. Here, we focus on reviewing the historical perspective as well as concepts regarding stem cells and CSCs in PCa. In addition, we will report possible strategies and new clinical approaches that address the CSC-based concept of tumorigenesis in PCa.