Longer genotypically-estimated leukocyte telomere length is associated with increased adult glioma risk

Telomere maintenance has emerged as an important molecular feature with impacts on adult glioma susceptibility and prognosis. Whether longer or shorter leukocyte telomere length (LTL) is associated with glioma risk remains elusive and is often confounded by the effects of age and patient treatment. We sought to determine if genotypically-estimated LTL is associated with glioma risk and if inherited single nucleotide polymorphisms (SNPs) that are associated with LTL are glioma risk factors. Using a Mendelian randomization approach, we assessed differences in genotypically-estimated relative LTL in two independent glioma case-control datasets from the UCSF Adult Glioma Study (652 patients and 3735 controls) and The Cancer Genome Atlas (478 non-overlapping patients and 2559 controls). LTL estimates were based on a weighted linear combination of subject genotype at eight SNPs, previously associated with LTL in the ENGAGE Consortium Telomere Project. Mean estimated LTL was 31bp (5.7%) longer in glioma patients than controls in discovery analyses (P = 7.82×10-8) and 27bp (5.0%) longer in glioma patients than controls in replication analyses (1.48×10-3). Glioma risk increased monotonically with each increasing septile of LTL (O.R.=1.12; P = 3.83×10-12). Four LTL-associated SNPs were significantly associated with glioma risk in pooled analyses, including those in the telomerase component genes TERC (O.R.=1.14; 95% C.I.=1.03-1.28) and TERT (O.R.=1.39; 95% C.I.=1.27-1.52), and those in the CST complex genes OBFC1 (O.R.=1.18; 95% C.I.=1.05-1.33) and CTC1 (O.R.=1.14; 95% C.I.=1.02-1.28). Future work is needed to characterize the role of the CST complex in gliomagenesis and further elucidate the complex balance between ageing, telomere length, and molecular carcinogenesis.     

端粒维持机制及肿瘤治疗新策略

端粒是位于真核细胞染色体末端的特殊蛋白结构,对维持线性染色体的稳定性至关重要,如珑何抑制端粒功能成为改进肿瘤治疗的新亮点.基于抑制端粒酶活性的抗肿瘤治疗存在不足,以端粒维持机制和端粒结构为靶点的抗肿瘤新策略已越来越受到重视,是富有前景的研究方向.     

Telomere maintenance mechanisms in malignant peripheral nerve sheath tumors: expression and prognostic relevance

This study investigated the prevalence and the prognostic relevance of the 2 known telomere maintenance mechanisms (TMMs), telomerase activity (TA) and alternative lengthening of telomeres (ALT), in malignant peripheral nerve sheath tumors (MPNST). In 57 specimens from 49 patients with MPNST (35 sporadic, 14 neurofibromatosis type 1-related), TA was determined using the telomeric repeat amplification protocol, and ALT was detected by assaying ALT-associated promyelocytic leukemia bodies (APB) and terminal restriction fragment (TRF) length distribution. TA or ALT (defined on the basis of APB) alone was found in 24.6% or 26.3% of the lesions, respectively, whereas 6 cases (10.5%) were TA+/ALT+. A concordance between APB and TRF results in defining the ALT status was observed in 44 of 57 cases (77.2%; P < .0001). TA was more frequently expressed in samples from patients with neurofibromatosis type 1 than in those with sporadic disease (60% vs 29.4%, P = 0.087). In the overall series, TA proved to be prognostic for 5-year disease-specific death (hazard ratio, 3.78; 95% confidence interval [CI], 1.60-8.95; P = .002), even when adjusted for the presence of neurofibromatosis type 1 (hazard ratio, 4.22; 95% CI, 1.804-9.874; P = .001) and margin status after surgery (hazard ratio, 5.78; 95% CI, 2.19-15.26; P < .001). Conversely, ALT did not significantly affect clinical outcome of MPNST using either APB expression (hazard ratio, 1.25; 95% CI 0.54-2.89; P = 0.605) or TRF distribution (hazard ratio, 0.57; 95% CI, 0.17-1.96; P = .375) as the detection approach. Our results indicate for the first time that both TMMs, TA and ALT, are present in MPNST and differentially affect patient prognosis.     

Genetic variants in telomerase-related genes are associated with an older age at diagnosis in glioma patients: evidence for distinct pathways of gliomagenesis

Background

Genome-wide association studies have implicated single nucleotide polymorphisms (SNPs) in 7 genes as glioma risk factors, including 2 (TERT, RTEL1) involved in telomerase structure/function. We examined associations of these 7 established glioma risk loci with age at diagnosis among patients with glioma.

Methods

SNP genotype data were available for 2286 Caucasian glioma patients from the University of California, San Francisco (n = 1434) and the Mayo Clinic (n = 852). Regression analyses were performed to test for associations between “number of risk alleles” and “age at diagnosis,” adjusted for sex and study site and stratified by tumor grade/histology where appropriate.

Results

Four SNPs were significantly associated with age at diagnosis. Carrying a greater number of risk alleles at rs55705857 (CCDC26) and at rs498872 (PHLDB1) was associated with younger age at diagnosis (P = 1.4 × 10⁻²² and P = 9.5 × 10⁻⁷, respectively). These SNPs are stronger risk factors for oligodendroglial tumors, which tend to occur in younger patients, and their association with age at diagnosis varied across tumor subtypes. In contrast, carrying more risk alleles at rs2736100 (TERT) and at rs6010620 (RTEL1) was associated with older age at diagnosis (P = 6.2 × 10⁻⁴ and P = 2.5 × 10⁻⁴, respectively). These SNPs are risk factors for all glioma grades/histologies, and their association with age at diagnosis was consistent across tumor subgroups.

Conclusions

Carrying a greater number of risk alleles might be expected to decrease age at diagnosis. However, glioma susceptibility conferred by variation in telomerase-related genes did not follow this pattern. This supports the hypothesis that telomerase-related mechanisms of telomere maintenance are more associated with gliomas that develop later in life than those utilizing telomerase-independent mechanisms (ie, alternative lengthening of telomeres).     

Telomere length and telomerase activity in malignant lymphomas at diagnosis and relapse

Telomere length maintenance, in the vast majority of cases executed by telomerase, is a prerequisite for long-term proliferation. Most malignant tumours, including lymphomas, are telomerase-positive and this activity is a potential target for future therapeutic interventions since inhibition of telomerase has been shown to result in telomere shortening and cell death in vitro. One prerequisite for the suitability of anti-telomerase drugs in treating cancer is that tumours exhibit shortened telomeres compared to telomerase-positive stem cells. A scenario is envisioned where the tumour burden is reduced using conventional therapy whereafter remaining tumour cells are treated with telomerase inhibitors. In evaluating the realism of such an approach it is essential to know the effects on telomere status by traditional therapeutic regimens. We have studied the telomere lengths in 47 diagnostic lymphomas and a significant telomere shortening was observed compared to benign lymphoid tissues. In addition, telomere length and telomerase activity were studied in consecutive samples from patients with relapsing non-Hodgkin's lymphomas. Shortened, unchanged and elongated telomere lengths were observed in the relapse samples. The telomere length alterations found in the relapsing lymphomas appeared to be independent of telomerase and rather represented clonal selection random at the telomere length level. These data indicate that anti-telomerase therapy would be suitable in only a fraction of malignant lymphomas.     

A preclinical mouse model of glioma with an alternative mechanism of telomere maintenance (ALT)

Glioblastoma multiforme is the most aggressive primary tumor of the central nervous system. Glioma stem cells (GSCs), a small population of tumor cells with stem‐like properties, are supposedly responsible for glioblastoma multiforme relapse after current therapies. In approximately thirty percent of glioblastoma multiforme tumors, telomeres are not maintained by telomerase but through an alternative mechanism, termed alternative lengthening of telomere (ALT), suggesting potential interest in developing specific therapeutic strategies. However, no preclinical model of ALT glioma was available until the isolation of TG20 cells from a human ALT glioma. Herein, we show that TG20 cells exhibit a high level of telomeric recombination but a stable karyotype, indicating that their telomeres retain their protective function against chromosomal instability. TG20 cells possess all of the characteristic features of GSCs: the expression of neural stem cell markers, the generation of intracerebral tumors in NOD‐SCID‐IL2Rγ (NSG) mice as well as in nude mice, and the ability to sustain serial intracerebral transplantations without expressing telomerase, demonstrating the stability of the ALT phenotype in vivo. Furthermore, we also demonstrate that 360B, a G‐quadruplex ligand of the pyridine derivative series that impairs telomere replication and mitotic progression in cancer cells, prevents the development of TG20 tumors. Together, our results show that intracerebral grafts of TG20 cells in immunodeficient mice constitute an efficient preclinical model of ALT glioblastoma multiforme and that G‐quadruplex ligands are a potential therapy for this specific type of tumor.     

Occupation and the risk of adult glioma in the United States

Objective: Previous studies have observed increased glioma incidence associated with employment in the petroleum and electrical industries, and in farming. Several other occupations have also been associated with increased risk, but with inconsistent results. We evaluated associations between occupational title and glioma incidence in adults. Methods: Cases were 489 patients with glioma diagnosed from 1994 to 1998 at three United States hospitals. Controls were 799 patients admitted to the same hospitals for non-malignant conditions. An experienced industrial hygienist grouped occupations that were expected to have similar tasks and exposures. The risk of adult glioma was evaluated for those subjects who ever worked in an occupational group for at least six months, those who worked longer than five years in the occupation, and those with more than ten years latency since starting work in the occupation. Results: Several occupational groups were associated with increased glioma incidence for having ever worked in the occupation, including butchers and meat cutters (odds ratio [OR] = 2.4; 95% confidence limits [CL]: 1.0, 6.0), computer programmers and analysts (OR = 2.0; 95% CL: 1.0, 3.8), electricians (OR = 1.8; 95% CL: 0.8, 4.1), general farmers and farmworkers (OR = 2.5; 95% CL: 1.4, 4.7), inspectors, checkers, examiners, graders, and testers (OR = 1.5; 95% CL: 0.8, 2.7), investigators, examiners, adjustors, and appraisers (OR = 1.7; 95% CL: 0.8, 3.7), physicians and physician assistants (OR = 2.4; 95% CL: 0.8, 7.2), and store managers (OR = 1.6; 95% CL: 0.8, 3.1), whereas occupation as a childcare worker was associated with decreased glioma incidence (OR = 0.4; 95% CL: 0.2, 0.9). These associations generally persisted when the subjects worked longer than five years in the occupation, and for those with more than ten years latency since starting to work in the occupation. Conclusions: This is our first analysis of occupation and will guide future exposure-specific assessments.     

Telomere maintenance and dysfunction predict recurrence in paediatric ependymoma

We have recently described the enzymatic subunit of telomerase (hTERT) as an important prognostic marker for paediatric ependymoma. Because of the lack of good, representative pre-clinical models for ependymoma, we took advantage of our large cohort of ependymoma patients, some with multiple recurrences, to investigate telomere biology in these tumours. Our cohort consisted of 133 ependymomas from 83 paediatric patients and included 31 patients with recurrences. Clinical outcome was measured as overall survival, progression-free survival and response to therapy. In all 133 tumours, hTERT expression correlated with proliferative markers, including MIB-1 index (P<0.0001) and mitotic index (P=0.005), as well as overall tumour grade (P=0.001), but not with other markers of anaplasia. There was no correlation between telomere length and hTERT expression or survival. Surprisingly, prior radiation or chemotherapy neither induced sustained DNA damage nor affected telomere maintenance in recurrent tumours. There was an inverse correlation between hTERT expression and telomere dysfunction as measured by gamma H2AX expression (P=0.016). Combining gamma H2AX and hTERT expressions could segregate tumours into three different survival groups (log rank, P<0.0001) such that those patients whose tumours expressed hTERT and showed no evidence of DNA damage had the worst outcome. This study emphasises the importance of telomere biology as a prognostic tool and telomerase inhibition as a therapeutic target for paediatric ependymoma. Furthermore, we have demonstrated that analysing tumours as they progress in vivo is a viable approach to studying tumour biology in humans.     

Glioma risk associated with extent of estimated European genetic ancestry in African Americans and Hispanics

Glioma incidence is highest in non-Hispanic Whites, and to date, glioma genome-wide association studies (GWAS) to date have only included European ancestry (EA) populations. African Americans and Hispanics in the US have varying proportions of EA, African (AA) and Native American ancestries (NAA). It is unknown if identified GWAS loci or increased EA is associated with increased glioma risk. We assessed whether EA was associated with glioma in African Americans and Hispanics. Data were obtained for 832 cases and 675 controls from the Glioma International Case–Control Study and GliomaSE Case–Control Study previously estimated to have <80% EA, or self-identify as non-White. We estimated global and local ancestry using fastStructure and RFMix, respectively, using 1,000 genomes project reference populations. Within groups with ≥40% AA (AFR_≥0.4), and ≥15% NAA (AMR_≥0.15), genome-wide association between local EA and glioma was evaluated using logistic regression conditioned on global EA for all gliomas. We identified two regions (7q21.11, p = 6.36 × 10⁻⁴; 11p11.12, p = 7.0 × 10⁻⁴) associated with increased EA, and one associated with decreased EA (20p12.13, p = 0.0026) in AFR_≥0.4. In addition, we identified a peak at rs1620291 (p = 4.36 × 10⁻⁶) in 7q21.3. Among AMR_≥0.15, we found an association between increased EA in one region (12q24.21, p = 8.38 × 10⁻⁴), and decreased EA in two regions (8q24.21, p = 0. 0010; 20q13.33, p = 6.36 × 10⁻⁴). No other significant associations were identified. This analysis identified an association between glioma and two regions previously identified in EA populations (8q24.21, 20q13.33) and four novel regions (7q21.11, 11p11.12, 12q24.21 and 20p12.13). The identifications of novel association with EA suggest regions to target for future genetic association studies.     

Telomere uncapping by the G-quadruplex ligand RHPS4 inhibits clonogenic tumour cell growth in vitro and in vivo consistent with a cancer stem cell targeting mechanism

The pentacyclic acridinium methosulfate salt RHPS4 induces the 3'single-stranded guanine-rich telomeric overhang to fold into a G-quadruplex structure. Stabilisation of the latter is incompatible with an attachment of telomerase to the telomere and thus G-quadruplex ligands can effectively inhibit both the catalytic and capping functions of telomerase. In this study, we examined mechanisms underlying telomere uncapping by RHPS4 in uterus carcinoma cells (UXF1138L) with short telomeres and compared the susceptibility of bulk and clonogenic cancer cells to the G-quadruplex ligand. We show that treatment of UXF1138L cells with RHPS4 leads to the displacement of the telomerase catalytic subunit (hTERT) from the nucleus, induction of telomere-initiated DNA-damage signalling and chromosome fusions. We further report that RHPS4 is more potent against cancer cells that grow as colonies in soft agar than cells growing as monolayers. Human cord blood and HEK293T embryonic kidney cell colony forming units, however, were more resistant to RHPS4. RHPS4-treated UXF1138L xenografts had a decreased clonogenicity, showed loss of nuclear hTERT expression and an induction of mitotic abnormalities compared with controls. Although single-agent RHPS4 had limited in vivo efficacy, a combination of RHPS4 with the mitotic spindle poison Taxol caused tumour remissions and further enhancement of telomere dysfunction.     

Association between leukocyte telomere length and glioma risk: a case-control study

Background

Compelling epidemiological evidence indicates that alterations of telomere length are associated with risks of many malignancies in a tumor-specific manner, such as lung cancer, breast cancer, and non-Hodgkin''s lymphoma. However, the association between leukocyte telomere length and glioma risk has not been investigated.

Methods

Relative telomere length (RTL) of peripheral blood leukocytes from 467 glioma patients and 467 healthy controls, matched by age and sex, was measured using the real-time PCR-based method in a case-control study. An unconditional multivariate logistic regression model was applied to estimate the association between RTL and glioma risk.

Results

Glioma patients showed notably longer RTL than controls (median, 0.555 vs 0.444; P > .04). RTL was negatively correlated with age in both cases (ρ = −0.430; P < .001) and controls (ρ = −0.388; P < .001). After adjusting for age, sex, smoking status and family history of cancer, multivariate logistic regression analysis showed that there was a U-shaped association between RTL and glioma risk (P for nonlinearity <.001). Compared with individuals in the second tertile of RTL, the odds ratios (95% CI) for participants in the first and third tertiles were 2.16 (range, 1.52–3.09) and 3.51 (range, 2.45–5.00), respectively. Stratified analysis showed that the association between RTL and glioma risk was not modulated by major host characteristics.

Conclusions

Our study demonstrates for the first time that either shorter or longer RTL in peripheral blood leukocytes is associated with increased glioma risk, which warrants further investigation in the future.     

端粒酶的起源、调控及与肿瘤关系的研究进展

端粒酶的激活与恶性肿瘤发生发展之间存在着密切的关系。有效地抑制端粒酶活性有可能导致肿瘤治疗方面的重大突破。有关端粒酶自身起源及结构的深入研究有助于这一问题的解决。本文对端粒酶的起源、组分、功能调控研究方面的最新进展进行了综述，并将端凿酶与肿瘤的关系予以总结和展望。     

Telomere dysfunction and chromothripsis

Chromothripsis is a recently discovered form of genomic instability, characterized by tens to hundreds of clustered DNA rearrangements resulting from a single dramatic event. Telomere dysfunction has been suggested to play a role in the initiation of this phenomenon, which occurs in a large number of tumor entities. Here, we show that telomere attrition can indeed lead to catastrophic genomic events, and that telomere patterns differ between cells analyzed before and after such genomic catastrophes. Telomere length and telomere stabilization mechanisms diverge between samples with and without chromothripsis in a given tumor subtype. Longitudinal analyses of the evolution of chromothriptic patterns identify either stable patterns between matched primary and relapsed tumors, or loss of the chromothriptic clone in the relapsed specimen. The absence of additional chromothriptic events occurring between the initial tumor and the relapsed tumor sample points to telomere stabilization after the initial chromothriptic event which prevents further shattering of the genome.     

端粒保护蛋白TRF2在肿瘤发生和治疗中作用机制的研究进展

端粒重复序列结合因子2（telomeric-repeat binding factor 2,TRF2）是一种重要的端粒保护蛋白,与端粒保护蛋白1（protection of telomeres 1,POT1）、端粒重复序列结合因子1（telomeric-repeat binding factor 1,TRF1）、相互作用核蛋白2（TRF1-interacting nuclear protein 2,TIN2）、阻滞活化蛋白1（repressor activator protein 1,Rap1）以及TPP1,5个核心蛋白通过一系列相互作用共同形成端粒保护蛋白复合体（shelterin）以维持端粒结构和功能的稳定性和完整性。越来越多的研究显示TRF2在多种肿瘤中异常表达并与肿瘤的发生,肿瘤细胞的耐药以及肿瘤血管的生成密切相关。因此,本文就TFR2的结构与生理功能,以及其在肿瘤发生,发展与治疗中的作用研究进展进行综述,以期为肿瘤的预防和治疗提供新的思路。      

蟾蜍灵通过抑制端粒酶活性诱导人脑胶质瘤细胞死亡

目的:探讨蟾蜍灵对人脑胶质瘤的作用。方法:将不同浓度的蟾蜍灵作用于胶质瘤细胞后,用CCK-8法检测细胞增殖抑制率,克隆形成实验检测细胞法检测增殖抑制,Western-blot测定人端粒酶逆转录酶（hTERT）的蛋白表达,Real time-PCR检测hTERT基因表达水平,CCK-8法检测蟾蜍灵对人原代胶质瘤细胞的杀伤作用。结果:蟾蜍灵对胶质瘤细胞株细胞U251、U87增殖具有抑制作用,且呈时间和剂量依赖性,并且能够有效抑制U251及U87细胞的克隆形成。Western-blot结果显示hTERT蛋白表达随药物浓度增加而递减,Real time-PCR结果证实蟾蜍灵降低了hTERT基因表达。同时体外实验证明,蟾蜍灵对人原代胶质瘤细胞具有明显杀伤作用。结论:蟾蜍灵能够抑制胶质瘤细胞系细胞及人原代胶质瘤细胞增殖,同时通过抑制hTERT表达降低端粒酶活性以发挥其抗肿瘤的作用。     

Exophytic malignant brainstem mixed glioma in an adult: A case report

Anaplastic mixed gliomas are rare tumors that occur mostly in the cerebral hemispheres. They have a distinctive histological appearance characterized by the presence of two or more glial cellular constituents. The incidence of malignant mixed glioma of the brainstem and posterior fossa is extremely low. The authors report an unusual case of an exophytic malignant mixed glioma. Following subtotal resection, the patient received conventional radiotherapy, but continued to deteriorate, and died five months after surgery. The extensive literature review focuses on histopathology, clinical features, natural history, and possible treatment modalities of this unusual neoplasm.