hTERT Gene Amplification Research Articles

Hepatitis C virus and schistosomiasis as a causative factor for hTERT amplification in hepatocellular carcinoma

Telomere abnormalities appear to play a role in carcinogenesis. hTERT is the catalytic component of the telomerase complex. It is regulates telomerase activity in cancers such as hepatocellular carcinoma (HCC). Amplification of the hTERT gene have been detected in about 21% of HCC. HCC occurs frequently in patients with chronic viral hepatitis or liver cirrhosis. Hepatitis B virus (HBV) or hepatitis C virus (HCV) induce HCC. The schistosomal parasite plays a role in the development of HCC through modify the course of hepatitis C accelerating progression to hepatitis C-associated fibrosis and thus quicker progression to HCC. The present study was conducted to evaluate hTERT amplification in HCC in association with HCV and schistosomiasis. Fifteen surgically resected HCC (Grade two) and fourteen cirrhotic resected liver were examined using FISH technique. Amplification of hTERT gene were found in 80% (12/15) of HCC which is much higher than the previously reported data. Eight samples were HCV and schistosomiasis positive, 4 were HCV positive, 2 were schistosomiasis positive and one was HCV and schistosomiasis negative. Amplification of the hTERT gene was not observed in cirrhotic tissues except two cases which is probably due to extension of adjacent HCC. From our results, the coexistence of HCV and/or schistosomiasis with hTERT amplification, we can conclude that the HCV and schistosomiasis may play an important role in upregulation of this gene. Consequently, this may lead to the development of HCC rather than just a marker for cancer development. However this conclusion might need further investigation.

Multitarget fluorescence in-situ hybridization assays in detecting abnormal cells in bronchial washing and biopsied lung cancer cells

Objective Lung carcinomas are associated with molecular abnormalities. We performed a pilot study to determine the feasibility of an interphase fluorescence in-situ hybridization (FISH) assay for the detection of lung carcinoma in bronchial washing specimens and tissue biopsy. Materials and methods Twenty bronchial washing specimens were analyzed. The LAVysion multicolor probe set, which contains locus-specific probes to 5p15, 7p12 (EGFR), and 8q24 (CMYC) and a centromeric probe to chromosome 6, was used; hTERT gene (5p15) red and control region EGR1 gene (5q31) green probes were also used. Results FISH positivity in the samples of this study was 60% (12/20) for LAVysion probes and only 33.3% (6/20) for hTERT probe. Conclusion The current study has indicated that multitarget FISH assays have shown higher sensitivity in detecting abnormal cells in bronchial washing and biopsied cells from patients with lung cancer; in addition, hTERT gene amplification or high copy number could be a marker for very poor prognosis in early-stage non-small-cell lung cancer patients.

GENETIC OF THYROID CANCER FAMILIAL NON MEDULLARY THYROID CANCER

Differentiated non-medullary thyroid cancer (NMTC) is mostly sporadic, but the recurrence of familial form of the disease has been reported. Short or dysfunctional telomeres have been associated with familial benign diseases and familial breast cancer. We aimed to study the telomere-telomerase complex in familial NMTC (FNMTC). The genetic analysis included the measurement in the peripheral blood of relative telomere length (RTL), telomerase reverse transcriptase (hTERT) gene amplification, hTERT mRNA expression, telomerase protein activity and search of hTERT or TERC (telomerase RNA component) gene mutations. We, also, studied telomeric fusions and associations as well as other chromosomal fragility features by conventional and molecular cytogenetic analyses, in phytohemagglutinin stimulated T-lymphocytes from familial patients, unaffected family members, sporadic PTC patients and healthy subjects. We found that, telomere lenght was significantly shorter in the blood of familial patients compared to sporadic PTCs, healthy subjects, nodular goiter and unaffected siblings. hTERT gene amplification was significantly higher in FNMTC patients compared to the other groups and, in particular, it was significantly greater in offspring with respect to parents. hTERT mRNA expression as well as telomerase activity were significantly higher in FNMTC patients compared to sporadic In addition, we demonstrated that familial patients have a significant increase in spontaneous telomeric associations and telomeric fusions compared to healthy subjects and sporadic cases. Q-FISH analysis demonstrated that familial cases display a significant decrease in the telomeric PNA-FISH signal intensity in metaphase chromsome. Our study demonstrates that patients with FNMTC display an imbalance of the telomeretelomerase complex in the peripheral blood.

GENETIC OF THYROID CANCER FAMILIAL NON MEDULLARY THYROID CANCER

Differentiated non-medullary thyroid cancer (NMTC) is mostly sporadic, but the recurrence of familial form of the disease has been reported. Short or dysfunctional telomeres have been associated with familial benign diseases and familial breast cancer. We aimed to study the telomere-telomerase complex in familial NMTC (FNMTC). The genetic analysis included the measurement in the peripheral blood of relative telomere length (RTL), telomerase reverse transcriptase (hTERT) gene amplification, hTERT mRNA expression, telomerase protein activity and search of hTERT or TERC (telomerase RNA component) gene mutations. We, also, studied telomeric fusions and associations as well as other chromosomal fragility features by conventional and molecular cytogenetic analyses, in phytohemagglutinin stimulated T-lymphocytes from familial patients, unaffected family members, sporadic PTC patients and healthy subjects. We found that, telomere lenght was significantly shorter in the blood of familial patients compared to sporadic PTCs, healthy subjects, nodular goiter and unaffected siblings. hTERT gene amplification was significantly higher in FNMTC patients compared to the other groups and, in particular, it was significantly greater in offspring with respect to parents. hTERT mRNA expression as well as telomerase activity were significantly higher in FNMTC patients compared to sporadic In addition, we demonstrated that familial patients have a significant increase in spontaneous telomeric associations and telomeric fusions compared to healthy subjects and sporadic cases. Q-FISH analysis demonstrated that familial cases display a significant decrease in the telomeric PNA-FISH signal intensity in metaphase chromsome. Our study demonstrates that patients with FNMTC display an imbalance of the telomeretelomerase complex in the peripheral blood.

HTERT Gene Amplification and Clinical Significance in Pleural Effusions of Patients With Lung Cancer

Patients and MethodsHuman telomerase reverse transcriptase (hTERT) gene amplification was detected in pleural effusions of patients with lung cancer (n = 69) and in patients with benign lung disease (n = 46) when using a quantitative polymerase chain reaction (qPCR) technique. ResultshTERT gene relative copy numbers were significantly higher in effusions from patients with malignant, adenocarcinoma and small-cell lung cancer than in effusions from patients with benign lung disease (P < .01). By using a threshold value of 1.39, hTERT gene amplification was significantly more frequent in malignant effusions compared with benign effusions and more likely to be positive for malignant effusions, compared with cytology (P < .01). The diagnostic performance of qPCR of hTERT gene amplification was significantly higher than that of cytology, in terms of sensitivity (91.3% vs. 56.5%), negative predictive value (87.8% vs. 60.5%), and accuracy (92.2% vs. 73.9%). ConclusionsDetecting hTERT gene amplification by qPCR appears suitable for distinguishing carcinoma cells from reactive mesothelial cells in pleural effusions. hTERT gene amplification was more sensitive than cytology and may be useful for diagnosing pleural micrometastases.

Characterization of arsenic-induced cytogenetic alterations in acute promyelocytic leukemia cell line, NB4

Gain or loss of genes plays important roles in leukemogenesis of APL via cooperation with PML-RARA. Fluorescence in situ hybridization (FISH) was applied to investigate the DNA copy number changes of hTERT, ERG, CDKN1B (P27), CDKN2A (P16), and TP53 genes in an acute promyelocytic leukemia (APL) cell line (NB4). Five bacterial artificial chromosome probes (BAC) for 9p21.3, 17p13.1, 12p13.2, 5p15.33, 21q22.2 regions were prepared using sequence independent amplification (SIA) and were hybridized to NB4 cells treated with different doses of arsenic trioxide (As(2)O(3); ATO) at various time intervals. NB4 cells were also karyotyped by G-banded chromosome analysis 24 h after culture initiation. FISH analysis prior to treatment showed CDKN1B, CDKN2A, and TP53 gene deletion but ERG and hTERT gene amplification. After treatment with ATO, the number of the NB4 cells with deleted CDKN1B and CDKN2A as well as the counts of the cells with hTERT amplification was significantly reduced in time- and does-dependent manners. In addition, we observed expressive increase in signal patterns of CDKN1B and CDKN2A along with significant decline in hTERT signal patterns in ATO-treated cells as compared with the control group (in time- and dose-dependent manners). On the other hand, no difference in signal patterns for Erg and p53 was observed in response to ATO exposure. The results of the present study show the cytogenetic alteration in hTERT, CDKN1B, and CDKN2A in NB4 cells after treatment with ATO might introduce a new mechanism of antitumor activities of ATO in APL cell line, NB4.

Short Telomeres, Telomerase Reverse Transcriptase Gene Amplification, and Increased Telomerase Activity in the Blood of Familial Papillary Thyroid Cancer Patients

Differentiated papillary thyroid cancer is mostly sporadic, but the recurrence of the familial form has been reported. Short or dysfunctional telomeres have been associated with familial benign diseases and familial breast cancer. The aim of our work was to study the telomere-telomerase complex in the peripheral blood of patients with familial papillary thyroid cancer (FPTC), including the measurement of relative telomere length (RTL), telomerase reverse transcriptase (hTERT) gene amplification, hTERT mRNA expression, telomerase protein activity, and search of hTERT or telomerase RNA component gene mutations. Cumulating a series of patients seen at the University of Siena and a series at the University of Rome, the experiments were conducted in 47 FPTC patients, 75 sporadic papillary thyroid cancer (PTC) patients, 20 patients with nodular goiter, 19 healthy subjects, and 20 unaffected siblings of FPTC patients. RTL, measured by quantitative PCR, was significantly (P < 0.0001) shorter in the blood of FPTC patients, compared with sporadic PTCs, healthy subjects, nodular goiter subjects, and unaffected siblings. Also by fluorescence in situ hybridization analysis, the results confirmed shorter telomere lengths in FPTC patients (P = 0.01). hTERT gene amplification was significantly (P < 0.0001) higher in FPTC patients, compared with the other groups, and in particular, it was significantly (P = 0.03) greater in offspring with respect to parents. hTERT mRNA expression, as well as telomerase activity, was significantly higher (P = 0.0003 and P < 0.0001, respectively) in FPTC patients, compared with sporadic PTCs. RTL, measured in cancer tissues, was shorter (P < 0.0001) in FPTC patients, compared with sporadic PTCs. No mutations of the telomerase RNA component and hTERT genes were found. Our study demonstrates that patients with FPTC display an imbalance of the telomere-telomerase complex in the peripheral blood, characterized by short telomeres, hTERT gene amplification, and expression. These features may be implicated in the inherited predisposition to develop FPTC.

Amplification of telomerase (hTERT) gene is a poor prognostic marker in non-small-cell lung cancer

Telomerase reactivation is a hallmark of human carcinogenesis. Increased telomerase activity may result from gene amplification and/or overexpression. This study evaluates the prognostic value of hTERT gene amplification and mRNA overexpression in 144 resectable non-small-cell lung cancer (NSCLC) specimens. The hTERT gene copy number was assessed by quantitative polymerase chain reaction (qPCR) on laser-capture microdissected tumour cells of 81 tumours, and by fluorescence in situ hybridisation (FISH) on a subset of 59 tumours. hTERT mRNA level was determined by reverse transcription (RT)–qPCR in 130 tumours. In total, 57% of (46 out of 81) primary NSCLC specimens demonstrated hTERT amplification, which was significantly more common (P<0.001) in adenocarcinoma (30 out of 40) than in squamous cell carcinoma (13 out of 37). The hTERT mRNA overexpression was noted in 74% (94 out of 130) of tumours; it was more frequent in squamous cell than in adenocarcinoma (87 vs 68%, P=0.03). Overexpression was significantly associated with amplification (P=0.03), especially in adenocarcinoma. The hTERT gene amplification was prognostic for shorter recurrence-free survival (hazard ratio=2.16, P=0.03). These data indicate that gene amplification is an important mechanism for hTERT overexpression in lung adenocarcinoma and is an independent poor prognostic marker for disease-free survival in NSCLC.

Telomerase reverse transcriptase gene amplification in hepatocellular carcinoma.

Telomerase activation is essential for the immortality of cancer cells. The expression of telomerase reverse transcriptase (hTERT), the catalytic component of the telomerase complex, regulates telomerase activity in human cancers. Amplification of the hTERT gene, located at chromosome 5p, is thought to be a potential genetic event contributing to telomerase activation in sporadic tumors. The amplification of the hTERT gene was examined in 46 surgically resected hepatocellular carcinomas (HCC) by real-time polymerase chain reaction and the status was compared with the expression of hTERT mRNA and clinicopathological parameters. Amplified hTERT genes were found in 21.7% (10/46) of HCC. The incidence of amplified hTERT genes in poorly differentiated HCC (6/12, 50%) was significantly higher than that in highly to moderately differentiated HCC (4/34, 11.8%; P = 0.012). Tumor size in those cases with hTERT gene amplification was larger compared to those cases with no amplification (P = 0.047). Amplification of the hTERT gene was not observed in non-cancerous tissues. The hTERT mRNA level did not correlate with the number of hTERT genes. Based on these results, it is thought that hTERT gene amplification is a cancer-specific event, and may furthermore contribute to the dedifferentiation and development of HCC. However, hTERT gene overexpression was rarely due to an increased hTERT gene copy number in HCC.

HTERT Gene Amplification and Increased mRNA Expression in Central Nervous System Embryonal Tumors

High-level gains at 5p15, a chromosomal region including the human telomerase catalytic protein subunit (hTERT) gene, have been documented in several medulloblastomas. We therefore analyzed hTERT gene dosage in a group of medulloblastomas and other embryonal brain tumors using differential PCR. Amplification of the hTERT locus was detected in 15 of 36 (42%) tumors examined. To correlate gene amplification with message level, we used real-time quantitative PCR to measure hTERT mRNA in 50 embryonal brain tumors. hTERT mRNA was detected in all but one of these cases, and mRNA level correlated significantly with gene dosage (r = 0.82). Log-rank analysis of survival data revealed a trend toward poor clinical outcomes in patients with medulloblastomas containing high hTERT mRNA levels, but clinical follow-up was relatively short and the association was not statistically significant (P = 0.078). Comparative genomic hybridization was used to further analyze the tumor with the greatest hTERT gene dosage and mRNA level, a recurrent medulloepithelioma. hTERT was amplified in the recurrent tumor but not in the primary lesion, suggesting this locus can be involved in tumor progression. Our data indicate that hTERT gene amplification is relatively common in embryonal brain tumors, and that increased expression of hTERT mRNA may be associated with biologically aggressive tumor behavior.

Amplification of the telomerase reverse transcriptase (hTERT) gene in cervical carcinomas.

The expression of telomerase reverse transcriptase (hTERT), the catalytic component of the telomerase complex, is required for activation of telomerase during immortalization and transformation of human cells. However, the biochemical and genetic mechanisms governing hTERT expression remain to be elucidated. In the present study, we examined hTERT amplification as a potential genetic event contributing to telomerase activation in cervical carcinomas. An amplification of the hTERT gene was found in 1/4 cervical cancer cell lines and 21/88 primary tumor samples derived from the patients with cervical carcinomas. An increase in the hTERT copy number was significantly correlated with higher levels of hTERT protein expression. Moreover, the hTERT alterations with the enhanced hTERT expression were exclusively observed in those tumors with high-risk human papillomavirus infection. Taken together, the hTERT gene amplification, directly or indirectly targeted by human papillomavirus, may be one of the driving forces responsible for upregulation of hTERT expression and activation of telomerase in cervical cancers.