title: Real-time RT-PCR quantification of human telomerase reverse transcriptase splice variants in tumor cell lines and non-small cell lung cancer.
abstract: BACKGROUND
  We developed and validated a real-time reverse transcription (RT)-PCR for the quantification of 4 individual human telomerase reverse transcriptase (TERT) splice variants (alpha+beta+, alpha-beta+, alpha+beta-, alpha-beta-) in tumor cell lines and non-small cell lung cancer (NSCLC).
  
  
  METHODS
  We used in silico designed primers and a common TaqMan probe for highly specific amplification of each TERT splice variant, PCR transcript-specific DNA external standards as calibrators, and the MCF-7 cell line for the development and validation of the method. We then quantified TERT splice variants in 6 tumor cell lines and telomerase activity and TERT splice variant expression in cancerous and paired noncancerous tissue samples from 28 NSCLC patients.
  
  
  RESULTS
  In most tumor cell lines, we observed little variation in the proportion of TERT splice variants. The alpha+beta- splice variant showed the highest expression and alpha-beta+ and alpha-beta- the lowest. Quantification of the 4 TERT splice variants in NSCLC and surrounding nonneoplastic tissues showed the highest expression percentage for the alpha+beta- variant in both NSCLC and adjacent nonneoplastic tissue samples, followed by alpha+beta+, with the alpha-beta+ and alpha-beta- splice variants having the lowest expression. In the NSCLC tumors, the alpha+beta+ variant had higher expression than other splice variants, and its expression correlated with telomerase activity, overall survival, and disease-free survival.
  
  
  CONCLUSIONS
  Real-time RT-PCR quantification is a specific, sensitive, and rapid method that can elucidate the biological role of TERT splice variants in tumor development and progression. Our results suggest that the expression of the TERT alpha+beta+ splice variant may be an independent negative prognostic factor for NSCLC patients.
authors.affiliation: Laboratory of Analytical Chemistry, University of Athens, Athens, Greece.
authors.forename: Eleni
authors.initials: E
authors.lastname: Mavrogiannou
authors.forename: Areti
authors.initials: A
authors.lastname: Strati
authors.forename: Aliki
authors.initials: A
authors.lastname: Stathopoulou
authors.forename: Emily G
authors.initials: EG
authors.lastname: Tsaroucha
authors.forename: Loukas
authors.initials: L
authors.lastname: Kaklamanis
authors.forename: Evi S
authors.initials: ES
authors.lastname: Lianidou
mesh_terms: D002289:Carcinoma, Non-Small-Cell Lung / Q000201:enzymology* / Q000401:mortality / Q000473:pathology; D045744:Cell Line, Tumor; D018572:Disease-Free Survival; D006801:Humans; D008175:Lung Neoplasms / Q000201:enzymology* / Q000401:mortality / Q000473:pathology; D008875:Middle Aged; D009364:Neoplasm Recurrence, Local; D020033:Protein Isoforms; D015996:Survival Rate; D019098:Telomerase / Q000235:genetics* / Q000378:metabolism
chemical_list: D020033:Protein Isoforms; D019098:Telomerase