Krebswissenschaft und -therapie

Rapid Quantification of Telomerase Activity Employing an Improved Real-time Telomeric Repeat Amplification Protocol in Clinical Tissue Samples Eliminates Interference by PCR Inhibitors

Abstract

Telomerase, a ribonucleoprotein with reverse transcriptase activity, enables human cells to maintain chromosomal stability and to proliferate without limits. Various studies demonstrated telomerase activation in human cancer, including hepatocellular carcinoma. Therefore, quantification of telomerase activity has been proposed as diagnostic and prognostic tool. In this study, we optimized a 1-step real-time quantitative telomeric repeat amplification protocol for the robust and rapid quantification of telomerase activity in clinical tissue samples. To ensure undisturbed PCR kinetics even in samples with high telomerase activity, we initially determined optimal sample dilution for our assay. Next, we assessed highly diluted samples and did not observe relevant interference by tissue inhibitors of PCR, which constitute a major problem analyzing clinical tissue samples with end-point assays. To test our real-time assay, we evaluated human liver samples and detected increased telomerase activity in malignant liver lesions, whereas benign liver tissue displayed only minimal telomerase activity. In conclusion, our optimized assay is suitable to quantify telomerase activity in clinical tissue samples without interference by PCR inhibitors. The assay may be employed to detect telomerase activity during carcinogenesis and to monitor telomerase activity during cancer progression and treatment.