Department of Pathology and Laboratory Medicine, Memorial Sloan Kettering Cancer Center, New York, NY, USA.
Department of Pathology, Vancouver General Hospital, Vancouver, BC Canada.
Pathol Res Pract. 2022 May;233:153892. doi: 10.1016/j.prp.2022.153892. Epub 2022 Apr 16.
Telomerase reverse transcriptase (TERT) activation has been shown to be an important cancer hallmark; the activation and expression of TERT has been documented in >90% of tumors and TERT activation has been touted as a prognostic marker in many cancers. However, there is currently no simple testing modality to detect TERT mRNA expression in surgical pathology specimens. In this study we aim to evaluate and validate the utility and reliability of the TERT RNAscope® in-situ hybridization (ISH) assay for the detection of TERT mRNA expression in formalin-fixed, paraffin embedded tissue.
RNAscope® detection for TERT was performed on a Leica Biosystems BOND III research staining robot using the Hs-TERT-O1 (ACD, 481968) probe. Twenty three samples containing 48 tissue types were assessed. TERT genomic alterations were determined by targeted next generation sequencing (NGS), while TERT mRNA expression was determined by both targeted RNA-sequencing and TERT RNAscope® and the results compared. Manual vs automated TERT expression quantification methodologies were evaluated for the ISH assay. The expression levels in normal vs. neoplastic tissues were also compared.
The RNAscope® assay showed high TERT expression in neoplastic tissues, while most normal tissues have no or very low expression levels (p-value= 0.0001, AUC: 0.99). In addition, there was good correlation of TERT expression between the RNAscope® assay and RNA-sequencing. For RNAscope® quantification, manual calculation of TERT signal/cell ratio based on a count of 100 cells was superior compared to automated signal detection.
TERT RNAscope® assay is a simple and reliable tool for the evaluation of TERT mRNA expression. TERT signal/cell ratio based on a count of 100 cells is a reproducible and accurate interpretation approach for evaluation of TERT expression.
端粒酶逆转录酶（TERT）的激活已被证明是一个重要的癌症标志；TERT 的激活和表达已在超过 90%的肿瘤中得到证实，TERT 的激活已被吹捧为许多癌症的预后标志物。然而，目前没有简单的检测方法来检测手术病理标本中的 TERT mRNA 表达。在这项研究中，我们旨在评估和验证 TERT RNAscope®原位杂交（ISH）检测在检测福尔马林固定、石蜡包埋组织中 TERT mRNA 表达的实用性和可靠性。
使用 Leica Biosystems BOND III 研究染色机器人对 TERT 进行 RNAscope®检测，使用 Hs-TERT-O1（ACD，481968）探针。评估了 23 个包含 48 种组织类型的样本。通过靶向下一代测序（NGS）确定 TERT 基因组改变，同时通过靶向 RNA-seq 和 TERT RNAscope®确定 TERT mRNA 表达，并比较结果。评估了 ISH 检测中手动与自动 TERT 表达定量方法。还比较了正常组织与肿瘤组织中的表达水平。
RNAscope®检测显示肿瘤组织中 TERT 表达较高，而大多数正常组织表达水平较低或无（p 值=0.0001，AUC：0.99）。此外，RNAscope®检测与 RNA-seq 之间存在良好的 TERT 表达相关性。对于 RNAscope®定量，基于对 100 个细胞进行计数的 TERT 信号/细胞比值的手动计算优于自动信号检测。
TERT RNAscope®检测是评估 TERT mRNA 表达的简单可靠工具。基于对 100 个细胞进行计数的 TERT 信号/细胞比值是评估 TERT 表达的一种可重复且准确的解释方法。
