Juhasz Agnes, Vassilakos Aikaterini, Chew Helen K, Gandara David, Yen Yun
Department of Medical Oncology and Therapeutics Research, City of Hope Comprehensive Cancer Center, Duarte, CA 91010, USA.
Oncol Rep. 2006 May;15(5):1299-304.
This study describes the development of a rapid and practical real-time RT-PCR method to quantify ribonucleotide reductase M2 (RRM2) mRNA in tumor and peripheral white blood cells (WBCs) from patients treated with GTI-2040, an antisense drug currently in clinical trials. In order to assess target down-regulation by GTI-2040, RRM2 mRNA expression levels were analyzed in pre- and post-treatment samples from a phase II clinical trial of GTI-2040 combined with capecitabine in patients with metastatic breast cancer. Target gene RRM2 mRNA levels were evaluated using quantitative RT-PCR method: real-time PCR (TaqMan) with fluorescein labeled probes on an ABI 7900HT instrument, with additional post-processing of the data to adjust for differences in total RNA in-put across the samples. Data are presented from a patient for whom both biopsy and PBMC samples were available, demonstrating applicability of this reproducible, highly sensitive real-time RT-PCR method for the detection and quantification of mRNAs for RRM2 in human WBC and tissue samples. By providing quantitative measurement of changes in target gene expression, this method may provide an opportunity to determine the correlation between target response to GTI-2040 antisense and clinical response in patients. Furthermore this assay may assess whether WBC samples are an appropriate surrogate tissue for approximating target down-regulation in the tumor.
本研究描述了一种快速实用的实时逆转录聚合酶链反应（RT-PCR）方法的开发，该方法用于定量检测接受GTI-2040（一种目前正在进行临床试验的反义药物）治疗的患者肿瘤组织和外周血白细胞（WBC）中的核糖核苷酸还原酶M2（RRM2）信使核糖核酸（mRNA）。为了评估GTI-2040对靶标的下调作用，在一项GTI-2040联合卡培他滨用于转移性乳腺癌患者的II期临床试验的治疗前和治疗后样本中，分析了RRM2 mRNA的表达水平。使用定量RT-PCR方法评估靶基因RRM2 mRNA水平：在ABI 7900HT仪器上使用荧光素标记探针进行实时PCR（TaqMan），并对数据进行额外的后处理，以校正各样本中总RNA输入量的差异。数据来自一位同时有活检样本和外周血单个核细胞（PBMC）样本的患者，证明了这种可重复、高度灵敏的实时RT-PCR方法可用于检测和定量人类WBC及组织样本中RRM2的mRNA。通过提供靶基因表达变化的定量测量，该方法可能为确定GTI-2040反义药物的靶标反应与患者临床反应之间的相关性提供机会。此外，该检测方法可评估WBC样本是否是用于近似肿瘤中靶标下调的合适替代组织。
