Lacroix J, Becker H D, Woerner S M, Rittgen W, Drings P, von Knebel Doeberitz M
Division of Molecular Diagnostics and Therapy, Department of Surgery, University of Heidelberg, Heidelberg, Germany.
Int J Cancer. 2001 Apr 1;92(1):1-8.
RT-PCR-based amplification of transcripts expressed in cancer but not in normal non-neoplastic cells is increasingly used for the sensitive detection of rare disseminated or exfoliated cancer cells to improve cancer staging and early detection protocols. However, these assays are frequently hampered by false-positive test results due to low-level transcription of the marker genes in normal cells. To overcome these limitations, target transcripts have to be identified that are tightly suppressed in normal non-neoplastic tissues, whereas they should be actively transcribed in the respective cancer cells. Here, we tested RT-PCR assays for 7 neuroendocrine marker transcripts including NCAM, PGP 9.5, gastrin, gastrin receptor, synaptophysin, preprogastrin-releasing peptide (preproGRP) and GRP-receptor to detect rare exfoliated tumor cells in peripheral venous blood and sputum samples from patients with lung cancer. Among these preproGRP RT-PCR was the only assay with which illegitimate transcription in blood or sputum samples from healthy donors or patients with unrelated diseases did not interfere. However, it reproducibly detected up to 10 small-cell lung cancer cells diluted in either 10 ml blood or 5 ml sputum samples. Single blood and sputum samples were collected directly before diagnostic bronchoscopy from 175 patients suspected to have lung cancer. Twenty-six of these had small-cell lung cancer (SCLC). Thereof, 13 patients (50%) tested positive in the blood sample and 5 of 23 patients (22%) tested positive in the sputum sample. Moreover, among 92 patients with non-small-cell lung cancer (NSCLC) 25 patients (27%) had disseminated cancer cells in peripheral blood. Amplification of preproGRP transcripts from clinical samples is a sensitive and specific assay to detect disseminated or exfoliated lung cancer cells either in peripheral blood or sputum samples.
基于逆转录聚合酶链反应（RT-PCR）扩增在癌症中表达但在正常非肿瘤细胞中不表达的转录本，越来越多地用于罕见播散性或脱落癌细胞的灵敏检测，以改善癌症分期和早期检测方案。然而，由于正常细胞中标记基因的低水平转录，这些检测常常受到假阳性检测结果的阻碍。为克服这些局限性，必须鉴定出在正常非肿瘤组织中被严格抑制，而在相应癌细胞中被积极转录的靶转录本。在此，我们检测了用于7种神经内分泌标记转录本的RT-PCR检测方法，包括神经细胞黏附分子（NCAM）、蛋白基因产物9.5（PGP 9.5）、胃泌素、胃泌素受体、突触素、前胃泌素释放肽原（preproGRP）和胃泌素释放肽受体（GRP受体），以检测肺癌患者外周静脉血和痰液样本中罕见的脱落肿瘤细胞。在这些检测方法中，preproGRP RT-PCR是唯一一种健康供体或患有无关疾病患者的血液或痰液样本中的非法转录不干扰检测结果的检测方法。然而，它可重复性地检测到稀释在10毫升血液或5毫升痰液样本中的多达10个小细胞肺癌细胞。在175例疑似患有肺癌的患者中，在诊断性支气管镜检查前直接采集单个血液和痰液样本。其中26例患有小细胞肺癌（SCLC）。其中，13例患者（50%）血液样本检测呈阳性，23例患者中的5例（22%）痰液样本检测呈阳性。此外，在92例非小细胞肺癌（NSCLC）患者中，25例患者（27%）外周血中有播散癌细胞。从临床样本中扩增preproGRP转录本是一种灵敏且特异的检测方法，可检测外周血或痰液样本中播散性或脱落的肺癌细胞。
