Nollau P, Moser C, Weinland G, Wagener C
Abteilung für Klinische Chemie, Medizinische Klinik, Universitätskrankenhaus Eppendorf, Hamburg, Germany.
Int J Cancer. 1996 May 3;66(3):332-6. doi: 10.1002/(SICI)1097-0215(19960503)66:3<332::AID-IJC11>3.0.CO;2-D.
Mutant-enriched PCR was applied to the detection of mutations at codons 12 and 13 of K-ras genes in the stools of patients with colorectal cancer. Mutations were analyzed in stool samples obtained prior to surgery. Resected tumor specimens were screened for K-ras mutations by PCR-mediated RFLP analysis. Using normal stool samples, assay conditions were adjusted to optimal sensitivity and specificity. The following specimens were included in the study: 16 stool samples corresponding to carcinomas in which K-ras mutations had been identified; 7 randomly selected stool samples corresponding to carcinomas which were negative for K-ras mutations; 1 stool sample from a patient with non-Hodgkin's lymphoma. In 13 of the 16 stool samples (81%) corresponding to tumors in which K-ras mutations had been identified previously, K-ras mutations were detected. In 2 of the 7 stool samples corresponding to tumors in which K-ras mutations had not been detected by previous PCR-mediated RFLP analysis, K-ras mutations were also present. Reanalyses of the tumors corresponding to these 2 positive stool samples by mutant-enriched PCR revealed a K-ras mutation in one of the tumors. The stool and tumor of the patient with non-Hodgkins lymphoma were negative for K-ras mutations. DNA sequence analysis revealed that, for each of the K-ras mutations identified in stool samples, identical base substitutions were present in the corresponding tumor tissue. The results indicate that tumor cells harboring K-ras mutations can be detected in the stools of patients with colorectal cancer by mutant-enriched PCR with high sensitivity and specificity. Because of the simplicity of the technique, it may be suitable for screening of stool samples for mutations of the K-ras gene.
采用富含突变体的聚合酶链反应（PCR）检测结直肠癌患者粪便中K-ras基因第12和13密码子的突变情况。对手术前采集的粪便样本进行突变分析。通过PCR介导的限制性片段长度多态性（RFLP）分析对切除的肿瘤标本进行K-ras突变筛查。使用正常粪便样本将检测条件调整至最佳灵敏度和特异性。本研究纳入了以下标本：16份与已鉴定出K-ras突变的癌组织相对应的粪便样本；7份随机选取的与K-ras突变阴性的癌组织相对应的粪便样本；1份来自非霍奇金淋巴瘤患者的粪便样本。在之前已鉴定出K-ras突变的16份肿瘤相对应的粪便样本中，有13份（81%）检测到K-ras突变。在之前通过PCR介导的RFLP分析未检测到K-ras突变的7份肿瘤相对应的粪便样本中，有2份也存在K-ras突变。对这2份粪便样本呈阳性的肿瘤组织通过富含突变体的PCR重新分析，发现其中1份肿瘤存在K-ras突变。非霍奇金淋巴瘤患者的粪便和肿瘤组织K-ras突变均为阴性。DNA序列分析显示，粪便样本中鉴定出的每个K-ras突变在相应的肿瘤组织中都存在相同的碱基替换。结果表明，通过富含突变体的PCR可在结直肠癌患者粪便中高灵敏度和特异性地检测到携带K-ras突变的肿瘤细胞。由于该技术操作简便，可能适用于粪便样本中K-ras基因突变筛查。
