van Es J M, Polak M M, van den Berg F M, Ramsoekh T B, Craanen M E, Hruban R H, Offerhaus G J
Department of Pathology, Academic Medical Centre, University of Amsterdam, The Netherlands.
J Clin Pathol. 1995 Mar;48(3):218-22. doi: 10.1136/jcp.48.3.218.
To determine the potential efficiency of molecular markers specific for neoplastic change--mutations of the K-ras oncogene and the p53 tumour suppressor gene--in diagnosing pancreatic carcinoma.
Archival cytology samples obtained from 17 patients with established pancreatic carcinoma were assayed for alterations in K-ras and p53. To detect changes in p53 expression, immunocytochemistry with polyclonal antibody CM1 was performed on the archival cytology slides after destaining. Mutations in K-ras codon 12 were then analysed on the scrapings of the same slides using mutant enriched polymerase chain reaction (PCR) amplification and restriction fragment length polymorphism analysis with allele specific oligonucleotide hybridisation for confirmation and characterisation.
False negative results were recorded for five of the cytology slides when compared with p53 immunostaining of the surgical resection specimen. These five cases had been stained previously with Giemsa which interacts adversely with the immunostaining in contrast to the Papanicolaou procedure. The K-ras codon 12 mutations followed the well established distribution frequency and spectrum for pancreatic cancer and corresponded with the findings in the resection specimens in all cases. Two scrapings yielded insufficient DNA for PCR. Importantly, for two cases with an inconclusive cytology diagnosis on routine light microscopy, the diagnosis was confirmed by one of the molecular markers. The application of the molecular markers increased the diagnostic accuracy of cytology in this small study from 76 to 89%.
The study indicates that assessment of alterations in the K-ras and p53 genes may be a valuable adjunct to diagnostic cytopathology of the head region of the pancreas, although there are some difficulties which will have to be overcome.
确定用于诊断胰腺癌的肿瘤性改变特异性分子标志物——K-ras癌基因和p53肿瘤抑制基因的突变——的潜在效率。
对17例确诊为胰腺癌患者的存档细胞学样本进行K-ras和p53改变的检测。为检测p53表达的变化，在对存档细胞学玻片进行脱色后，用多克隆抗体CM1进行免疫细胞化学检测。然后使用突变富集聚合酶链反应（PCR）扩增和等位基因特异性寡核苷酸杂交的限制性片段长度多态性分析，对同一张玻片的刮取物进行K-ras密码子12突变分析，以进行确认和特征分析。
与手术切除标本的p53免疫染色相比，5张细胞学玻片出现假阴性结果。这5例之前用吉姆萨染色，与巴氏染色法相比，吉姆萨染色与免疫染色存在不良相互作用。K-ras密码子12突变遵循胰腺癌已确定的分布频率和谱，且在所有病例中均与切除标本的结果一致。两次刮取得到的DNA不足以进行PCR。重要的是，对于2例在常规光学显微镜下细胞学诊断不明确的病例，其中一种分子标志物确诊了诊断。在这项小型研究中，分子标志物的应用将细胞学诊断的准确性从76%提高到了89%。
该研究表明，评估K-ras和p53基因的改变可能是胰腺头部诊断性细胞病理学的一项有价值的辅助手段，尽管仍有一些困难需要克服。
