Goggins M, Hruban R H, Kern S E
Department of Pathology, Johns Hopkins Medical Institutions, Baltimore, Maryland 21205-2196, USA.
Am J Pathol. 2000 May;156(5):1767-71. doi: 10.1016/S0002-9440(10)65047-X.
Patients harboring germline BRCA2 mutations are at an increased risk of developing pancreatic cancer. We investigated the prevalence of biallelic inactivation of BRCA2 in the presumed precursors to invasive pancreatic ductal carcinomas, pancreatic intraepithelial neoplasia (PanIN). Surgical resection specimens from three patients with germline BRCA2 mutations who developed pancreatic ductal adenocarcinoma were studied. Fourteen PanINs were needle-microdissected from paraffin-embedded tissue. DNA was isolated from these microdissected tissues and amplified by primer-mediated pre-amplification. Loss of heterozygosity at the BRCA2 locus was determined by polymerase chain reaction amplification and cycle sequencing. The presence of the wild-type alleles was evaluated at the nucleotide positions of the germline BRCA2 mutations. The K-ras gene was sequenced at codon 12 and 13 to confirm the efficacy of microdissection. By histological evaluation the prevalence of PanINs in these patients was not notably elevated. Loss of the wild-type allele of BRCA2 was present in one high-grade PanIN (PanIN 3), but in none of 13 low-grade PanINs (PanIN 1). In contrast, K-ras mutations were detectable in 7 of the 14 PanINs. These results suggest that biallelic inactivation of the BRCA2 gene is a relatively late event in pancreatic tumorigenesis. In contrast to classical molecular progression models of tumorigenesis, the inactivation of the wild-type allele in a carrier of a recessive tumor susceptibility gene may not always be the first somatic event during the molecular evolution of a cancer. The necessity for earlier genetic alterations before biallelic inactivation of a recessive tumor susceptibility gene such as BRCA2 may explain why affected carriers have normal numbers of neoplastic precursor lesions, a relatively low phenotypic penetrance, and late age of onset of pancreatic and other cancers.
携带种系BRCA2突变的患者患胰腺癌的风险增加。我们研究了侵袭性胰腺导管癌的假定前体——胰腺上皮内瘤变（PanIN）中BRCA2双等位基因失活的发生率。对三名发生胰腺导管腺癌的种系BRCA2突变患者的手术切除标本进行了研究。从石蜡包埋组织中针吸微切割出14个PanIN。从这些微切割组织中分离DNA，并通过引物介导的预扩增进行扩增。通过聚合酶链反应扩增和循环测序确定BRCA2基因座的杂合性缺失。在种系BRCA2突变的核苷酸位置评估野生型等位基因的存在情况。对K-ras基因的第12和13密码子进行测序，以确认微切割的效果。通过组织学评估，这些患者中PanIN的发生率没有明显升高。在一个高级别PanIN（PanIN 3）中存在BRCA2野生型等位基因缺失，但在13个低级别PanIN（PanIN 1）中均未发现。相比之下，在14个PanIN中有7个可检测到K-ras突变。这些结果表明，BRCA2基因的双等位基因失活在胰腺肿瘤发生过程中是一个相对较晚的事件。与经典的肿瘤发生分子进展模型不同，隐性肿瘤易感基因携带者中野生型等位基因的失活可能并不总是癌症分子进化过程中的第一个体细胞事件。在隐性肿瘤易感基因如BRCA2双等位基因失活之前需要更早的基因改变，这可能解释了为什么受影响的携带者肿瘤前体病变数量正常、表型外显率相对较低以及胰腺癌和其他癌症发病较晚。
