Larson A A, Kern S, Sommers R L, Yokota J, Cavenee W K, Hampton G M
Ludwig Institute for Cancer Research, University of California at San Diego, La Jolla, 92093-0660, USA.
Cancer Res. 1996 Mar 15;56(6):1426-31.
Infection of epithelial cells with human papillomavirus is an important early event in the development of cervical dysplasia. However, progression to overt malignancy appears dependent upon further genetic and/or epigenetic events. We have recently developed methodologies for the simultaneous analysis of loss of heterozygosity (LOH) at multiple PCR-based microsatellite loci using semiautomated fluorescent DNA sequencing technology to determine the locations of tumor suppressor genes which are inactivated during tumor progression. While examining 30 microsatellite loci for LOH on chromosomes 3p, 4, and 11q, we detected novel tumor-specific alleles indicative of microsatellite instability (MI). The methodology allowed rapid and accurate comparison of over 3000 genotypes from 89 primary tumors and 10 cervical carcinoma-derived cell lines and showed that five tumors (5.6%) and one human papillomavirus-negative cell line, C33A, had genetic features consistent with a replication error (RER+) phenotype as defined by MI at two or more loci. In each of the RER+ tumors, LOH was also observed at one or more loci on each of the three chromosomes examined. These findings suggest that defects in DNA repair-associated genes are rarely acquired and do not supersede allelic loss during cervical carcinogenesis. In addition, the semiautomated multiplex approach has proven unequivocal in the detection and interpretation of MI and should greatly accelerate the rapidity and accuracy of analysis of such defects in tumors. Moreover, the number of loci that can be relatively easily examined in this way will also allow a detailed statistical consideration of the importance of such events.
人乳头瘤病毒感染上皮细胞是宫颈发育异常发展过程中的一个重要早期事件。然而，进展为明显的恶性肿瘤似乎依赖于进一步的基因和/或表观遗传事件。我们最近开发了一些方法，利用半自动荧光DNA测序技术同时分析多个基于PCR的微卫星位点的杂合性缺失（LOH），以确定在肿瘤进展过程中失活的肿瘤抑制基因的位置。在检查3号染色体短臂、4号和11号染色体长臂上的30个微卫星位点的LOH时，我们检测到了指示微卫星不稳定性（MI）的新的肿瘤特异性等位基因。该方法能够快速、准确地比较来自89个原发性肿瘤和10个宫颈癌衍生细胞系的3000多个基因型，并显示5个肿瘤（5.6%）和1个人乳头瘤病毒阴性细胞系C33A具有与复制错误（RER+）表型一致的遗传特征，该表型由两个或更多位点的MI定义。在每个RER+肿瘤中，在所检查的三条染色体的每一条上的一个或多个位点也观察到了LOH。这些发现表明，与DNA修复相关的基因缺陷很少出现，并且在宫颈癌发生过程中不会取代等位基因缺失。此外，半自动多重分析方法已被证明在检测和解释MI方面是明确无误的，并且应该会大大加快对肿瘤中此类缺陷分析的速度和准确性。此外，以这种方式相对容易检查的位点数量也将允许对这些事件的重要性进行详细的统计学考量。
