Loss of heterozygosity (LOH) was detected in morphologically normal lobules adjacent to breast cancers. The most frequent aberration was at chromosome 3p22-25; of ten cases with this LOH in the carcinoma, six displayed the same LOH in adjacent normal lobules. This suggests that in a subset of sporadic breast cancers, a tumor suppresser gene at 3p22-25 may be important in initiation or early progression of tumorigenesis. Among sixteen breast cancers with LOH at 17p13.1 and five breast cancers with LOH at 11p15.5, one case each displayed the same LOH in adjacent normal lobules. Thus the molecular heterogeneity that characterizes invasive breast cancers may occur at the earliest detectable stages of progression.The mature breast contains lobules, clusters of closed glandular spaces that produce milk during lactation. These lobules are connected to the nipple-areolar complex by a system of branching ducts that are surrounded by varying amounts of fat and connective tissue. Breast cancer is thought to develop within a terminal ductal-lobular unit (TDLU), which includes the lobule and its most proximal ducts (1).Breast cancer evolves by clonal selection of cells that acquire multiple molecular changes. One model suggests that breast cancer, like colon cancer (2), develops through a defined progression of morphologically distinguishable stages beginning with benign hyperplasia, which progresses to atypical hyperplasia, then to in situ carcinoma, and finally to invasive cancer (1). This sequential progression may not be the only way that breast cancers develop, however. Many small invasive cancers do not have atypical components, which suggests that they may have developed directly from morphologically normal epithelium. If this were true, one might expect to find evidence of a "field effect" in which at least some of the genetic aberrations found in invasive cancers are also present in the morphologically normal epithelium.To test this hypothesis, we carefully microdissected hematoxylin-eosin-stained sections of breast cancers so as to isolate morphologically discrete regions (Fig. 1A). DNA was prepared from malignant areas of the section and from adjacent normal TDLUs. As a control for each case, DNA was also prepared from normal breast skin (usually from a separate section) that had been similarly microdissected.We studied LOH at chromosome 3p24, 11p15.5, 13q13, and 17p13.1 because these loci show LOH in a high percentage (ϳ30 to 60%) of invasive ductal breast cancers (3, 4). For the carcinomatous regions, the frequency of LOH at 3p24 (48%) and 11p15.5 (29%) was similar to that previously reported (4). The frequency of LOH in the invasive components was higher than the literature values for 13q13 (64% here versus ϳ40%) and for 17p13.1 (80% here versus ϳ60%). These discrepancies may be due to random variation because our sample size was small.In 8 of 30 cases we detected LOH in the adjacent morphologically normal TDLUs (Table 1). In all eight cases, the same allele was missing in the adjacent carcinoma (Fig. 2, A an...

Previously we found differences in the distribution of the individual human papillomavirus types in cervical cancers and high-grade squamous intraepithelial lesions. This suggested that there were differences in risk for progression of high-grade squamous intraepithelial lesions that were related to human papillomavirus type within the category of oncogenic genotypes. In this work, we add additional cases including low-grade squamous intraepithelial lesions. ThinPrep s samples from 282 squamous intraepithelial lesions and invasive cervical cancers were categorized morphologically by consensus interpretation and genotyped for 27 individual human papillomavirus types by polymerase chain reaction-based reverse line blot analysis using PGMY09/ PGMY11 consensus primers for the L1 open reading frame. The 27 human papillomavirus types were divided into three categories: high risk 16, 18, 31, 45; intermediate risk 33, 35, 39, 51, 52, 56, 58, 59, 68, 73, 82, 83; and low risk: 6, 11, 26, 40, 42, 53, 54, 55, 57, 66, and 84. Of the 282 cases of cancer and squamous intraepithelial lesions, 95.7% were positive for one or more of 27 human papillomavirus types and 38.7% had two or more genotypes. Three major categories of squamous intraepithelial lesions were identified based upon the combination of consensus diagnosis and human papillomavirus category: (1) high-grade squamous intraepithelial lesions associated with high-risk human papillomavirus types that appear to be at increased risk for progression to carcinoma; (2) squamous intraepithelial lesions (typically low-grade intraepithelial lesions and high-grade lesions consistent with moderate dysplasia) associated with intermediate risk human papillomavirus types with limited or indeterminate risk for progression; (3) low-grade squamous intraepithelial lesions associated with low-risk human papillomavirus types with little or no risk for progression. Only a subset of human papillomavirus genotypes commonly considered to be oncogenic were closely associated with invasive cervical cancer and high-grade squamous intraepithelial lesions classed as severe dysplasia. Other oncogenic types were closely associated with high-grade squamous intraepithelial lesions of moderate dysplasia and lowgrade squamous intraepithelial lesions. This suggests that risk for progression to invasion in squamous intraepithelial lesions is closely related to human papillomavirus genotype. Knowledge of the associated human papillomavirus type in women with morphologic squamous intraepithelial lesions may help to clarify risk for progression. Keywords: cervical carcinoma; human papillomavirus; low-grade squamous intraepithelial lesion; high-grade squamous intraepithelial lesion; HPV genotyping; cervical carcinogenesis Traditionally, cervical carcinogenesis was thought to evolve through a series of increasingly abnormal intraepithelial patterns (ie, mild-moderate-severe dysplasia) followed, in a minority of cases, by acquisition of the ability to invade and metastasize. The identification of human papillom...

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Microsatellite instability (MSI) is a hallmark of the DNA replication error phenotype, due to the inactivation of mismatch repair genes. MSI has been implicated in colon and many other gastrointestinal cancers. MSI usually can be analyzed by PCR amplification of microsatellite markers followed by electrophoresis and detected using autoradiography or fluorescence techniques. We report here a novel method for high-throughput detection of MSI using denaturing high performance liquid chromatography (DHPLC). Amplification of two mononucleotide markers (BAT25 and BAT26) by polymerase chain reaction (PCR) is followed by DHPLC analysis to display alteration in the length of repetitive sequences. These two markers were tested in 84 colorectal cancer samples confirmed to be 44 MSI-H and 40 MSI-L or MSS, previously defined by multiple microsatellite markers and/or by immunohistochemical analyses of MLH1 and MSH2 proteins. Among 44 MSI-H samples, sequence variations in BAT26 and BAT25 were detected in 44 (100%) and 43 (98%), respectively, while no sequence variation in the two markers was detected in 40 MSI-L or MSS samples. A total of 96 gastric cancers and their matched normal tissues were then analyzed for MSI-H using this method. Sequence variations in BAT26 and BAT25 were detected in nine (9.4%) samples. Seven of the nine cases were shown unstable at both BAT26 and BAT25; one each was unstable at BAT26 or BAT25. These results were confirmed by autoradiography analyses. Together, our results demonstrate high sensitivity and specificity of DHPLC in the analysis of sequence variations in BAT25 and BAT26 to determine MSI status. This simple, efficient, and high-throughput approach will facilitate analysis of MSI in large sample sets of any cancers.