Neoplastic progression is characterized by clonal expansion of tumor cells associated with accumulation of mutational damage. The timing of mutation acquisition could be of value in distinguishing preneoplastic conditions from early and advanced cancer as well as characterizing tumor aggressiveness and treatment response. Using quantitative methods applied to microdissected cell clusters selected according to cytomorphologic features, we sought to demonstrate the feasibility and efficacy for determining the time and course of mutation accumulation in pancreatobiliary cytology specimens. In all, 40 pancreatic duct and 21 biliary brushing cytology specimens were retrieved from the cytology database. Xylene-resistant markings were placed on the slide underside and coverslips removed. Clusters of benign, atypical and malignant cells were manually microdissected and DNA extracted. Mutations (allelic imbalance) (loss of heterozygosity) were quantitatively determined for a broad panel of 15 markers (1p, 3p, 5q, 9p, 10q, 17p, 17q, 21q, 22q) as well as point mutation in K-ras-2 using PCR/capillary electrophoresis. Time course was based on earlier mutations having a higher proportion of mutant DNA for a particular marker. The descending frequency of detectable mutational involvement in pancreatic cytology was K-ras-2 point mutation (58%), 3p25-26 and 17q21 (35%), 5q23 (33%), 1p36 (28%), followed by the remaining molecular markers. The descending frequency of mutational content in bile duct cytology was 17p13, 1p36, 3p25-26, and 5q23 followed by remaining molecular markers. K-ras-2 point mutation was not seen in bile duct specimens. While there was overlap in the spectrum of mutational markers in pancreatic duct and biliary brushing cytology, the temporal profile was significantly different (Po0.001). Pancreatic and biliary neoplasia progression involves distinct subset of accumulated defined mutations. Determination of timing of the mutational damage in cytologic material could be incorporated in the work-up and help in making a more definitive diagnosis of malignancy in pancreatobiliary cytology specimens.

P63, a homologue of p53, was recently identified as a useful basal cell-specific marker. We compared the sensitivity and specificity of p63 with the widely used high-molecular-weight keratin 34betaE12 for the diagnosis of prostate carcinoma in needle biopsies. We selected 100 consecutive prostate carcinoma diagnosed by needle biopsies with an adequate number of cancerous glands on the slide. We chose 1 representative hematoxylin and eosin-stained slide from each case and gave it a Gleason score. The same paraffin block was retrieved for 34betaE12 and p63 stains. We compared staining patterns of 34betaE12 and p63 on both malignant glands and benign glands and recorded basal cell density (percentage of basal cells with positive staining in the benign glands). The cases were divided into 3 groups according to the Gleason score: 5 to 6 (31 cases), 7 (46 cases), and 8 to 10 (23 cases). In 20 cases, focal and patchy staining in a basal cell distribution in malignant glands (range, 1%-20%; mean, 6.6%) was demonstrated (19 by both stains and 1 by 34betaE12 only). In 1 case with a Gleason score of 9, the cancer cells, not the basal cells, were stained focally by p63 but not by 34betaE12. Higher-grade tumors demonstrated higher numbers of malignant glands with basal cell staining (1.65% for Gleason 7, 1.26% for Gleason 8-10, compared with 0.42% for Gleason 5-6). The overall specificity of the absence of basal cell staining in the malignant glands for 34betaE12 and p63 was 98.63% and 98.60%, respectively. In 17 cases, both stains revealed total absence of basal cell staining in some benign glands (range, 1%-10%; mean, 3.5%). The overall sensitivity in identifying basal cells in benign glands was 99.48% and 99.44% for 34beta12 and p63, respectively. Basal cell density was higher for 34betaE12 in comparison with p63 (92% vs. 87%). For diagnosing prostate carcinoma in the needle biopsies, p63 is as specific and sensitive Hospital as 34betaE12 and therefore can be used as a complementary basal cell-specific stain for 34betaE12 in difficult cases.