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An immunohistochemical differential staining of cancerous cells with anti-cytidine antibody after denaturation of nuclear DNA by acid hydrolysis with 2N HCl at 30 degree C for 20 min (DNA-instability test) has been used as a marker of malignancy. The test was applied to bioptic tissues of human gastric polyp assessed histopathologically as foveolar hyperplastic polyp (13 cases), mild (58 cases), moderate (86 cases), and severe (20 cases) dysplasia, and adenocarcinomas (14 cases). The serial sections of the same tissues were also subjected to immunohistochemical staining for Ki67, p53, DNA-fragmentation factor (DFF45), and basic fibroblast growth factor (bFGF). The DNA-instability test was positive in 14 (100%) adenocarcinoma cases, 20 (100%) severe dysplasia cases, 52 (60.5%) moderate dysplasia cases, and 12 (20.7%) mild dysplasia cases, indicating malignancy. All foveolar hyperplastic polyps were negative to the DNA-instability testing. Furthermore, the percentage of glands positive in the DNA-instability test steadily increased in going from mild (10%), to moderate (40%), to severe (100%) dysplasia, and adenocarcinoma (100%). All other biological markers tested in the present study showed significantly higher values in the adenoma glands, being positive to DNA-instability testing, irrespective of the dysplasia grade, as compared to those in the adenoma glands that were negative to DNA-instability testing. Furthermore, the former values were comparable to those in adenocarcinoma. These results indicate that cancer cell clones are already present at the adenoma stages showing a positive DNA-instability test, enhanced proliferative activity, p53 mutation, induction of DFF45 and bFGF. These factors allow cancer cell proliferation, producing heterogeneous subclones due to DNA-instability, enhancing their survival by escaping apoptosis, and providing abundant nutrients during the early-stage progression of gastric cancer. Based on these findings, we herein propose the concept of "procancer" (as opposed to "pre-cancer") as being a unique stage during the course of carcinogenesis and cancer progression. We designate the term to cancer clones at the very early stages of malignant progression that do not show distinguishable morphological atypia but do show positive DNA-instability testing and positive staining for various biomarkers such as Ki67, p53, DFF45, and bFGF. We also define the abnormal positive staining of these biomarkers, including the DNA-instability test as "functional atypia", compared to the ordinary morphological atypia.

Detection of cancer clones in human gastric adenoma by increased DNA-instability and other biomarkers