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-67- Carcinogenesis/Tomor/Oncogene(2) 9/4 Room-D
OVEREXPRESSION AND GENE MUTATION OF P53 IN SQUAMOUS CELL CARCINOMAS OF THE SKIN.
Mayumi Matsuta1, Gen Kosegawa1, Toshihide Akasaka1, Morimasa Matsuta2.
1Department of Dermatology, 2Department of Obstetrics & Gynecology, Iwate Medical University School of Medicine, Morioka, Japan.
We investigated the expression of p53 protein in 31 cases of squamous cell carcinoma of the skin, immunohistochemocally. Sbsequently, the p53 gene mutation of exons 5 to 8 was analyzed by PCR-FSSCP. Twenty nine cases overexpressed p53 protein, and 42%(13/31) showed mutations within the exons 5-8 of the p53 gene; the mutation of one exon was detected in 10 cases of these cases, and of 2 or 3 of exons in three cases. However, no mutation was detected in eighteen cases, but 2 cases of these cases were negative immunohistochemically. The data suggested that unsequenced p53 gene mutations might occur in SCC of the skin or have wild type p53 protein that accumulates for other reasons.
-68- Carcinogenesis/Tomor/Oncogene(2) 9/4 Room-D
ANALYSIS OF p53 GENE PRODUCT AND PCNA IN SKIN CANCER.
Masako Udono, Yoko Abe, Ichiro Katayama.
Department of Dermatology, Nagasaki University School of Medicine, Nagasaki, Japan.
It has been suggested that mutation of p53 gene is involved in transformation of cells in various types of cancers and PCNA is a functional marker of proliferation in cancer. The relationship between mutational changes of p53 gene and the expression of PCNA in cells dreived from 10 cases of each of actinic keratosis, Bowen's disease, SCC, and seborrheic keratosis, was investigated by double hist-staining with specific monoclonal antibodies. Further, PCR-SSCP analysis was performed for detection of mutation of p53 gene in these cells. Overexpression of both p53 gene product and PCNA was observed in all cases except seborrhic keratosis and only one case each of actinic keratosis and SCC was recognized as abnormal p53 with band-shifting in exon 5. Although the functional relationship between p53 and PCNA in transfromation and cell cycle regulation of skin cancer is not well understood, our data suggests that the PCNA expression appears to be accompanied by overexpression of wild type but not mutational p53 gene product.
-69- Carcinogenesis/Tomor/Oncogene(2) 9/4 Room-D
ASSOCIATION OF OVEREXPRESSION OF p53 PROTEIN with THE PROLIFERATIVE ACTIVITY IN SQUAMOUS CELL CARCINOMA OF THE SKIN.
Takahiro Shimizu, Masahiko Muto, Hiroko Furumoto, Junji Nakano, Chidori Asagami.
Department of Dermatology, Yamaguchi University School of Medicine, Yamaguchi, Japan.
Although overexpression of p53 protein has been demonstrated in a variety of skin cancers, its significance is still unclear. Ki-67 antigen is generally accepted as a reliable marker of proliferating cells. Sixteen cases of well-differentiated squamous cell carcinoma (SCC) were immunohistochemically analyzed for the expression of p53 protein and Ki-67 antigen. p53 protein was detected in 12 out of the 16 cases. Cases with higher p53 indices (the percentage of p53 protein positive tumor cells) tended to display higher Ki-67 indices (the percentage of Ki-67 antigen positive tumor cells) and the correlation was statistically significant. The present study suggests that the presence of detectable p53 protein confers a proliferative advantage to SCC of the skin.
-70- Carcinogenesis/Tomor/Oncogene(2) 9/4 Room-D
ABERRATIONS OF p53 GENE AND RAS GENES IN SOLAR KERATOSIS.
Masahito Taguchi1, Tetsuya Tsuchida1, Shigeo Ikeda1, Shaw Watanabe2, Takao Sekiya3.
1Department of Dermatology, Saitama Medical School, Saitama, 2Department of Nutrition and Epidemiology, Tokyo University of Agriculture, Tokyo, 3Oncogene Division, National Cancer Center Research Institute, Tokyo, Japan.
In order to clarify the multiple step progress from solar keratosis (SKs) to human epidermal skin cancer, we used PCR-SSCP analysis to investigate aberrations of the p53 gene (exons 2 to 11) and the ras genes (exons 1 and 2) in SKs and squamous cell carcinomas (SCCs). In a series of Japanese patients, 8 out of 27 (30%) cases of SKs and 2 out of 5 (40%) SCCs showed structural abnormalities in the p53 gene. One out of 27 (3.7%) cases of SKs showed a point mutation in the H-ras gene. In our cases, no mutations of the ras genes could be detected in SCCs. Mutation of the ras genes was not detected from SKs and SCCs where mutations were shown to occur in the p53 gene. We concluded that the aberration of the p53 gene and the ras genes are due to independent processes of ultraviolet ray carcinogenesis in the course of carcinogenic change to SCC.
-71- Carcinogenesis/Tomor/Oncogene(2) 9/4 Room-D
EXPRESSIONS OF MMP-1, P53, METALLOTHIONEIN AND PCNA IN SOLAR KERATOSIS.
Reiko Tsukifuji, Hiroshi Shinkai.
Department of Dermatology, Chiba University School of Medicine, Chiba, Japan.
We studied expressions for MMP-1mRNA, p53 protein, metallothionein(MT) and PCNA(proliferating-cell nuclear antigen) in solar keratosis (SK) to understand the progression toward invasive tumors. Transcripts for MMP-1 were examined by in situ hybridization using digoxigenin-labeled RNA probes and immunostaining for p53, MT and PCNA was done using the streptavidin-biotin method. MMP-1mRNA was detected in 4 of 14 cases, and p53, MT and PCNA were overexpressed in 7, 4 and 5 of 14 cases, respectively. SKs associated with SCC demonstrated more expressions for these four markers. The results suggest that SK demonstrates a diversity in the expressions for MMP-1, p53, MT and PCNA in the development of SCC.
-72- Carcinogenesis/Tomor/Oncogene(2) 9/4 Room-D
p53 GENE MUTATION OF SKIN TUMORS DETECTED BY p53 FUNCTIONAL ASSAY.
Toshinori Bito1, Masato Ueda1, Masamitsu Ichihashi1, Mitsuhiro Tada2, Hideyuki Saya3.
1Department of Dermatology, Kobe University School of Medicine, Kobe, 2Department of Neurosurgery, Hokkaido University School of Medicine, Hokkaido, 3Department of Oncology, Kumamoto University School of Medecine, Kumamoto, Japan.
A simple p53 functional assay using yeast was used to detect the p53 gene mutation in skin tumors. This is the assay for p53 mutation in which human p53 expressed in Saccharomyces cereviae activates transcription of the ADE2 gene, cloned to downstream of a CYC1 minimal promoter containing p53 binding site. Consequently, yeast colonies containing wild-type p53 become white and colonies containing mutant p53 become red. Twenty skin tumors (5 actnic keratosis, 1 keratoacanthoma, 6 Bowen's disease, 4 basal cell carcinoma and 4 squamous cell carcinoma) and 2 normal skin were examined in this assay. PCR using synthesized cDNA was performed with primers for p53 gene. An unpurified human p53 RT-PCR product is cloned into a constitutive yeast expression vector by homologous recombination in vivo after cotransformation of PCR product and vector into yeast. Same tumor tissues were also immunohistochemically stained with p53 antibody. Seventeen of 22 samples formed red colonies, and 13 of these showed overexpression of p53 protein. Four of 5 samples which formed white colonies showed no protein expression. One actinic keratosis formed white colonies, while protein overexpression was seen. These data indicate that the protein expression in tumor tissue is relatively reliable indicator of genetic alteration of p53.
-73- Carcinogenesis/Tomor/Oncogene(2) 9/4 Room-D
EXPRESSION OF p53 PROTEIN IN DLE.
Hijiri Kato1, Suzuo Hashizume1, Masaru Ito1, Takako Baba1, Eiichi Shibayama2, Toshihumi Takakuwa2, Masako Mizoguchi1.
1Department of Dermatology, 2Department of Pathology, St. Marianna University, School of Medicine, Kanagawa, Japan.
EXPRESSION OF p53 PROTEIN IN DLE. H. Kato*1, S. Hashizume1, M. Ito1, T. Baba1, E. Shibayama2, T. Takakuwa2, M. Mizoguchi1. 1Department of Dermatology and 2Department of Pathology, St. Marianna University, School of Medicine, Kanagawa, Japan. Mutation of p53 gene is a common genetic alteration in human malignancies. Overexpression of p53 protein in skin cancers or precancerosis has been reported, but the expression of p53 protein in DLE, which is known as an inflammatory auto immune disease, has never been reported. We have investigated the expression of p53 and bcl-2 protein in DLE in comparison with SLE and SCC by immunohistochemical staining. Immunohistochemical staining for p53 protein was observed in 8/10 (80%) DLEs, 3/10 (30%) SLEs and 8/10 (80%) SCCs. And bcl-2 protein was observed 8/10 (80%) DLEs, 10/10 (100%) SLEs and 0/10 (0%) SCCs. We examined for p53 mutation using polymerase chain reaction and single-strand conformation polymorphism analysis (PCR-SSCP) techniques in 8 cases of DLE which had p53 protein detected by immunohistochemical staining.