Inhibitory effects on tumor invasion and metastasis induced by transfection of anti-sense E1AF,an ets-oncogene family transcription factor.

• Project/Area Number: 08672063
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Morphological basic dentistry
• Research Institution: HOKKAIDO UNIVERSITY
• Principal Investigator: SHINDOH Masanobu HOKKAAIDO UNIVERSITY SCHOOL OF DENTISTRY,DEPARTMENT OF ORAL PATHOLOGY ASSOCIATE PROFESSOR, 歯学部, 助教授 (20162802)
• Co-Investigator(Kenkyū-buntansha): FUJINAGA Kei SAPPORP MEDICAL UNIVERSITY SCHOOL OF MEDICINE DEPARTMENT OF MOLECULAR BIOLOGY,CA, 医学部・附属がん研究所, 名誉教授 (10045338) ; TOTSUKA Yasunori HOKKAAIDO UNIVERSITY SCHOOL OF DENTISTRY,DEPARTMENT OF ORAL SURGERY,PROFESSOR, 歯学部, 教授 (00109456) ; CHIBA Itsuo HOKKAAIDO UNIVERSITY SCHOOL OF DENTISTRY,DEPARTMENT OF ORAL SURGERY,ASSISTANT PR, 歯学部, 助手 (50250460)
• Project Period (FY): 1996 – 1997
• Project Status: Completed (Fiscal Year 1997)
• Budget Amount: ¥2,200,000 (Direct Cost: ¥2,200,000); Fiscal Year 1997: ¥900,000 (Direct Cost: ¥900,000); Fiscal Year 1996: ¥1,300,000 (Direct Cost: ¥1,300,000)
• Keywords: E1AF / ets-oncogene / antisense / tumor invasion / p21^<waf1 / cip1> / ets-omcgene / timor innasion / cip1> / EIAF / Matrix metalloproteinase / Oral Squamows cell carvinoma / imrasion / anti sense vector

Research Abstract

E1AF is a newly identified human ets-family transcription factor. We have reported that E1AF can up-requlate transcription of matrix metalloproteinase (MMP) genes and confers invasive phenotype on human cancer cells. HSC3 is an oral squamous cell carcinoma-derived cell line, and it manifests high levels of E1AF,and MMP-1 and -9 gene expression that are associated with invasive potential. We reconstructed an E1AF antisense expression vector, transfected HSC3 cells with the vector and obtained HSC3AS cells which express E1AF antisense RNA.HSC3AS showed decreasing m, -3 and -9. moreover, HSC3AS showed lower invasive potential in in vitro three-dimensional raft culture and in vivo implantation into nude mice. These results imply that transfection of antisense E1AF inhibits tumor invasion by down-regulating MMP genes.
Thus, E1AF is thought to be highly correlated with malignant potentials of cancer cells, however, little is known about E1AF expression and cancer cell malignancies in in vivo tumors. We examined 27 oral SCC speciment using RT-PCR,Southern blot hybtidization and in situ hybridization (ISH) and compared to the clinico-pathological parameters. Among the 27 patients, E1AF was detected in 15 cases. E1AF mRAN was detected in 13 of 17 invasive SCC_s, whereas the majority of SCC_s not expressing E1AF showed an expansive growth pattern. Increased prevalence of E1AF-positive oral SCC was observed in cases with nodal metastasis. These results indicate that E1AF may be involved in cancer cell malignancies through its ability to promote invasive potential
p21^<Waf1/Cip1> is one of the key requlatory proteins in cell cycle, terminal differentiation and apoptosis. its promoter was shown to be transactivated by the wild-type p53 protein as well as in a p53-independent manner. We demonstrate that E1AF,an ets-related transcription factor, activates the human p21^<Waf1/Cip1> promoter by interacting with the ets-binding sites located close to the two previously identified p53-responsive elements. Northern blot analysis revealed that p21^<Waf1/Cip1> and E1AF were correlatively uprequlated in response to cisplatin treatment in SiHa cells. Transient expression assays demonstrated that E1AF can activate the p21^<Waf1/Cip1> promoter-driven luciferase reporter gene in SiHa cells. The p21^<Waf1/Cip1> promoter activity was also increased in p53-null Saos2 osteosarcoma cells, but was markedly reduced when the etsbinding sites were deleted. These results indicate that E1AF positively requlates transcription from the p21^<Waf1/Cip1> promoter in response to genotoxic stresses.

Research Products

• [Publications] Hida, K., Shindoh, M. et al.: "Antisense E1AF Transfection Restrains Oral Cancer Invasion by Reducing Matrix Metalloproteinase Activities." American Journal of Pathology. 150・6. 2125-2132 (1997)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] Hida, K., Shindoh, M. et al.: "Expression of E1AF,an ets-Family Transcription Factor, is Correlated with the Invasive Phenotyne of Oral Squarnvao cell Carcinomas." European Journal of Cancer (Part B), Oral Oncol.33・6. 426-430 (1997)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] Funaoka, K., Shindoh, M. et al.: "Activation of the p21^<wafl/cipl> Promoter by the ets Oncogene Family Transcriptton Factor E1AF." Biochemical and Biophysical Research Communications. 236. 79-82 (1997)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] Hida, K., Shindoh, M., Yoshida, K., Yasuda, M., Hanzawa, M., Funaoka, K., Kohgo, T., Amemiya, A., Totsuka, Y., Yoshida, K., Fujinaga, K: "Antisense E1AF transfection restrains oral cancer invasion by reducing matrix metalloprotease activities." Am.J.Pathol. 150. 2125-2132 (1997)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Hida, K., Shindoh, M., Yoshida, K., Kohgo, T., Fujinaga, K., Totsuka, Y: "Expression of E1AF,an ets-family transcription factor, is correlated with the invasive phenotype of oral squamous cell carcinomas." Oral Oncol. 33. 426-430 (1997)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Funaoka, K., Shindoh, M., Yoshida, K., Nishikata, S., Totsuka, Y., Fujinaga, K.: "Activation of the p21Waf1/Cip1 promoter by the ets-oncogene family transcription factor E1AF." Biochem. Biophys. Res.Commun.236. 79-82 (1997)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Hida,K., Shindoh,M.et al.: "Antisenes E1AF Transfectlon Restrains Oral Cancer/nrasion by Reducing Matrix Metalloproteinase Activities" American Journal of Pathology. 150・6. 2125-2132 (1997)
• [Publications] Hida,K., Shindoh,M.et al.: "Expression of E1AF,an ets-Family Transcription Factor,is Correlated with the invasive Phenotyre of Oral Squamoos lell Carcinomas." European Journal of Cancer(Rart B),Oral Omol.33・6. 426-430 (1997)
• [Publications] Funaoka,K., Shindoh,M.et al.: "Activation of the p21^<wafl/cip1> Promoter by the ets Oncogene Family Transoription Factor E1AF" Biochemical and Biophysical Research Communications. 236. 79-82 (1997)
• [Publications] Shindoh, M., Higashino, F. et al.: "Correlated expression of matrix netalloproteinases and ets-family transcription on factor ELA-F in imvasive oral-squamous-ull-carinoma-usived cell lins." Am. J. Pathol.148 (3). 693-700 (1996)
• [Publications] Hida, k., Shindoh, M. et al.: "Antisense EIAF transfectioin restrains cral cancer cell in vasion by reducing MMPs acttrities." Am. J. Pathol.151 (in press). (1997)