Effect of connective tissue growth factor on hypoxia-inducible factor 1α degradation and tumor angiogenesis

Cheng Chi Chang, Ming Tsai Lin, Been Ren Lin, Yung Ming Jeng, Szu Ta Chen, Chia Yu Chu, Robert J. Chen, King Jen Chang, Pan Chyr Yang, Min Liang Kuo

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Abstract

Background: Connective tissue growth factor (CTGF) inhibits the metastatic activity of human lung cancer cells in a mouse model; however, the mechanism of this modulation is unclear. We investigated the role of angiogenesis in this process. Methods: CL1-5 and A549 human lung adenocarcinoma cells were stably transfected with vectors containing CTGF or hypoxia-inducible factor (HIF) 1α or with vector controls. Transfected cells were injected into nude mice (n = 10 per group), and tumor growth, metastasis, and mouse survival were measured. Excised xenograft tumors and primary human lung adenocarcinomas (n = 24) were subjected to immunohistochemistry with antibodies to the endothelial cell marker CD31 and to CTGF. Expression of HIF-1α and vascular endothelial growth factor (VEGF) A was assessed in vitro by using reporter gene assays. Cells were transiently transfected with HIF-1α mutant and antisense arrest-defective 1 protein (ARD-1), and HIF-1α acetylation was assayed by immunoprecipitation. All statistical tests were two-sided. Results: Xenograft tumors derived from CTGF transfectants grew more slowly than those from control-transfected cells and had reduced expression of HIF-1α and VEGF-A, vascularization (as assessed by CD31 expression), and metastasis (all P <.001). Xenograft tumors derived from CTGF-overexpressing cells that were transfected with HIF-1α had higher VEGF-A expression than CTGF-overexpressing xenografts. Mice with CTGF/HIF-1α xenografts had lower survival than mice carrying CTGF-overexpressing xenografts (CL1-5/Neo, mean = 69.6 days, 95% confidence interval [CI] = 53.9 to 85.3 days versus CL1-5/ CTGF, mean = 102.1 days, 95% CI = 92.1 to 112.1 days; P = .001, CL1-5/ CTGF, mean = 102.1 days, 95% CI = 92.1 to 112.1 days versus CL1-5/CTGF/ HIF-1α, mean = 81.7 days, 95% CI = 66.5 to 96.9 days; P = .011, CL1-5/Neo, mean = 69.6 days, 95% CI = 53.9 to 85.3 days versus CL1-5/ CTGF/HIF-1α, mean = 81.7 days, 95% CI = 66.5 to 96.9 days; P = .122). Tumors of patients with the same disease stage but with high CTGF protein expression had reduced microvessel density compared with tumors with low expression. Transfection with antisense-ARD1 decreased the level of acetylated HIF-1α and restored HIF-1α and VEGF-A expression in CTGF-overexpressing cells. Conclusion: CTGF inhibition of metastasis involves the inhibition of VEGF-A-dependent angiogenesis, possibly by promoting HIF-1α protein degradation.

Original languageEnglish
Pages (from-to)984-995
Number of pages12
JournalJournal of the National Cancer Institute
Volume98
Issue number14
DOIs
Publication statusPublished - Jul 19 2006
Externally publishedYes

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

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  • Cite this

    Chang, C. C., Lin, M. T., Lin, B. R., Jeng, Y. M., Chen, S. T., Chu, C. Y., Chen, R. J., Chang, K. J., Yang, P. C., & Kuo, M. L. (2006). Effect of connective tissue growth factor on hypoxia-inducible factor 1α degradation and tumor angiogenesis. Journal of the National Cancer Institute, 98(14), 984-995. https://doi.org/10.1093/jnci/djj242