Supplementary Figure 2 from Hypoxia-Independent Gene Expression Mediated by SOX9 Promotes Aggressive Pancreatic Tumor Biology

Camaj, Peter; Jäckel, Carsten; Krebs, Stefan; DeToni, Enrico N.; Blum, Helmut; Jauch, Karl-Walter; Nelson, Peter J.; Bruns, Christiane J.

doi:10.1158/1541-7786.22509117.v1

Supplementary Figure 2 from Hypoxia-Independent Gene Expression Mediated by SOX9 Promotes Aggressive Pancreatic Tumor Biology

journal contribution

posted on 2023-04-03, 16:00 authored by Peter Camaj, Carsten Jäckel, Stefan Krebs, Enrico N. DeToni, Helmut Blum, Karl-Walter Jauch, Peter J. Nelson, Christiane J. Bruns

PDF file - 70K, S2. A) Differential methylation of the Sox9-promoter in NMET and HMET cells. B) Methylation and hypoxia. NMET and HMET cells were cultivated under hypoxia or normoxia, respectively. (C) Effect of methylation inhibition on VEGF expression. Relative transcription of VEGF in LMET and HMET cells is demonstrated. D) Effect of methylation inhibition on Sox9 expression.

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ARTICLE ABSTRACT

Pancreatic cancer aggressiveness is characterized by its high capacity for local invasion, ability to promote angiogenesis, and potential to metastasize. Hypoxia is known to represent a crucial step in the development of aggressive malignant features of many human cancers. However, micrometastatic tumors are not typically subjected to hypoxic events during early stages of dissemination; therefore, it is unclear how these tumors are able to maintain their aggressive phenotype. Thus, the identification of regulators of hypoxia-related genes in aggressive/metastatic tumors represents a fundamental step for the design of future therapies to treat pancreatic cancer. To this end, transcriptomic profiles were compared between the nonmetastatic pancreatic cancer cell line FG (LMET) and its angiogenic/metastatic derivate L3.6pl (HMET) under normoxic or hypoxic conditions. Cluster analysis revealed a number of transcripts that were induced by hypoxia in nonmetastatic cancer cells. Strikingly, this cluster was determined to be constitutively activated under normoxia in the metastatic cancer cells and could not be further induced by hypoxia. A subset of these transcripts were regulated by the transcription factor SOX9 in the aggressive-metastatic cells, but driven by hypoxia-inducible factor-1α (HIF-1α) in the parental nonmetastatic cell line. Moreover, these transcripts were enriched in cancer-related networks including: WNT, CXCR4, retinoic acid, and (FAK) focal adhesion kinase, gene PTK2 signaling pathways. In functional assays, inhibition of SOX9 expression in HMET cells led to increased apoptosis and reduced migration in vitro and a significant reduction in primary tumor growth, angiogenesis, and metastasis following orthotopic tumor cell injection. At the molecular level, the control of SOX9 expression was associated with changes in the methylation status of the SOX9 promoter. Finally, SOX9 upregulation was verified in a series of tumor specimens of patients with pancreatic carcinoma.Implications: SOX9 represents a novel target for pancreatic cancer therapy. Mol Cancer Res; 12(3); 421–32. ©2013 AACR.