The inflammatory cytokine TNFα cooperates with Ras in elevating metastasis and turns WT-Ras to a tumor-promoting entity in MCF-7 cells

BMC Cancer. 2014 Mar 6;14:158. doi: 10.1186/1471-2407-14-158.

Abstract

Background: In the present study we determined the relative contribution of two processes to breast cancer progression: (1) Intrinsic events, such as activation of the Ras pathway and down-regulation of p53; (2) The inflammatory cytokines TNFα and IL-1β, shown in our published studies to be highly expressed in tumors of >80% of breast cancer patients with recurrent disease.

Methods: Using MCF-7 human breast tumor cells originally expressing WT-Ras and WT-p53, we determined the impact of the above-mentioned elements and cooperativity between them on the expression of CXCL8 (ELISA, qRT-PCR), a member of a "cancer-related chemokine cluster" that we have previously identified. Then, we determined the mechanisms involved (Ras-binding-domain assays, Western blot, luciferase), and tested the impact of Ras + TNFα on angiogenicity (chorioallantoic membrane assays) and on tumor growth at the mammary fat pad of mice and on metastasis, in vivo.

Results: Using RasG12V that recapitulates multiple stimulations induced by receptor tyrosine kinases, we found that RasG12V alone induced CXCL8 expression at the mRNA and protein levels, whereas down-regulation of p53 did not. TNFα and IL-1β potently induced CXCL8 expression and synergized with RasG12V, together leading to amplified CXCL8 expression. Testing the impact of WT-Ras, which is the common form in breast cancer patients, we found that WT-Ras was not active in promoting CXCL8; however, TNFα has induced the activation of WT-Ras: joining these two elements has led to cooperative induction of CXCL8 expression, via the activation of MEK, NF-κB and AP-1. Importantly, TNFα has led to increased expression of WT-Ras in an active GTP-bound form, with properties similar to those of RasG12V. Jointly, TNFα + Ras activities have given rise to increased angiogenesis and to elevated tumor cell dissemination to lymph nodes.

Conclusions: TNFα cooperates with Ras in promoting the metastatic phenotype of MCF-7 breast tumor cells, and turns WT-Ras into a tumor-supporting entity. Thus, in breast cancer patients the cytokine may rescue the pro-cancerous potential of WT-Ras, and together these two elements may lead to a more aggressive disease. These findings have clinical relevance, suggesting that we need to consider new therapeutic regimens that inhibit Ras and TNFα, in breast cancer patients.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line, Tumor
  • Chick Embryo
  • Female
  • Gene Expression Regulation, Neoplastic
  • Guanosine Triphosphate / metabolism
  • Humans
  • Interleukin-1beta
  • Interleukin-8 / metabolism
  • MAP Kinase Signaling System
  • MCF-7 Cells
  • Mice
  • NF-kappa B / genetics
  • NF-kappa B / metabolism
  • Neoplasm Metastasis
  • Neoplasms / genetics
  • Neoplasms / metabolism*
  • Neoplasms / pathology*
  • Neovascularization, Pathologic / genetics
  • Neovascularization, Pathologic / metabolism
  • Protein Binding
  • Protein Interaction Domains and Motifs
  • Proto-Oncogene Proteins p21(ras) / chemistry
  • Proto-Oncogene Proteins p21(ras) / metabolism*
  • Signal Transduction
  • Transcription Factor AP-1 / genetics
  • Transcription Factor AP-1 / metabolism
  • Transcription, Genetic
  • Tumor Necrosis Factor-alpha / metabolism*
  • Tumor Suppressor Protein p53 / genetics
  • Tumor Suppressor Protein p53 / metabolism

Substances

  • Interleukin-1beta
  • Interleukin-8
  • NF-kappa B
  • Transcription Factor AP-1
  • Tumor Necrosis Factor-alpha
  • Tumor Suppressor Protein p53
  • Guanosine Triphosphate
  • Proto-Oncogene Proteins p21(ras)