Activated Oncogenic Pathway Modifies Iron Network in Breast Epithelial Cells: A Dynamic Modeling Perspective

Abstract

Dysregulation of iron metabolism in cancer is well documented and it has been suggested that there is interdependence between excess iron and increased cancer incidence and progression. In an effort to better understand the linkages between iron metabolism and breast cancer, a predictive mathematical model of an expanded iron homeostasis pathway was constructed that includes species involved in iron utilization, oxidative stress response and oncogenic pathways. The model leads to three predictions. The first is that overexpression of iron regulatory protein 2 (IRP2) recapitulates many aspects of the alterations in free iron and iron-related proteins in cancer cells without affecting the oxidative stress response or the oncogenic pathways included in the model. This prediction was validated by experimentation. The second prediction is that iron-related proteins are dramatically affected by mitochondrial ferritin overexpression. This prediction was validated by results in the pertinent literature not used for model construction. The third prediction is that oncogenic Ras pathways contribute to altered iron homeostasis in cancer cells. This prediction was validated by a combination of simulation experiments of Ras overexpression and catalase knockout in conjunction with the literature. The model successfully captures key aspects of iron metabolism in breast cancer cells and provides a framework upon which more detailed models can be built.

Fig 1. Intracellular iron network.

Iron homeostasis pathway depicted in green (LIP, TfR1, Fpn, Ft, IRP1, IRP2, Hep), iron utilization depicted in orange (Mfrn, LIPmt, Ftmt, ALAS1, heme, HO-1), oxidative stress response depicted in blue (ROS, Keap1, Nfr2, Antioxidant enzymes), and oncogenic pathway depicted in pink (EGFR, SOS, GAPs, Ras, ERK, c-Myc). IL-6, in yellow, is the only inflammatory cytokine in the network. Arrows represent activation/upregulation and hammer heads represent inhibition/downregulation. Dashed connections are explained in the iron utilization subsection of the introduction. Rectangular shapes represent proteins/enzymes, circular; molecules, hexagon-like; receptors. CellDesigner [67] was used for visualization.

Fig 2. Simulation results of the intracellular iron network.

Heatmap of point and cycle attractors of seven different knockout (k/o) and overexpression (o/e) models. Catalase low bioactivity (CAT k/o) is modeled by setting the AE group to zero inside the update rule for ROS (see cancer phenotype subsection for more details).

Fig 3. Effect of IRP2 overexpression (o/e) in MCF10A cells.

(a) One representative experiment. Proteins were analyzed by Western blotting. Loading was assessed with an antibody to GAPDH. (b) Proteins in empty vector cells and IRP2 overexpressing cells. Graphs show mean and standard deviation of three separate experiments.

Fig 4. Intracellular iron network with iron-sensing node.

(a) Simplified version of Fig 1 that includes a hypothesized mitochondrial iron-sensing node, which is depicted as a question mark. (b) Heat map of point attractors of three different knockout and overexpression models. (iron homeostasis (IH); oxidative stress response (OSR); oncogenic (Onc.); knockout (k/o); overexpression (o/e).)