A Quantitative Systems Biology Framework of the NRF2–KEAP1–sMAF–BACH1–ARE Network Dynamics for Drug-Target Testing

The KEAP1–NRF2 pathway is the principal regulator of the cellular antioxidant response and an actively pursued drug target, with both activators and, more recently, inhibitors entering clinical development. Quantitative models of the pathway have treated the upstream redox sensor and the downstream competition of NRF2, small MAF and BACH1 for antioxidant response elements (AREs) separately, so that the feedback between transcriptional output and the oxidant that drives it has not been represented. Here we integrate an experimentally parameterised NRF2–KEAP1 redox model with a measured-constant NRF2–sMAF–BACH1–ARE competition module into a single 28-species, 24-reaction ordinary differential equation model, encoded in SBML and calibrated against NRF2 time courses in PEO1 ovarian cancer cells. The integrated model reproduces the source model exactly in the absence of BACH1. ARE activation follows a non-cooperative hyperbolic dependence on nuclear NRF2 (Hill coefficient ≈ 1) that BACH1 shifts competitively, raising the half-maximal nuclear NRF2 concentration about sevenfold at 1 µM BACH1 without changing the maximal occupancy. Modelling of hydrogen peroxide boluses reveals a threshold between 5 and 400 µM above which clearance slows rather than accelerates and the system can escape to a high-oxidant state. Global sensitivity analysis shows that control resides mainly in a lumped ratio of oxidant production to thioredoxin regeneration capacity, which predicts loss of a steady state with 96.6% accuracy, followed by NRF2 availability and, third, the ARE competition layer. Screening mechanism-level perturbations against these two separable blocks ranks disruption of the KEAP1–NRF2 interaction above reduced KEAP1-dependent ubiquitination, places KEAP1 degradation outside the useful region, and predicts near-multiplicative benefit from combining NRF2 activation with BACH1 inhibition in proportion to BACH1 expression. Perturbing the same axis in the opposite direction reproduces the expected behaviour of the recently described covalent allosteric molecular glue class of NRF2 inhibitors. We also identify a structural feature of the inherited backbone, in which oxidation of KEAP1 carries most of the H2O2 flux, which limits the oxidant-load predictions and points to a specific model correction. The framework, its SBML files and analysis scripts are provided as a reusable platform for mechanism-level target testing.