Hyperoside impairs mitochondrial respiration in chronic myeloid leukemia by promoting STUB1-mediated ubiquitination and degradation of NOX4.

Chronic myeloid leukemia (CML) cells exhibit a distinct reliance on oxidative phosphorylation (OXPHOS), presenting a metabolic vulnerability for therapeutic intervention. While Hyperoside, a natural flavonol, demonstrates anti-leukemic activity, its precise molecular targets and impact on mitochondrial bioenergetics remain elusive. In this study, we combined in silico modeling and biophysical validation to identify NADPH oxidase 4 (NOX4) as a direct molecular target of Hyperoside. Surface plasmon resonance and cellular thermal shift assays confirmed high-affinity binding between Hyperoside and NOX4. Hyperoside promotes the proteasomal degradation of NOX4 by enhancing its interaction with the E3 ubiquitin ligase STUB1. This Hyperoside-induced recruitment of STUB1 accelerates NOX4 ubiquitination and turnover in K562 and Meg-01 cells. Functionally, Hyperoside treatment resulted in a severe impairment of mitochondrial respiration, including reduced oxygen consumption rates, diminished ATP production, and inhibitory phosphorylation of the pyruvate dehydrogenase complex. Importantly, these metabolic defects were reversed by NOX4 overexpression or STUB1 knockdown. In summary, our findings characterize a novel STUB1/NOX4 signaling axis and establish Hyperoside as a targeted metabolic modulator that dismantles the bioenergetic machinery essential for CML cell survival.