Traditional medicines provide a fertile ground to explore potent lead compounds, yet their transformation into modern drugs is fraught with challenges in deciphering the target that is mechanistically valid for its biological activity. Herein we reveal that (Z)‐(+)‐isochaihulactone (1) exhibited significant inhibition against multiple‐drug‐resistant (MDR) cancer cell lines and mice xenografts. NMR spectroscopy showed that 1 resisted an off‐target thiolate, thus indicating that 1 was a target covalent inhibitor (TCI). By identifying the pharmacophore of 1 (α,β‐unsaturated moiety), a probe derived from 1 was designed and synthesized for TCI‐oriented activity‐based proteome profiling. By MS/MS and computer‐guided molecular biology approaches, an affinity‐driven Michael addition of the noncatalytic C247 residue of GAPDH was found to control the “ON/OFF” switch of apoptosis through non‐canonically nuclear GAPDH translocation, which bypasses the common apoptosis‐resistant route of MDR cancers. Overcoming resistance: A new class of inhibitors that target the noncatalytic C247 residue of GAPDH in an affinity‐driven manner to initiate SIAH1‐dependent apoptosis and androgen‐receptor degradation has been discovered (see picture). The affinity‐driven Michael addition of isochaihulactone was found to control the “ON/OFF” switch of apoptosis by a mechanism that bypasses the common apoptosis‐resistant route of multiple‐drug‐resistant cancers.