AbstractEpidermal growth factor receptor (EGFR) is an intensively focused drug target for non‐small cell lung cancer (NSCLC). JBJ‐04–125–02 is an effective ATP‐noncompetitive and T790M/L858R‐selective inhibitor of EGFR, but the implied negative regulation mechanism is not fully clarified. Here, computational approaches were employed to address this. We find that JBJ‐04–125–02 induces contrary effects on the binding of adenosine and phosphate moieties of ATP. The allosteric inhibitor lowers the stability of the hinge region, affecting the anchor of the adenosine portion of ATP, while a more closed conformation of P‐loop is observed and might be unfavorable for the phosphotransfer and product release. The umbrella sampling simulations further demonstrate that less free energy is needed for the initial dissociation of ATP (the adenosine group) from the inactive EGFR in the presence of JBJ‐04–125–02, but more for the phosphate groups egressing from the active cavity. Together, these findings provide a deeper understanding of the negative regulation of JBJ‐04–125–02. Moreover, the key inter‐molecular interactions contributing to ATP binding are identified. Our work might pave the way for designing allosteric drugs targeting EGFR for lung cancer patients, and also suggests that computational techniques are effective for investigating the allosteric mechanism.