Complex crystal structure determination and anti-non-small-cell lung cancer activity of the Hsp90^N inhibitor Debio0932

Debio0932 is a promising lead compound in phase I clinical trials targeting the N-terminal ATP-binding pocket of the molecular chaperone heat-shock protein 90 (Hsp90^N). The absence of a crystal structure of the Hsp90^N–Debio0932 complex, however, has impeded further structural optimization of Debio0932 and understanding of the molecular-interaction mechanism. Here, a high-resolution crystal structure of the Hsp90^N–Debio0932 complex was successfully determined (resolution limit 2.20 Å; PDB entry 6lr9) by X-ray diffraction and the molecular-interaction mechanism was analysed in detail, which suggested that Debio0932 suppresses cancer cells by accommodating itself in the ATP-binding pocket of Hsp90^N, disabling its molecular-chaperone capability. The results of a thermal shift assay (ΔT_m = 8.83 ± 0.90°C) and isothermal titration calorimetry (K_d = 15.50 ± 1.30 nM) indicated strong binding and favourable thermodynamic changes in the binding of Hsp90^N and Debio0932. Based on the crystal structure of the complex and on molecular-interaction analysis, 30 new Debio0932 derivatives were designed and nine new derivatives exhibited increased binding to Hsp90^N, as determined by molecular-docking evaluation. Additionally, Debio0932 suppressed cell proliferation (IC₅₀ values of 3.26 ± 2.82 µM for A549, 20.33 ± 5.39 µM for H1299 and 3.16 ± 1.04 µM for H1975), induced cell-cycle arrest and promoted apoptosis in three non-small-cell lung cancer (NSCLC) cell lines. These results provide novel perspectives and guidance for the development of new anti-NSCLC drugs based on the lead compound Debio0932.