Studies On The In Vivo & In Vitro Antitumour Activity Of SNX-2112 And Its Mechanism To Induce A-375 Cell Apoptosis

Objective:To identify the antitumor activity of HSP90 small molecule inhibitors-SNX-2112 in vivo& in vitro, and using human melanoma A-375 cell line as a study model, to investigate its molecule mechanism of cell apoptosis induced SNX-2112, providing theoretical foundation for further development.Methods:MTT assay was employed to determine the antiproliferation effect of SNX-2112 against eight different solid tumor cells. Cell cycle and apoptosis were analyzed by flow cytometry afte PI staining or Annexin-V/PI staining. Mice H-22 orthotopic transplantation tumor model was constructed to observe the preliminary pharmacodynamics of SNX-2112 in vivo. The morphological change of apoptosis was characterized by DAPI staining and TUNEL. Western Blot, immunoprecipitation and immunofluorescence were applied to detect the influence of SNX-2112 in Caspase family protein expression, different pathways of apoptosis related protein expression and the expression of HSP90 client proteins after SNX-2112 administration.Results:We characterized that SNX-2112 has a broad-spectrum anti-solid tumor activity as evidenced by potently inducing growth inhibition and apoptosis in eight human solid cancer cells. SNX-2112 inhibition showed concentration and time dependent, IC50 values ranging from 0.7 to 22μM. SNX-2112 caused the cell cycle progression bloking in G2/M phase. In vivo, SNX-2112 could significantly inhibit the growth of mice implanted solid tumor of H-22 hepatoma cells.Inhibition rates of high, middle, low-dose group were:68.6%,31.4% and 15.1%. By laser confocal microscope, we observed representative morphological change of cell apoptosis, including chromatin condensation and marginalization and/or DNA fragmentation. We have witnessed that the SNX-2112-induced apoptosis-driving event was Caspase-dependent and Akt, B-Raf/Erk(1/2), IKKa/NF-kB-dependent signaling pathways were all involved in SNX-2112-mediated cell apoptosis. In addition, SNX-2112 induced degradation of Bcl-2, Gsk3βand p53, which played a critical regulation effect in tumor formation and devolopment. Our study showed that SNX-2112 tightly regulated the stability of Trap-1, without obviously impaction on Hsp90a/βand Grp94.Conclusions:Small-molecule target inhibitor, SNX-2112 potentially has significant anti-tumor activity in vivo& in vitro. Our results demonstrated that SNX-2112 induced tumor cell apoptosis through affecting related multi-pathway and multi-target.