snx-7081 and fludarabine

Clinical trials of heat shock protein 90 (Hsp90) inhibitors have been limited by high toxicity. We previously showed that the Hsp90 inhibitor, SNX-7081, synergizes with and restores sensitivity to fludarabine nucleoside (2-FaraA) in human chronic lymphocytic leukemia (CLL) cells with lesions in the p53 pathway (Best OG, et al., Leukemia Lymphoma 53:1367-75, 2012). Here, we used label-free quantitative shotgun proteomics and comprehensive bioinformatic analysis to determine the mechanism of this synergy. We propose that 2-FaraA-induced DNA damage is compounded by SNX-7081-mediated inhibition of DNA repair, resulting in enhanced induction of apoptosis. DNA damage responses are impaired in part due to reductions in checkpoint regulators BRCA1 and cyclin D1, and cell death is triggered following reductions of MYC and nucleolin and an accumulation of apoptosis-inducing NFkB2 p100 subunit. Loss of nucleolin can activate Fas-mediated apoptosis, leading to the increase of pro-apoptotic proteins (BID, fas-associated factor-2) and subsequent apoptosis of p53-negative, 2-FaraA refractory CLL cells. A significant induction of DNA damage, indicated by increases in DNA damage marker γH2AX, was observed following the dual drug treatment of additional cell lines, indicating that a similar mechanism may operate in other p53-mutated human B-lymphoid cancers. These results provide valuable insight into the synergistic mechanism between SNX-7081 and 2-FaraA that may provide an alternative treatment for CLL patients with p53 mutations, for whom therapeutic options are currently limited. Moreover, this drug combination reduces the effective dose of the Hsp90 inhibitor and may therefore alleviate any toxicity encountered.

Topics: Antineoplastic Agents; Apoptosis; Benzamides; Blotting, Western; BRCA1 Protein; Cell Line, Tumor; Chromatography, Liquid; Cyclin D1; DNA Damage; DNA Repair; Drug Synergism; HSP90 Heat-Shock Proteins; Humans; Leukemia, Lymphocytic, Chronic, B-Cell; Mutation; NF-kappa B p52 Subunit; Nucleolin; Phosphoproteins; Protein Interaction Maps; Proteomics; Proto-Oncogene Proteins c-myc; RNA-Binding Proteins; Signal Transduction; Tandem Mass Spectrometry; Tumor Suppressor Protein p53; Vidarabine

Drug resistance in chronic lymphocytic leukemia (CLL) associated with lesions in the ATM/TP53 pathway represents a major challenge in clinical management. Evidence suggests that heat shock protein-90 (Hsp90) inhibitors may represent a therapeutic option in combination with more conventional therapies. We explored the effects of combining the Hsp90 inhibitor, SNX-7081, with fludarabine in vitro against CLL cells and hematological cell lines. In seven cell lines and 23 patient samples synergy between SNX-7081 and fludarabine (2-FaraA) was apparent in the three TP53 mutated cell lines and at significantly lower concentrations in TP53 or ATM dysfunctional patient cells. In 11/13 2-FaraA-resistant patient samples, SNX-7081 reduced the 50% inhibitory concentration to within a clinically achievable range. Synergy between SNX-7081 and 2-FaraA was evident in both the cell lines and patient samples as a significant decrease in cell viability. Our data suggest that combining SNX-7081 and fludarabine may be effective in the treatment of fludarabine-refractory CLL.

Topics: Antineoplastic Agents; Apoptosis; Ataxia Telangiectasia Mutated Proteins; Benzamides; Cell Cycle; Cell Cycle Proteins; Cell Line; Cell Line, Tumor; Cell Survival; DNA-Binding Proteins; Dose-Response Relationship, Drug; Drug Resistance, Neoplasm; Drug Synergism; Flow Cytometry; HL-60 Cells; HSP90 Heat-Shock Proteins; Humans; Leukemia, Lymphocytic, Chronic, B-Cell; Mutation; Protein Serine-Threonine Kinases; Signal Transduction; Tumor Cells, Cultured; Tumor Suppressor Protein p53; Tumor Suppressor Proteins; Vidarabine