Aurora Kinase Inhibitors as a Novel Targeted Drug Therapy for Bladder Cancer
Ning Zhou, Kamini Singh, Alex Almasan, Donna E Hansel. Cleveland Clinic, Cleveland, OH
Background: Conventional chemotherapy for invasive bladder cancer has limited efficacy and is generally associated with poor patient prognoses. This study aims to evaluate the potential of targeting the aurora kinases, a family of mitotic regulators, with pharmacologic inhibitors as a novel treatment for bladder cancer.
Design: Expression of genes associated with the mitotic spindle checkpoint, including the aurora kinases A and B, in clinical tissue samples of urothelial and squamous cell carcinomas was evaluated by RNA microarray and reverse-transcriptase PCR. Urothelial carcinoma cell lines UM-UC-3 and T24 were treated with the nonspecific aurora kinase inhibitor ZM447439 and the aurora kinase A inhibitor MLN8237, either alone or in combination with gemcitabine or paclitaxel. Effects of drug treatments were evaluated by flow cytometry with propidium iodide staining, immunofluorescence microscopy, MTS proliferation assay, and TUNEL labeling.
Results: RNA microarray analysis comparing human tissue specimens of urothelial (N=8) and squamous cell carcinomas (N=9) of the bladder to normal urothelium (N=10) identified overexpression of 13 gene transcripts related to the mitotic spindle checkpoint, including aurora kinases A and B, in the cancer specimens. Upregulation of these genes was validated by RT-PCR on a separate set of clinical samples (N=4 for each group). Next, we evaluated the impact of aurora kinase inhibitors on the bladder cancer cell lines in vitro. ZM447439 and MLN8237 treatment of UM-UC-3 and T24 cell lines at concentrations of 10 nM to 1 μM induced G2/M cell cycle arrest and aneuploidy (>4N DNA content) in a dose dependent manner. Immunofluorescence microscopy revealed abnormal mitotic figures with multipolar spindle apparatuses in treated cells. Both cell lines also exhibited cell death by positive TUNEL staining 48 hours after treatment with either inhibitor. Simultaneous treatment of T24 cells with MLN8237 and paclitaxel and subsequent MTS proliferation assay revealed an antagonistic interaction between paclitaxel and MLN8237, whereas simultaneous treatment with MLN8237 and gemcitabine resulted in an additive effect.
Conclusions: Several mitotic spindle checkpoint proteins, including the aurora kinases, are overexpressed in bladder cancer. Our data indicate that aurora kinase inhibitors have significant potential as a novel therapy for bladder cancer, as they induce abnormal mitosis, cell cycle arrest, and eventual death of bladder cancer cells in vitro.
Monday, March 19, 2012 11:30 AM
Platform Session: Section G2, Monday Morning