The effect of hsp90 inhibitors on histone methylation in bladder cancer

Heat shock protein 90 (HSP90) is an ATP-dependent molecular chaperone that required for the stability and function of numerous oncogenic signaling proteins that determine the hallmarks of cancer. HSP90 has been shown to play an privotal role in epigenetic pathways in recent years. Bladder cancer constitutes the second most common type of cancer among urogenital cancers. Bladder cancer has traditionally been accepted as a genetic disease, however it is known that the epigenetic mechanisms also play an important role in the etiology of this disease. The aim of this thesis project to explain how the HSP90 protein affects to inactivating histone methylation markers levels in bladder cancers. Cytotoxic effect of HSP90 spesific inhibitor Geldanamycin (GA) in human bladder cancer cell line T24 was detected time and dose dependent manner with WST1, expression aberration of target genes DNMT1, HDAC1 and WIF-1 using real-time PCR and protein levels of DNMT1, H3K27me3, Histone H4, H4K20me1, H4K20me3, Akt1, pAkt1(S473), SET8, KMT5C, Bax and Bcl2 using western blot. Our results demostrated that GA caused an decreased cell proliferation at high dose (10 µM). In both low and high dose cases, GA was decreased DNMT1 at both transcriptional and translational levels via inhibition of Akt1 and therewithal HDAC1 expression. However, reexpression of WIF1 gene was observed after treatment GA based on decreased DNMT1 and HDAC1 gene levels. Furthermore we showed that histone post-translational modifications (H3K27me3, H4K20me1 and H4K20me3) and enzymes (SET8 and KMT5C) responsible for this modifications was up regulated or downregulated depend on the dose and time following GA treatment and also not observe significantly any changes at Histone H4 protein level. We demonstrated to induce apoptosis following reexpression of WIF. Our findings may offer a new approach determination of molecular effect of HSP90 on epigenetic regulation would allow us to identify new molecular targets (HSP90 client proteins) for treatment of the invasive bladder cancer.