Targeted inhibition of heat shock protein 90 disrupts multiple oncogenic signaling pathways, thus inducing cell cycle arrest and programmed cell death in human urinary bladder cancer cell lines

Panagiotis K Karkoulis,Eumorphia G Konstantakou,Dimitrios J Stravopodis,Gerassimos E Voutsinas

doi:10.1186/1475-2867-13-11

Abstract

BackgroundGeldanamycin (GA) can be considered a relatively new component with a promising mode of action against human malignancies. It specifically targets heat shock protein 90 (Hsp90) and interferes with its function as a molecular chaperone.MethodsIn this study, we have investigated the effects of geldanamycin on the regulation of Hsp90-dependent oncogenic signaling pathways directly implicated in cell cycle progression, survival and motility of human urinary bladder cancer cells. In order to assess the biological outcome of Hsp90 inhibition on RT4 (grade I) and T24 (grade III) human urinary bladder cancer cell lines, we applied MTT assay, FACS analysis, Western blotting, semi-quantitative (sq) RT-PCR, electrophoretic mobility shift assay (EMSA), immunofluorescence and scratch-wound assay.ResultsWe have herein demonstrated that, upon geldanamycin treatment, bladder cancer cells are prominently arrested in the G1 phase of cell cycle and eventually undergo programmed cell death via combined activation of apoptosis and autophagy. Furthermore, geldanamycin administration proved to induce prominent downregulation of several Hsp90 protein clients and downstream effectors, such as membrane receptors (IGF-IR and c-Met), protein kinases (Akt, IKKα, IKKβ and Erk1/2) and transcription factors (FOXOs and NF-κΒ), therefore resulting in the impairment of proliferative -oncogenic- signaling and reduction of cell motility.ConclusionsIn toto, we have evinced the dose-dependent and cell line-specific actions of geldanamycin on cell cycle progression, survival and motility of human bladder cancer cells, due to downregulation of critical Hsp90 clients and subsequent disruption of signaling -oncogenic- integrity.

Highlights

Geldanamycin (GA) can be considered a relatively new component with a promising mode of action against human malignancies
In the bladder cancer cell lines examined in the present study, geldanamycin administration primarily leads to a dose-dependent G1-checkpoint cell cycle arrest, while analysis of expression and activation profiles of several determinants of the cell cycle (Cdk4, pRb, Cyclin D1 and E2F1) correlate well with the observed block in cycle progression
heat shock protein 90 (Hsp90) inhibition due to geldanamycin administration proved to induce a severe reduction in the expression dynamics of several bona fide Hsp90 protein clients involved in cell cycle progression, subsequently diminishing bladder cancer cell proliferation potential

Summary

Introduction

Geldanamycin (GA) can be considered a relatively new component with a promising mode of action against human malignancies. It targets heat shock protein 90 (Hsp90) and interferes with its function as a molecular chaperone. Heat shock protein 90 (Hsp90) is an abundant intracellular protein that has emerged as a significant target of modern cancer therapeutic protocols. Hsp normally accounts for ~1-2% of the total protein content of the cell, while under stress conditions its levels increase up and a functional capacity. Cancer initiation, tumor progression and its clonal evolution are associated with a requirement for increased intracellular levels of Hsp90 [6,7,8]. Due to cancer cell dependence upon specific -oncogenic- Hsp protein clients, Hsp inhibition is thought to negatively interfere with critical oncogenic signaling pathways involved in the hallmark traits of cancer (i.e. sustaining cell proliferation, resisting cell death, and promoting invasion and metastasis) [9], demonstrating exciting prospects in the future of cancer therapeutics [10]

Methods

Results

Conclusion