Identifying HSP90a as a therapeutic target in BCGrefractory disease.

Abstract

Introduction: The current standard of care for high-grade non-muscle invasive bladder cancer (NMIBC) is intravesical instillation of Bacillus Calmette-Gu_erin (BCG). While BCG therapy has been shown to reduce recurrence and progression, treatment failure occurs in up to 45% of patients. Those who progress after BCG therapy have poor prognoses despite curative interventions. Therefore, identifying novel therapeutic strategies to enhance BCG efficacy and reduce bladder cancer recurrence is of critical clinical importance. We aimed to identify a targetable pathway in patients with BCG refractory bladder cancer. Method(s): Matched post BCG clinical samples from eight BCG-responsive NMIBC and eight refractory patients were identified. RNA sequencing was used to assess differentially expressed genes (DEGs) between responders and non-responders. Based on the DEGs, gene expression of HSP90AA1 in MB49-LUC mouse bladder cancer cell line was used to assess viability after STA-9090, a heat shock protein 90 (HSP90) inhibitor, using PrestoBlue cell viability reagent. Result(s): Analysis of DEGs identified 16 significantly altered signalling pathways between post-BCG responsive and refractory NMIBC tissues (Fig. 1). Majority of these pathways, such as Interferon Gamma Response, IL6-JAK-STAT3 Signalling, Natural Killer Cells, Inflammatory Response, TNFa Signalling via NF-jB, E2F Targets, MYC Targets V1, and Oxidative Phosphorylation, were modulated by HSP90a (HSP90 family), a multifunctional chaperone protein. RNA expression of HSP90AA1 was also significantly higher in refractory NMIBC compared to responsive tissues (log2 fold change = 1.644, P = 0.001). HSP90AA1 gene expression in MB49-LUC cells was significantly higher than in normal C57 mouse bladder tissue (P < 0.05). Treatment of MB49-LUC cells with STA-9090 significantly suppressed tumour cell growth compared to vehicle. 1 nM STA-9090 treatment reduced cell viability by 30.82% compared with vehicle (P < 0.01); 5 nM STA-9090 treatment reduced cell viability by 92.66% compared with vehicle (P < 0.0001); 10 nM STA-9090 treatment reduced cell viability by 92.44% compared with vehicle (P < 0.0001); 15 nM STA-9090 treatment reduced cell viability by 95.03% compared with vehicle (P < 0.0001); 20 nM STA-9090 treatment reduced cell viability by 95.99% compared with vehicle (P < 0.0001). Additionally, treatment with 5 nM STA-9090 reduced cell viability by 89.38% compared with 1 nM STA-9090 treatment (P < 0.0001). Conclusion(s): Our results demonstrate that the majority of pathways modulated by HSP90a show differential RNA expression in patients with BCG-refractory bladder cancer, with some pathways downregulated (Interferon Gamma Response, IL6-JAK-STAT3 Signalling, Natural Killer Cells, Inflammatory Response, and TNFa Signalling via NF-jB) and others upregulated (E2F Targets, MYC Targets V1, and Oxidative Phosphorylation). Our results demonstrate that the majority of HSP90 RNA-modulated pathways show differential regulation in patients with BCG-refractory bladder cancer. Inhibition of HSP90AA1 effectively reduced tumour cell viability in vitro using a mouse cancer model. Further in vivo studies are warranted to confirm the role of HSP90a in driving BCG-refractory disease and to evaluate its potential as a therapeutic target.