Abstract 967: Development of fluorescent-cell based high throughput assay to identify novel therapeutics of bladder cancer cells through upregulation of SPARC expression

Abstract

Abstract Bladder cancer is the most common malignancy affecting the urinary system with an estimated 76,960 new cases and projected 16,390 deaths in 2016 in the United States. The role of SPARC in various cancers is contextual and differs based on the tumor classification, malignancy and progression. We have recently reported the tumor suppressor effect of SPARC in urinary bladder cancer. We have shown that the loss of SPARC in bladder cancer resulted accelerated bladder carcinogenesis and metastasis multiple pathways involving cell cycle deregulation, inflammation and angiogenesis. We have also reported that SPARC protein expression significantly decreased in the cancerous compartment in advanced human bladder cancer as well as in carcinogen-induced murine urothelial cancer. However, the mechanism of the downregulation of SPARC expression in cancer in general and in bladder cancer in particular is still unraveled. The purpose of our study is to develop a robust, high throughput assay to identify therapeutic candidates as regulators of SPARC expression in bladder cancer. We cloned SPARC promoter in a third generation lentiviral vector coupled with fluorescent GFP or mCherry fluorescent protein to quantitatively measure SPARC expression. We generated stable bladder cancer cell lines expressing SPARC expression in bladder cancer cell lines. UMUC3 bladder cancer cells were transduced with lentiviral vector to generate SPARC-promoter reporter system to screen regulators of SPARC expression. Reporter cells were plated in 96 well plates and treated with 10uM of FDA-approved and natural product drug libraries. Real time monitoring of changes in SPARC expression was done by measuring GFP fluorescence intensity using IncuCyte® ZOOM Live Imaging Analysis System. We identified drugs that consistently exhibited dosage-dependent augmentation or down-regulation of SPARC expression in UMUC3 cells. Our screens identified novel up-regulators of SPARC expression that are both standard of care and novel bladder cancer therapeutics. Our efforts produced an efficient robust assay to identify novel therapeutics and regulators of the tumor suppressor SPARC in the bladder cancer ecosystem that can be used in the adjuvant and neoadjuvant settings. Citation Format: Chirayu M. Patel. Development of fluorescent-cell based high throughput assay to identify novel therapeutics of bladder cancer cells through upregulation of SPARC expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 967. doi:10.1158/1538-7445.AM2017-967