Serge A. Afeli, PhD, Presbyterian College
PROJECT TITLE: Role of MRP5 transporters in urothelial bladder cancer cells survival
In the U.S., bladder cancer accounts for nearly 76,000 new cancer cases, 17,000 deaths, and $4 billion associated costs annually. It is the fourth deadliest cancer in men and fifth in women. The combination therapy cisplatin-gemcitabine is currently the gold standard for bladder cancer treatment. Unfortunately, in patients with unresectable bladder cancer, cisplatin-gemcitabine response rate is only 49% with approximately 38% of patients surviving for up to 18 months. In most cancers, chemotherapy failure is multifactorial and may result from an increase in drug efflux transporters expression. ATP-binding cassette (ABC) transporters are drug influx/efflux proteins physiologically expressed in cellular and intracellular membranes. They can pump various drugs out of cancer cells before they reach their optimum concentration and cause treatment failure. ABCC5 (MRP5) transporter is a key member of the ABC transporter family which similarly to p-glycoprotein has been associated with chemoresistance in various cancers. In pancreatic cancer cells, 5-fluorouracil and gemcitabine treatment failure was correlated to MRP5 transporters overexpression. In non-small cell lung cancer and human embryonic kidney cells, MRP5 transporters accelerated the extrusion of gemcitabine and 5-fluorouracil cytotoxic metabolic from the cells which led to drug resistance. Unlike in other cancer types, information about MRP5 transporters in bladder cancer remains scarce. Our long-term goal is to understand the regulation of urothelial bladder cancer cells by MRP5 transporters and further identify the molecular pathways linked to MRP5 expression. This proposal aims to elucidate MRP5 transporters gene expression level in bladder cancer cells before and after chemotherapy as well as outline the genetic markers affected by cisplatin and gemcitabine treatment. We will utilize molecular biology techniques including qPCR, western blot, and immunocytochemistry with confocal imaging as well as microarray analysis on urothelial bladder cancer cells. Our central hypothesis is that MRP5 transporters are key regulators of urothelial bladder cancer cells survival, and therefore a better understanding of their physiological role may provide additional options to better manage treatment failure in bladder cancer patients.
With SC INBRE Bioinformatics Pilot Project Program funding, we were able to purchase reagents, perform cell imaging, and train students.
With the aid of this funding, we provided the first molecular evidence for MRP5 transporters proteins in urothelial bladder cancer (RT4) cells using western blot and immunocytochemistry. We have successfully created RT4 cells in which the MRP5 transporter gene has been silenced using shRNA approach. We are now in the process of using microarray analysis to span for genes that are up or downregulated upon chemotherapy intervention in both native RT4 cells as well as RT4 cells in which MRP5 transporters have been silenced.
September 22, 2017