Abstract
Activating mutation of KRas is a genetic alteration that occurs in the majority of pancreatic tumors and is therefore an ideal therapeutic target. The ability of reoviruses to preferentially replicate and induce cell death in transformed cells that express activated Ras prompted the development of a reovirus-based formulation for cancer therapy called Reolysin. We hypothesized that Reolysin exposure would trigger heavy production of viral products leading to endoplasmic reticular (ER) stress-mediated apoptosis. Here, we report that Reolysin treatment stimulated selective reovirus replication and decreased cell viability in KRas-transformed immortalized human pancreatic duct epithelial cells and pancreatic cancer cell lines. These effects were associated with increased expression of ER stress-related genes, ER swelling, cleavage of caspase-4, and splicing of XBP-1. Treatment with ER stress stimuli including tunicamycin, brefeldin A, and bortezomib (BZ) augmented the anticancer activity of Reolysin. Cotreatment with BZ and Reolysin induced the simultaneous accumulation of ubiquitinated and viral proteins, resulting in enhanced levels of ER stress and apoptosis in both in vitro and in vivo models of pancreatic cancer. Our collective results demonstrate that the abnormal protein accumulation induced by the combination of Reolysin and BZ promotes heightened ER stress and apoptosis in pancreatic cancer cells and provides the rationale for a phase I clinical trial further investigating the safety and efficacy of this novel strategy.
Highlights
Reoviruses are naturally occurring viruses that are nonpathogenic and have been reported to replicate in cancer cells with an activated Ras pathway but not in normal tissue.[4,5,6] To take advantage of this observation therapeutically, the reovirus-based anticancer agent Reolysin was recently developed and has already progressed into clinical trials,[7,8,9,10,11] but its mechanism(s) of action remains unclear
We demonstrate that Reolysin induces an accumulation of viral products in pancreatic cancer cells with activated Ras, which results in endoplasmic reticular (ER) stress and apoptosis
As expected based on earlier studies conducted in other cell types, exposure to Reolysin resulted in preferential reovirus replication in KRas-positive HPNE cells (Figure 1b)
Summary
Reoviruses are naturally occurring viruses that are nonpathogenic and have been reported to replicate in cancer cells with an activated Ras pathway but not in normal tissue.[4,5,6] To take advantage of this observation therapeutically, the reovirus-based anticancer agent Reolysin was recently developed and has already progressed into clinical trials,[7,8,9,10,11] but its mechanism(s) of action remains unclear. We hypothesized that unchecked viral replication in Ras-activated pancreatic cancer cells would promote endoplasmic reticular (ER) stress and apoptosis. We demonstrated that treatment with the proteasome inhibitor bortezomib (BZ) generated a Received 18.2.13; revised 07.6.13; accepted 17.6.13; Edited by C Munoz-Pinedo substantial accrual of ubiquitin-conjugated proteins and induced ER stress-mediated apoptosis in both in vitro and in vivo models of pancreatic cancer.[14,15,16] Considering that abnormal protein buildup can trigger pancreatic cancer cell death, the simultaneous accumulation of ubiquitinated proteins and viral products may be especially toxic to pancreatic cancer cells. We demonstrate that Reolysin induces an accumulation of viral products in pancreatic cancer cells with activated Ras, which results in ER stress and apoptosis. Further stimulation of ER stress with conventional ER stress-inducing agents (i.e., tunicamycin) or BZ augments the anticancer activity of Reolysin in both in vitro and in vivo models of pancreatic cancer
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