Oncogene (2005) 24, 7720–7728 & 2005 Nature Publishing Group All rights reserved 0950-9232/05 $30.00 www.nature.com/onc Unshackling the links between reovirus oncolysis, Ras signaling, translational control and cancer Maya Shmulevitz1, Paola Marcato1 and Patrick WK Lee*,1 1Department of Microbiology and Immunology, Dalhousie University, 7P Sir Charles Tupper Medical Building, 5850 College Street, Halifax, NS, Canada B3K 1X5 Reovirus has an inherent preference for replicating in cells cancer cells. Viruses have evolved intricate strategies to with dysregulated growth factor signaling cascades that ensure efficient replication in the host cell. Viral encoded comprise Ras activation. Precisely how reovirus exploits factors can alter the regulation of cellular processes the host cell Ras pathway is unclear, but there is evidence important for viral replication. Processes amicable to suggesting that activated Ras signaling is important for viral replication are provoked, while those that are efficient viral protein synthesis. Defining the molecular detrimental for replication are prevented. For example, mechanism of reovirus oncolysis will shed light on reovirus DNA viruses that require cell division for viral genome replication and important aspects of cellular transforma- replication have evolved strategies to ensure that cells tion, Ras signaling cascades and regulation of protein proliferate. Cellular responses that would clear viral translation. infections such as apoptosis, interferon (IFN)-induced Oncogene (2005) 24, 7720–7728. doi:10.1038/sj.onc.1209041 antiviral activities and presentation of viral antigens by multihistocompatibility class I (MHC-I) molecules are Keywords: cancer; oncolytic virus; oncolysis; reovirus; overcome to different extents by various viruses. In view translation; Ras of such highly evolved strategies to ensure efficient replication in cells, how can viruses be used specifically to kill cancer cells? With increased understanding of virus replication, viral-cell interactions and methods to manipulate viral Viruses with inherent or engineered preference for genomes, viruses have been generated that are no longer replication in cancer cells able to regulate specific host processes. The deletion of viral proteins important in regulating cellular processes Many exciting developments have been made in the last can, therefore, create viruses that are unable to replicate two decades with respect to the use of replication- in normal cells. Conversely, cellular events involved in competent viruses as specific oncolytic agents. A handful tumorigenesis, such as increased cell division and of oncolytic viruses have been identified. Some naturally dysregulated cell death programs, make tumor cells occurring viruses preferentially infect and kill trans- permissive to replication by these recombinant viruses. formed cells, such as vesicular stomatitis virus (VSV), ONYX-015 is an example of an engineered oncolytic herpes simplex virus (HSV), Newcastle disease virus virus, and is described thoroughly by Clodagh O’Shea (NDV) and reovirus (Norman and Lee, 2000; Stojdl and Frank McCormick in this edition of Oncogene et al., 2000; Sinkovics and Horvath, 2000; Farassati reviews. The engineered ONYX-015 virus is unable to et al., 2001). The Edmonston-B strain of measles virus make E1B-55K, an early adenoviral protein critical for (MV-Edm), originally attenuated for use as a live virus degrading p53 and thus overcoming p53-mediated cell vaccine, has potential oncolytic application (Grote et al., cycle arrest and apoptosis (Biederer et al., 2002). The 2001). Furthermore, several genetic modifications to selective oncolysis by ONYX-015 is likely in part due to adenoviruses, influenza viruses and herpes viruses have the dysregulation of p53 pathways in susceptible tumor been shown to confer selective oncolytic activity cells. Interestingly, it was recently discovered that (Martuza, 2000; Bergmann et al., 2001; Dobbelstein, following infection with ONYX-015 efficient export of 2004). A growing body of research demonstrates that late adenoviral RNA from the nucleus is restricted to these viruses can efficiently and selectively destroy tumor cells (O’Shea et al., 2004). This finding shows that transformed cell lines and also have antitumor proper- novel and unperceived differences between tumor and ties in vivo. Exciting developments have also been made normal cells can affect virus replication. in understanding the molecular bases of viral oncolysis. Remarkably, reoviruses show an inherent prefe- The use of replication-competent viruses for oncolysis rence for replication in many transformed cells. depends on their ability to selectively infect and destroy Possible explanations for why such a preference may have developed through the evolution of reovirus are discussed in this review. Reoviruses, the prototype *Correspondence: PWK Lee; E-mail: [email protected] member of the Reoviridae family, were first isolated Molecular mechanism of reovirus oncolysis M Shmulevitz et al 7721 from the intestinal tracts of apparently healthy indivi- replication, while transfection of EGFR into these cells duals. In fact, most people have been exposed to proved significantly advantageous for reovirus infection reovirus by the age of 5 with little or no manifestation (Strong et al., 1993). Interestingly, the involvement of of symptoms. Hence, there are fewer concerns about the EGFR in reovirus replication was found to be safety of using reovirus in cancer treatment. As this dependent on signaling pathways initiated through the review will describe in detail, an understanding of the tyrosine kinase receptor rather than receptor binding cellular events that contribute to the conditional specificity. A mutated EGFR devoid of signaling replication of reovirus is emerging. Growth factor capabilities was unable to confer permissiveness to receptor signaling is intimately related with tumor reovirus replication, while the v-erbB oncoprotein, progression. Genetic changes resulting in constitutive which lacks the extracellular ligand-binding domain activation of growth factor signaling pathways result in but has a constitutively active kinase domain, was uncontrolled proliferation, differentiation and/or me- sufficient to permit reovirus infection (Strong et al., tastasis, and are associated with most if not all human 1993; Strong and Lee, 1996). Therefore, what started as cancers. Intriguingly, reovirus replication is sensitive to a search for a secondary receptor for reovirus entry that the status of signaling cascades downstream of specific could permit selective replication in transformed cells tyrosine kinase receptors such as the epidermal growth turned into a very interesting connection between the factor receptor (EGFR). The pathways downstream of status of intracellular signaling pathways and the tyrosine kinase receptors are not only numerous, but are reovirus life cycle (Figure 1). complicated by extensive cross-talk. Untangling the large networks of interactions coupled with cell signal- Involvement of Ras/RalGEF/p38 signaling pathways in ing and phenotypic alterations associated with cancer is reovirus replication a challenge. Therefore, not only is reovirus a promising oncolytic agent but also reovirus replication offers a The interaction of growth factors with their cognate unique approach to understanding the components and receptors results in the activation of a cascade of outcomes of signal transduction pathways aberrantly intracellular biochemical events leading to a cohort of regulated in cancer. cellular functions. Which cellular functions can affect reovirus replication? Which components of the signaling pathway are important? The EGFR signaling cascade initiates with receptor oligomerization and subsequent Intracellular signaling pathways involved in selective activation of tyrosine kinase activity resulting in replication of reovirus autophosphorylation of specific tyrosine residues (Ull- How the link between selective reovirus replication and rich and Schlessinger, 1990; Fantl et al., 1993; Weiss and cancer was discovered Schlessinger, 1998). Adaptor molecules with phospho- tyrosine binding, src homology 2 (SH2) domains are Differences between virally or spontaneously trans- recruited to activated EGFR and in turn assemble a formed cell lines and primary or untransformed cells cohort of proteins particular to one of many downstream with respect to their susceptibility to reovirus cytotoxi- cascades (Pawson, 1994; Pawson and Scott, 1997; Buday, city were first noted in 1977 (Hashiro et al., 1977). 1999). The Ras signaling cascade is a major pathway Likewise, human lung fibroblast cells (WI-38) showed downstream of EGFR (Campbell et al., 1998). Ras enhanced permissibility to reovirus replication when proteins form a subfamily of small GTP-binding proteins they were transformed with the simian virus 40 T- involved in regulation of a wide variety of cellular antigen (Duncan et al., 1978). Nevertheless, it was not function such as cell growth, differentiation and cell until the 1990s that clues were obtained regarding the survival. Membrane-anchored Ras proteins cycle be- molecular basis of the selective replication of reovirus in tween the inactive GDP-bound state and the active GTP- transformed cells. bound state. The activation of Ras proteins is promoted Receptor specificity often dictates the tropism of by guanine nucleotide exchange factors such as the viruses (Cohen et al., 1988; Schneider-Schaulies, 2000; Son of Sevenless (Sos), which are