The emergence of combinatorial strategies in the development of RNA oncolytic virus therapies.

Oncolytic viruses (OVs) represent an exciting new biological approach to cancer therapy. In particular, RNA viruses have emerged as potent agents for oncolytic virotherapy because of their capacity to specifically target and destroy tumour cells while sparing normal cells and tissues. Several barriers remain in the development of OV therapy, including poor penetration into the tumour mass, inefficient virus replication in primary cancers, and tumour-specific resistance to OV-mediated killing. The combination of OVs with cytotoxic agents, such as small molecule inhibitors of signalling or immunomodulators, as well as stealth delivery of therapeutic viruses have shown promise as novel experimental strategies to overcome resistance to viral oncolysis. These agents complement OV therapy by unblocking host pathways, delivering viruses with greater efficiency and/or increasing virus proliferation at the tumour site. In this review, we summarize recent development of these concepts, the potential obstacles, and future prospects for the clinical utilization of RNA OVs in cancer therapy.

Targeting the apoptotic pathway with BCL-2 inhibitors sensitizes primary chronic lymphocytic leukemia cells to vesicular stomatitis virus-induced oncolysis.

Chronic lymphocytic leukemia (CLL) is characterized by clonal accumulation of CD5(+) CD19(+) B lymphocytes that are arrested in the G(0)/G(1) phase of the cell cycle and fail to undergo apoptosis because of overexpression of the antiapoptotic B-cell CLL/lymphoma 2 (BCL-2) protein. Oncolytic viruses, such as vesicular stomatitis virus (VSV), have emerged as potential anticancer agents that selectively target and kill malignant cells via the intrinsic mitochondrial pathway. Although primary CLL cells are largely resistant to VSV oncolysis, we postulated that targeting the apoptotic pathway via inhibition of BCL-2 may sensitize CLL cells to VSV oncolysis. In the present study, we examined the capacity of EM20-25--a small-molecule antagonist of the BCL-2 protein--to overcome CLL resistance to VSV oncolysis. We demonstrate a synergistic effect of the two agents in primary ex vivo CLL cells (combination index of 0.5; P < 0.0001). In a direct comparison of peripheral blood mononuclear cells from healthy volunteers with primary CLL, the two agents combined showed a therapeutic index of 19-fold; furthermore, the combination of VSV and EM20-25 increased apoptotic cell death in Karpas-422 and Granta-519 B-lymphoma cell lines (P < 0.005) via the intrinsic mitochondrial pathway. Mechanistically, EM20-25 blocked the ability of the BCL-2 protein to dimerize with proapoptotic BAX protein, thus sensitizing CLL to VSV oncolytic stress. Together, these data indicate that the use of BCL-2 inhibitors may improve VSV oncolysis in treatment-resistant hematological malignancies, such as CLL, with characterized defects in the apoptotic response.

Sequence analysis of a 5.1 kbp region of the Spodoptera frugiperda multicapsid nucleopolyhedrovirus genome that comprises a functional ecdysteroid UDP-glucosyltransferase (egt) gene.

The ecdysteroid UDP-glucosyltransferase gene from the Spodoptera frugiperda multicapsid nucleopolyhedrovirus (SfMNPV) was identified using degenerate primers whose sequence were derived from conserved regions of the EGT proteins encoded by other baculoviruses. Analysis of the gene sequence revealed the presence of an open reading frame (ORF) with potential to encode a polypeptide of 525 amino acids. Promoter sequences typical of baculovirus genes were found in the 5' region of this ORF. A polyadenylation signal was identified downstream the translation stop codon. A transient expression assay showed that the product of this ORF was able to conjugate glucose from UDP-glucose with ecdysone confirming that the gene identified was indeed the SfMNPV egt gene. The SfMNPV egt gene and the sequences of other baculovirus egt genes were used to infer a phylogenetic tree. The nucleotide sequence of the entire BamHI fragment that contains the SfMNPV egt gene was determined. Search of the available sequence databases suggested that, besides the egt gene, this region contains 5 ORFs similar to the baculovirus genes gp37 (fusolin), to ptp2 and to ORFs 28, 29, and 30 of Spodoptera exigua multicapsid nucleopolyhedrovirus. Both the phylogenetic analysis of the egt genes and the gene order of the region that flanks the egt gene indicated that SfMNPV is closely related to the baculoviruses that infects S. exigua and Mamestra configurata.