Molecular analysis of human cancer cells infected by an oncolytic HSV-1 reveals multiple upregulated cellular genes and a role for SOCS1 in virus replication

Abstract

Oncolytic herpes simplex viruses (oHSVs) are promising anticancer therapeutics. We sought to characterize the functional genomic response of human cancer cells to oHSV infection using G207, an oHSV previously evaluated in a phase I trial. Five human malignant peripheral nerve sheath tumor cell lines, with differing sensitivity to oHSV, were infected with G207 for 6 h. Functional genomic analysis of virus-infected cells demonstrated large clusters of downregulated cellular mRNAs and smaller clusters of those upregulated, including 21 genes commonly upregulated in all five lines. Of these, 7 are known to be HSV-1 induced and 14 represent novel virus-regulated genes. Gene ontology analysis revealed that a majority of G207-upregulated genes are involved in Janus kinase/signal transducer and activator of transcription signaling, transcriptional regulation, nucleic acid metabolism, protein synthesis and apoptosis. Ingenuity networks highlighted nodes for AP-1 subunits and interferon signaling via STAT1, suppressor of cytokine signaling-1 (SOCS1), SOCS3 and RANTES. As biological confirmation, we found that virus-mediated upregulation of SOCS1 correlated with sensitivity to G207 and that depletion of SOCS1 impaired virus replication by >10-fold. Further characterization of roles provided by oHSV-induced cellular genes during virus replication may be utilized to predict oncolytic efficacy and to provide rational strategies for designing next-generation oncolytic viruses.

Figure 1

Transcriptomic effect of G207 infection in human MPNST cells. (a) Hierarchical clustering of G207-regulated genes at 6 h post-infection of human MPNST cell lines (S462, ST8814, 90-8, STS26 T, t265p21). Red signifies upregulated and blue signifies downregulated genes. (b) Comparison of gene expression changes in each cell line centric on the MPNST cell line listed on the y axis.

Figure 2

G207 infection induces upregulation of a subset of cellular genes. 343 probe sets were >2.5 × upregulated by G207 infection. Of these probe sets, the number (■) and percentage (□) commonly upregulated by G207 in two, three, four or five cell lines are shown. Inset: Venn diagram with fivefold symmetry illustrating 38 probe sets commonly upregulated in 5/5 human MPNST cell lines.

Figure 3

Ingenuity software analysis of G207-upregulated genes involved in immune and antiviral response.

Figure 4

Quantitative real-time PCR validation of G207-upregulated genes. MPNST cells were infected with G207 and cellular RNA was harvested at 6 h post-infection. qRT-PCR for SOCS1, STAT1, GADD45-β, c-FOS and eIF4A1 are shown (*P<0.05, **P<0.005, ***P<0.0005).

Figure 5

Depletion of suppressor of cytokine signaling 1 (SOCS1) impairs G207 replication in MPNST cells. (a) Western blot showing levels of SOCS1 protein in parental S462 cells and cells expressing control and SOCS1-targeted shRNAs. (b) Virus replication assay demonstrated that SOCS1-depleted S462 cells have impaired virus replication by >10-fold at 48 and 72 h, compared with parental S462 cells or S462 cells expressing the control shRNA (*P<0.001).