Giving Oncolytic Vaccinia Virus More BiTE
Authors: Steven M Albelda, Steve H Thorne
Journal: Molecular Therapy (2014)

Two of the most exciting new areas of cancer therapy are the use of oncolytic viruses and immune-stimulating monoclonal antibodies. Each approach has shown great promise in animal models and some successes in early clinical trials, but there remain significant barriers to highly effective therapy. To date, the two technologies have been developed independently of each other, for the most part. However, in this issue of Molecular Therapy, Yu et al.1Yu F Wang X Guo ZS Bartlett DL Gottschalk SM Song X-T T-cell engager-armed oncolytic vaccinia virus significantly enhances antitumor therapy.Mol Ther. 2014; 22: 102-111Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar describe experiments that combine the strengths of both approaches by creating an oncolytic vaccinia virus (VV) that has the ability to home to tumors where it can replicate and induce tumor lysis but also has the ability to secrete a bispecific T-cell engager (BiTE) that can bind T cells to tumor cells and additionally induce immune-mediated tumor cell destruction (Figure 1). Oncolytic viruses, which are capable of selective or preferential replication within tumor cells, have been studied intensively over the past 15 years.2Eager RM Nemunaitis J Clinical development directions in oncolytic viral therapy.Cancer Gene Ther. 2011; 18: 305-317Crossref PubMed Scopus (93) Google Scholar A large variety of such viruses have been studied in both the preclinical and clinical settings. Both naturally occurring viruses with intrinsic tumor selectivity (such as reovirus, vesicular stomatitis virus, and Newcastle disease virus) and viruses genetically engineered to enhance tumor specificity (such as adenovirus, herpes simplex virus, measles virus, polio virus, and VV) are being developed.3Guo ZS Thorne SH Bartlett DL Oncolytic virotherapy: molecular targets in tumor-selective replication and carrier cell-mediated delivery of oncolytic viruses.Biochim Biophys Acta. 2008; 1785: 217-231PubMed Google Scholar One of the most promising approaches has been the use of oncolytic VVs,4Kirn DH Thorne SH Targeted and armed oncolytic poxviruses: a novel multi-mechanistic therapeutic class for cancer.Nat Rev Cancer. 2009; 9: 64-71Crossref PubMed Scopus (315) Google Scholar as this viral backbone is capable of rapid replication and spread that leads to profound localized tissue damage, and it has evolved to disseminate through the bloodstream and to induce a targeted immune response. Its safety was demonstrated during the smallpox-eradication program, and strains highly selective for tumor replication have been reported.5McCart JA Ward JM Lee J Hu Y Alexander HR Libutti SK et al.Systemic cancer therapy with a tumor-selective vaccinia virus mutant lacking thymidine kinase and vaccinia growth factor genes.Cancer Res. 2001; 61: 8751-8757PubMed Google Scholar,6Kirn DH Wang Y Le Boeuf F Bell J Thorne SH Targeting of interferon-beta to produce a specific, multi-mechanistic oncolytic vaccinia virus.PLoS Med. 2007; 4: e353Crossref PubMed Scopus (158) Google Scholar This viral backbone is the area of focus of the paper by Yu et al.1Yu F Wang X Guo ZS Bartlett DL Gottschalk SM Song X-T T-cell engager-armed oncolytic vaccinia virus significantly enhances antitumor therapy.Mol Ther. 2014; 22: 102-111Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar An important determinant in the success of any oncolytic virus is its interaction with the immune system. Not unexpectedly, injection of a virus induces strong innate and acquired immune responses with the goal of eradicating the virus. This immunologically mediated destruction of the virus and virally infected cells limits the ability of the virus to persist and spread, thus restricting its oncolytic potential and antitumor efficacy. It is therefore not surprising that the immune system was initially seen as a barrier that needed to be overcome in oncolytic viral therapy. However, it has more recently become clear that the relationship of the immune system to oncolytic virus therapy is far more complex.7Prestwich RJ Ilett EJ Errington F Diaz RM Steele LP Kottke T et al.Immune-mediated antitumor activity of reovirus is required for therapy and is independent of direct viral oncolysis and replication.Clin Cancer Res. 2009; 15: 4374-4381Crossref PubMed Scopus (129) Google Scholar The key observation was that destruction of tumor cells in a highly immunogenic environment saturated with high levels of exogenous (viral) and endogenous danger signals was an ideal environment in which to generate and attract polyclonal antitumor CD8+ T cells. Further experimentation has confirmed that this induction of antitumor immune responses is important, and in some cases is the most important mechanism by which oncolytic viruses eliminate tumors.8Wang LC Lynn RC Cheng G Alexander E Kapoor V Moon EK et al.Treating tumors with a vaccinia virus expressing IFNβ illustrates the complex relationships between oncolytic ability and immunogenicity.Mol Ther. 2012; 20: 736-748Abstract Full Text Full Text PDF PubMed Scopus (34) Google Scholar Finding the optimal balance between induction of antitumor immune responses and minimal inhibition of antiviral responses has thus become a goal of the field. Some of the most promising results in this area have come from studies using modified VVs.4Kirn DH Thorne SH Targeted and armed oncolytic poxviruses: a novel multi-mechanistic therapeutic class for cancer.Nat Rev Cancer. 2009; 9: 64-71Crossref PubMed Scopus (315) Google Scholar VV has been extensively used in immunotherapy studies. It has a large cloning capacity for cytokine or antigen expression, a long track record of safety when used as a vaccine, and a well-defined immune response. As such, a variety of transgenes designed to enhance the antitumor immune response induced by oncolytic VV have been examined. The most successful approach to date has been insertion of the gene encoding the immune-activating cytokine granulocyte–macrophage colony-stimulating factor (GM-CSF).9Kim JH Oh JY Park BH Lee DE Kim JS Park HE et al.Systemic armed oncolytic and immunologic therapy for cancer with JX-594, a targeted poxvirus expressing GM-CSF.Mol Ther. 2006; 14: 361-370Abstract Full Text Full Text PDF PubMed Scopus (227) Google Scholar Secretion of GM-CSF by tumor cells has long been known to reduce tumor formation in mice,10Dranoff G Jaffee E Lazenby A Golumbek P Levitsky H Brose K et al.Vaccination with irradiated tumor cells engineered to secrete murine granulocyte-macrophage colony-stimulating factor stimulates potent, specific, and long-lasting anti-tumor immunity.Proc Natl Acad Sci USA. 1993; 90: 3539-3543Crossref PubMed Scopus (2652) Google Scholar probably through its ability to attract and activate dendritic cells. Genetic incorporation of GM-CSF into irradiated tumor cells has been used in the vaccine setting and is being tested in clinical trials with some success.11DeFrancesco L Landmark approval for Dendreon's cancer vaccine.Nat Biotechnol. 2010; 28: 531-532Crossref PubMed Scopus (40) Google Scholar With the same rationale, the GM-CSF gene was engineered into a modified VV and developed as an oncolytic agent (JX-594).9Kim JH Oh JY Park BH Lee DE Kim JS Park HE et al.Systemic armed oncolytic and immunologic therapy for cancer with JX-594, a targeted poxvirus expressing GM-CSF.Mol Ther. 2006; 14: 361-370Abstract Full Text Full Text PDF PubMed Scopus (227) Google Scholar JX-594 has moved to clinical trials, where it has shown potential in hepatocellular carcinomas12Heo J Reid T Ruo L Breitbach CJ Rose S Bloomston M et al.Randomized dose-finding clinical trial of oncolytic immunotherapeutic vaccinia JX-594 in liver cancer.Nat Med. 2013; 19: 329-336Crossref PubMed Scopus (534) Google Scholar and is now in phase III clinical testing. Other immune-activating cytokines and chemokines—including interferon-β (refs. 6Kirn DH Wang Y Le Boeuf F Bell J Thorne SH Targeting of interferon-beta to produce a specific, multi-mechanistic oncolytic vaccinia virus.PLoS Med. 2007; 4: e353Crossref PubMed Scopus (158) Google Scholar, 8Wang LC Lynn RC Cheng G Alexander E Kapoor V Moon EK et al.Treating tumors with a vaccinia virus expressing IFNβ illustrates the complex relationships between oncolytic ability and immunogenicity.Mol Ther. 2012; 20: 736-748Abstract Full Text Full Text PDF PubMed Scopus (34) Google Scholar) and CCL5 (ref. 13Li J O'Malley M Urban J Sampath P Guo ZS Kalinski P et al.Chemokine expression from oncolytic vaccinia virus enhances vaccine therapies of cancer.Mol Ther. 2011; 19: 650-657Abstract Full Text Full Text PDF PubMed Scopus (105) Google Scholar)—have also been introduced into VVs and have shown enhanced efficacy in preclinical models. However, because mediators such as GM-CSF, IFN-β, and CCL5 are generalized activators of the immune system and have relatively little direct antitumor effect, they have at least three potential limitations. First, their efficacy is dependent on the development or enhancement of endogenous antitumor responses (which may be absent or compromised in many tumor patients). Second, the inflammation they produce may enhance viral clearance. Third, these activators may potentially trigger protumor immune effects (e.g., GM-CSF has been shown to induce the production of myeloid-derived suppressor cells). It would therefore be an advance if a VV cargo could be developed that would specifically induce antitumor immune responses without stimulating innate or acquired antiviral immune responses. It is in this context that Yu et al.1Yu F Wang X Guo ZS Bartlett DL Gottschalk SM Song X-T T-cell engager-armed oncolytic vaccinia virus significantly enhances antitumor therapy.Mol Ther. 2014; 22: 102-111Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar report experiments describing a VV designed to secrete a BiTE antibody (summarized in Figure 1). BiTEs consist of a tumor-targeting single-chain variable antibody fragment (scFv) translated in tandem with another scFv directed against the T-cell activation ligand, CD3 (ref. 14Choi BD Cai M Bigner DD Mehta AI Kuan CT Sampson JH Bispecific antibodies engage T cells for antitumor immunotherapy.Expert Opin Biol Ther. 2011; 11: 843-853Crossref PubMed Scopus (69) Google Scholar). These bispecific antibodies bind to both tumor-specific epitopes and the T-cell receptor (TCR) on nearby lymphocytes. Crosslinking of the TCR initiates an immuno­logical synapse and triggers T-cell activation with subsequent release of proteases and cytokines that kill tumor cells. The T cells do not need to be preactivated, costimulation does not appear to be necessary, and serial lysis by engaged T cells has been observed. They are thus extremely potent, with in vitro activity in the picomolar range. BiTEs directed against CD19 (to target B-cell malignancies) have shown success in clinical trials.15Bargou R Leo E Zugmaier G Klinger M Goebeler M Knop S et al.Tumor regression in cancer patients by very low doses of a T cell-engaging antibody.Science. 2008; 321: 974-977Crossref PubMed Scopus (822) Google Scholar Other targets on tumor cells being explored include EpCAM, EGFRvIII, and EphA2 (refs. 14Choi BD Cai M Bigner DD Mehta AI Kuan CT Sampson JH Bispecific antibodies engage T cells for antitumor immunotherapy.Expert Opin Biol Ther. 2011; 11: 843-853Crossref PubMed Scopus (69) Google Scholar, 16Hammond SA Lutterbuese R Roff S Lutterbuese P Schlereth B Bruckheimer E et al.Selective targeting and potent control of tumor growth using an EphA2/CD3-bispecific single-chain antibody construct.Cancer Res. 2007; 67: 3927-3935Crossref PubMed Scopus (74) Google Scholar). Despite great promise, however, there are still significant challenges in the administration of BiTes, including (i) the need for continuous intravenous administration by infusion pumps, (ii) difficulties in the manufacturing of these biological agents from mammalian producer cells, and (iii) troublesome systemic side effects (e.g., flulike symptoms and some worrisome central nervous system effects). Yu et al.1Yu F Wang X Guo ZS Bartlett DL Gottschalk SM Song X-T T-cell engager-armed oncolytic vaccinia virus significantly enhances antitumor therapy.Mol Ther. 2014; 22: 102-111Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar hypothesized that an oncolytic VV encoding a BiTE targeting the tumor surface antigen EphA2 would result in enhanced antitumor efficacy due to oncolysis plus bystander immune killing. They therefore generated a double-deleted VV (lacking the TK and VGF genes) expressing an EphA2-CD3 BiTE or control green fluorescent protein (GFP) by cloning these genes into the TK gene locus under the control of the F17R late promoter to allow sufficient viral replication before T-cell activation. Their tumor target was the human A549 lung cancer cell line, which expresses EphA2. They found that the BiTE did not interfere with either viral replication or the ability of the virus to induce A549 tumor cell lysis with no T cells present. Supernatant from the VV-BiTE–infected cells, but not VV-GFP–infected cells, was able to activate the T cells and induce T cell–mediated killing of uninfected A549 cells. Importantly, A549 lung cancer cells infected with the VV-BiTE in the presence of human T cells were killed more efficiently than A549 cells infected with the VV-GFP virus, establishing the bystander effect. Finally, the effect of the VV-BiTE virus was studied in two animal models. First, A549 cells were mixed with human peripheral blood mononuclear cells (PBMCs) and injected into the flanks of immunodeficient mice. Second, A549 cells were injected intravenously (to form lung tumors). Seven days later, the mice were injected intravenously with PBMCs alone, VVs alone, or an admixture of PBMCs with VVs. In both cases, systemic administration of VV-BiTEs, but not VV-GFP, resulted in significantly increased survival compared with controls. There are some limitations to this study. The authors studied only one BiTE target and one tumor cell target. Also, the animal models used were artificial in that PBMCs were coinjected with either the tumor cells or the virus. A more realistic model would require VV-BiTE treatment in mice in which the tumors are infiltrated with human T cells. It is also not clear how effectively BiTEs will activate tumor-infiltrating lymphocytes that show profound hypofunction and proximal defects in their TCR signaling machinery.17Vazquez-Cintron EJ Monu NR Frey AB Tumor-induced disruption of proximal TCR-mediated signal transduction in tumor-infiltration CD8+ lymphocytes inactivates antitumor effector phase.J Immunol. 2010; 185: 7133-7140Crossref PubMed Scopus (38) Google Scholar However, even given these limitations, the findings by Yu et al.1Yu F Wang X Guo ZS Bartlett DL Gottschalk SM Song X-T T-cell engager-armed oncolytic vaccinia virus significantly enhances antitumor therapy.Mol Ther. 2014; 22: 102-111Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar show the promise of combining two promising and previously separate new forms of anticancer therapy. By using the VV as a "carrier" and localized expression factory, it should be possible to overcome some of the limitations of system­ically injected BiTEs by ensuring very high local concentrations, which should minimize systemic side effects and may enhance efficacy. On the other hand, because the BiTEs should induce only minimal antiviral effects (perhaps due to local cytokine production), the oncolytic effects of the VV should be undiminished and perhaps allow persistence. Further exploration of the merger of these two new promising antitumor technologies seems warranted.