Jonathan Pol

HDAC Inhibition Suppresses Primary Immune Responses, Enhances Secondary Immune Responses, and Abrogates Autoimmunity During Tumor Immunotherapy

Histone deacetylase inhibitors (HDACi) can modulate innate antiviral responses and render tumors more susceptible to oncolytic viruses (OVs); however, their effects on adaptive immunity in this context are largely unknown. Our present study reveals an unexpected property of the HDACi MS-275 that enhances viral vector-induced lymphopenia leading to selective depletion of bystander lymphocytes and regulatory T cells while allowing expansion of antigen-specific secondary responses. Coadministration of vaccine plus drug during the boosting phase focuses the immune response on the tumor by suppressing the primary immune response against the vaccine vector and enhancing the secondary response against the tumor antigen. Furthermore, improvement of T cell functionality was evident suggesting that MS-275 can orchestrate a complex array of effects that synergize immunotherapy and viral oncolysis. Surprisingly, while MS-275 dramatically enhanced efficacy, it suppressed autoimmune pathology, profoundly improving the therapeutic index.

Maraba Virus as a Potent Oncolytic Vaccine Vector

The rhabdovirus Maraba has recently been characterized as a potent oncolytic virus. In the present study, we engineered an attenuated Maraba strain, defined as MG1, to express a melanoma-associated tumor antigen. Its ability to mount an antitumor immunity was evaluated in tumor-free and melanoma tumor-bearing mice. Alone, the MG1 vaccine appeared insufficient to prime detectable adaptive immunity against the tumor antigen. However, when used as a boosting vector in a heterologous prime-boost regimen, MG1 vaccine rapidly generated strong antigen-specific T-cell immune responses. Once applied for treating syngeneic murine melanoma tumors, our oncolytic prime-boost vaccination protocol involving Maraba MG1 dramatically extended median survival and allowed complete remission in more than 20% of the animals treated. This work describes Maraba virus MG1 as a potent vaccine vector for cancer immunotherapy displaying both oncolytic activity and a remarkable ability to boost adaptive antitumor immunity.

Immunogenic HSV-mediated oncolysis shapes the antitumor immune response and contributes to therapeutic efficacy.

Within the oncolytic virus field, the extent of virus replication that is essential for immune stimulation to control tumor growth remains unresolved. Using infected cell protein 0 (ICP0)-defective oncolytic Herpes simplex virus type 1 (HSV-1) and HSV-2 viruses (dICP0 and dNLS) that show differences in their in vitro replication and cytotoxicity, we investigated the inherent features of oncolytic HSV viruses that are required for potent antitumor activity. In vitro, the HSV-2 vectors showed rapid cytotoxicity despite lower viral burst sizes compared to HSV-1 vectors. In vivo, although both of the dICP0 vectors initially replicated to a similar level, HSV-1 dICP0 was rapidly cleared from the tumors. In spite of this rapid clearance, HSV-1 dICP0 treatment conferred significant survival benefit. HSV-1 dICP0-treated tumors showed significantly higher levels of danger-associated molecular patterns that correlated with higher numbers of antigen-presenting cells within the tumor and increased antigen-specific CD8+ T-cell levels in the peripheral blood. This study suggests that, at least in the context of oncolytic HSV, the initial stages of immunogenic virus replication leading to activation of antitumor immunity are more important than persistence of a replicating virus within the tumor. This knowledge provides important insight for the design of therapeutically successful oncolytic viruses.

Combining oncolytic HSV-1 with immunogenic cell death-inducing drug mitoxantrone breaks cancer immune tolerance and improves therapeutic efficacy.

Workenhe and colleagues show that the combination regimen of oncolytic virus with mitoxantrone significantly increases the efficacy of either treatment alone by enhancing the immunogenicity of and breaking immune tolerance against tumor-associated antigens, an Achilles heel of current cancer therapy. Although antitumor activity of herpes simplex virus 1 (HSV-1) ICP0 null oncolytic vectors has been validated in murine breast cancer models, oncolytic virus treatment alone is insufficient to break immune tolerance. Thus, we investigated enhancing efficacy through combination therapy with the immunogenic cell death–inducing chemotherapeutic drug, mitoxantrone. Despite a lack of enhanced cytotoxicity in vitro, HSV-1 ICP0 null oncolytic virus KM100 with 5 μmol/L mitoxantrone provided significant survival benefit to BALB/c mice bearing Her2/neu TUBO-derived tumors. This protection was mediated by increased intratumoral infiltration of neutrophils and tumor antigen-specific CD8+ T cells. Depletion studies verified that CD8-, CD4-, and Ly6G-expressing cells are essential for enhanced efficacy of the combination therapy. Moreover, the addition of mitoxantrone to KM100 oncolytic virus treatment broke immune tolerance in BALB-neuT mice bearing TUBO-derived tumors. This study suggests that oncolytic viruses in combination with immunogenic cell death–inducing chemotherapeutics enhance the immunogenicity of the tumor-associated antigens, breaking immunologic tolerance established toward these antigens. Cancer Immunol Res; 1(5); 309–19. ©2013 AACR.

Oncolytic vesicular stomatitis virus quantitatively and qualitatively improves primary CD8 + T-cell responses to anticancer vaccines

The ability of heterologous prime-boost vaccination to elicit robust CD8+ T cell responses has been well documented. In contrast, relatively little is known about how this immunotherapeutic strategy impacts the functional qualities of expanded T cells in the course of effector and memory responses. Using vesicular stomatitis virus (VSV) as a boosting vector in mice, we demonstrate that a massive secondary expansion of CD8+ T cells can be achieved shortly after priming with recombinant adenoviral vectors. Importantly, VSV-boosted CD8+ T cells were more potent than those primed by adenoviruses only, as measured by cytokine production, granzyme B expression, and functional avidity. Upon adoptive transfer, equivalent numbers of VSV-expanded CD8+ T cells were more effective (on a per-cell basis) in mediating antitumor and antiviral immunity than T cells only primed with adenoviruses. Furthermore, VSV boosting accelerated the progression of expanded CD8+ T lymphocytes to a central memory phenotype, thereby altering the effector memory profile typically associated with adenoviral vaccination. Finally, the functional superiority of VSV-expanded T cells remained evident 100 d after boosting, suggesting that VSV-driven immunological responses are of sufficient duration for therapeutic applications. Our data strongly support the choice of VSV as a boosting vector in prime-boost vaccination strategies, enabling a rapid amplification of CD8+ T cells and improving the quality of expanded T cells during both early and late immunological responses.

S6K-STING interaction regulates cytosolic DNA–mediated activation of the transcription factor IRF3

Cytosolic DNA–mediated activation of the transcription factor IRF3 is a key event in host antiviral responses. Here we found that infection with DNA viruses induced interaction of the metabolic checkpoint kinase mTOR downstream effector and kinase S6K1 and the signaling adaptor STING in a manner dependent on the DNA sensor cGAS. We further demonstrated that the kinase domain, but not the kinase function, of S6K1 was required for the S6K1-STING interaction and that the TBK1 critically promoted this process. The formation of a tripartite S6K1-STING-TBK1 complex was necessary for the activation of IRF3, and disruption of this signaling axis impaired the early-phase expression of IRF3 target genes and the induction of T cell responses and mucosal antiviral immunity. Thus, our results have uncovered a fundamental regulatory mechanism for the activation of IRF3 in the cytosolic DNA pathway.

Oncolytic viruses: a step into cancer immunotherapy

Correspondence: Brian Lichty McMaster Immunology Research Centre, Department of Pathology and Molecular Medicine, McMaster University, 1280 Main Street West, Hamilton, Ontario, Canada L8S 4L8 Tel +1 905 525 9140 ext 22478 Fax +1 905 522 6750 Email lichtyb@mcmaster.ca Abstract: Oncolytic virotherapy is currently under investigation in phase I–III clinical trials for approval as a new cancer treatment. Oncolytic viruses (OVs) selectively infect, replicate in, and kill tumor cells. For a long time, the therapeutic efficacy was thought to depend on the direct viral oncolysis (virocentric view). The host immune system was considered as a brake that impaired virus delivery and spread. Attention was paid primarily to approaches enhancing virus tumor selectivity and cytotoxicity and/or that limited antiviral responses. Thinking has changed over the past few years with the discovery that OV therapy was also inducing indirect oncolysis mechanisms. Among them, induction of an antitumor immunity following OV injection appeared to be a key factor for an efficient therapeutic activity (immunocentric view). Indeed, tumor-specific immune cells persist post-therapy and can search and destroy any tumor cells that escape the OVs, and thus immune memory may prevent relapse of the disease. Various strategies, which are summarized in this manuscript, have been developed to enhance the efficacy of OV therapy with a focus on its immunotherapeutic aspects. These include genetic engineering and combination with existing cancer treatments. Several are currently being evaluated in human patients and already display promising efficacy.

Privileged Antigen Presentation in Splenic B Cell Follicles Maximizes T Cell Responses in Prime-Boost Vaccination

Effector T cells (TEFF) are a barrier to booster vaccination because they can rapidly kill Ag-bearing APCs before memory T cells are engaged. We report in this study that i.v. delivery of rhabdoviral vectors leads to direct infection of follicular B cells in the spleen, where the earliest evidence of secondary T cell responses was observed. This allows booster immunizations to rapidly expand CD8+ central memory T cells (TCM) during the acute phase of the primary response that is dominated by TEFF. Interestingly, although the ablation of B cells before boosting with rhabdoviral vectors diminishes the expansion of memory T cells, B cells do not present Ags directly. Instead, depletion of CD11c+ dendritic cells abrogates secondary T cell expansion, suggesting that virus-infected follicular B cells may function as an Ag source for local DCs to subsequently capture and present the Ag. Because TCM are located within B cell follicles in the spleen whereas TEFF cannot traffic through follicular regions, Ag production and presentation by follicular APCs represent a unique mechanism to secure engagement of TCM during an ongoing effector response. Our data offer insights into novel strategies for rapid expansion of CD8+ T cells using prime-boost vaccines by targeting privileged sites for Ag presentation.

Immunogenic stress and death of cancer cells: Contribution of antigenicity vs adjuvanticity to immunosurveillance

Cancer cells are subjected to constant selection by the immune system, meaning that tumors that become clinically manifest have managed to subvert or hide from immunosurveillance. Immune control can be facilitated by induction of autophagy, as well as by polyploidization of cancer cells. While autophagy causes the release of ATP, a chemotactic signal for myeloid cells, polyploidization can trigger endoplasmic reticulum stress with consequent exposure of the “eat‐me” signal calreticulin on the cell surface, thereby facilitating the transfer of tumor antigens into dendritic cells. Hence, both autophagy and polyploidization cause the emission of adjuvant signals that ultimately elicit immune control by CD8+ T lymphocytes. We investigated the possibility that autophagy and polyploidization might also affect the antigenicity of cancer cells by altering the immunopeptidome. Mass spectrometry led to the identification of peptides that were presented on major histocompatibility complex (MHC) class I molecules in an autophagy‐dependent fashion or that were specifically exposed on the surface of polyploid cells, yet lost upon passage of such cells through immunocompetent (but not immunodeficient) mice. However, the preferential recognition of autophagy‐competent and polyploid cells by the innate and cellular immune systems did not correlate with the preferential recognition of such peptides in vivo. Moreover, vaccination with such peptides was unable to elicit tumor growth‐inhibitory responses in vivo. We conclude that autophagy and polyploidy increase the immunogenicity of cancer cells mostly by affecting their adjuvanticity rather than their antigenicity.

Panorama From the Oncolytic Virotherapy Summit

In June, the historical heart of Quebec City welcomed the 7th International Summit on Oncolytic Viral Therapeutics for an excellent opportunity to witness the growth of the field, in both size and maturity. Efforts are being made to better characterize interactions between oncolytic viruses (OVs) and host components, both inside and outside the tumor bed, with their relative contribution to the overall therapeutic efficacy. New approaches in improving OV delivery, tumor selectivity, spreading, and killing continue to flourish. Clinical evaluations, ongoing and upcoming, have never been this numerous. The present review draws on some of the unpublished and recently published findings reported during the meeting.