Combination immunotherapy with vaccine and oncolytic HSV virotherapy is time dependent.

PURPOSE: Oncolytic virotherapy or immunovirotherapy is a strategy that utilizes viruses to selectively infect and kill tumor cells while also stimulating an immune response against the tumor. Early clinical trials in both pediatric and adult patients using oncolytic herpes simplex viruses (oHSVs) have demonstrated safety and promising efficacy; however, combinatorial strategies designed to enhance oncolysis while also promoting durable T cell responses for sustaining disease remission are likely required. We hypothesized that combining the direct tumor cell killing and innate immune stimulation by oHSV with a vaccine that promotes T cell mediated immunity may lead to more durable tumor regression. EXPERIMENTAL DESIGN: To this end, we investigated the preclinical efficacy and potential synergy of combining oHSV with a self-assembling nanoparticle vaccine co-delivering peptide antigens and Toll-like receptor-7 and -8 agonists (TLR-7/8a) (referred to as SNAPvax™), that induces robust tumor specific T cell immunity. We then assessed how timing of the treatments (i.e., vaccine before or after oHSV) impacts T cell responses, viral replication, and preclinical efficacy. RESULTS: The sequence of treatments was critical, as survival was significantly enhanced when the SNAPvax™ vaccine was given prior to oHSV. Increased clinical efficacy was associated with reduced tumour volume and increases in virus replication and tumor antigen specific CD8+ T cells. CONCLUSIONS: These findings substantiate the criticality of combination immunotherapy timing and provide preclinical support for combining SNAPvax with oHSV as a promising treatment approach for both pediatric and adult tumors.

Poly-IC enhances the effectiveness of cancer immunotherapy by promoting T cell tumor infiltration.

BACKGROUND: Immunotherapies, such as immune checkpoint inhibitors and adoptive cell therapies, have revolutionized cancer treatment and resulted in complete and durable responses in some patients. Unfortunately, most immunotherapy treated patients still fail to respond. Absence of T cell infiltration to the tumor site is one of the major obstacles limiting immunotherapy efficacy against solid tumors. Thus, the development of strategies that enhance T cell infiltration and broaden the antitumor efficacy of immunotherapies is greatly needed. METHODS: We used mouse tumor models, genetically deficient mice and vascular endothelial cells (VECs) to study the requirements for T cell infiltration into tumors. RESULTS: A specific formulation of poly-IC, containing poly-lysine and carboxymethylcellulose (PICLC) facilitated the traffic and infiltration of effector CD8 T cells into the tumors that reduced tumor growth. Surprisingly, intratumoral injection of PICLC was significantly less effective in inducing tumor T cell infiltration and controlling growth of tumors as compared with systemic (intravenous or intramuscular) administration. Systemically administered PICLC, but not poly-IC stimulated tumor VECs via the double-stranded RNA cytoplasmic sensor MDA5, resulting in enhanced adhesion molecule expression and the production of type I interferon (IFN-I) and T cell recruiting chemokines. Expression of IFNαß receptor in VECs was necessary to obtain the antitumor effects by PICLC and IFN-I was found to directly stimulate the secretion of T cell recruiting chemokines by VECs indicating that this cytokine-chemokine regulatory axis is crucial for recruiting effector T cells into the tumor parenchyma. Unexpectedly, these effects of PICLC were mostly observed in tumors and not in normal tissues. CONCLUSIONS: These findings have strong implications for the improvement of all types of T cell-based immunotherapies for solid cancers. We predict that systemic administration of PICLC will improve immune checkpoint inhibitor therapy, adoptive cell therapies and therapeutic cancer vaccines.

Sustained Persistence of IL2 Signaling Enhances the Antitumor Effect of Peptide Vaccines through T-cell Expansion and Preventing PD-1 Inhibition.

Peptide vaccines can be a successful and cost-effective way of generating T-cell responses against defined tumor antigens, especially when combined with immune adjuvants such as poly-IC. However, strong immune adjuvants can induce a collateral increase in numbers of irrelevant, nonspecific T cells, which limits the effectiveness of the peptide vaccines. Here, we report that providing prolonged IL2 signaling in the form of either IL2/anti-IL2 complexes or pegylated IL2 overcomes the competitive suppressive effect of irrelevant T cells, allowing the preferential expansion of antigen-specific T cells. In addition to increasing the number of tumor-reactive T cells, sustained IL2 enhanced the ability of T cells to resist PD-1-induced negative signals, increasing the therapeutic effectiveness of the vaccines against established tumors. This vaccination strategy using peptides and sustained IL2 could be taken into the clinic for the treatment of cancer. Cancer Immunol Res; 6(5); 617-27. ©2018 AACR.

Designing therapeutic cancer vaccines by mimicking viral infections.

The design of efficacious and cost-effective therapeutic vaccines against cancer remains both a research priority and a challenge. For more than a decade, our laboratory has been involved in the development of synthetic peptide-based anti-cancer therapeutic vaccines. We first dedicated our efforts in the identification and validation of peptide epitopes for both CD8 and CD4 T cells from tumor-associated antigens (TAAs). Because of suboptimal immune responses and lack of therapeutic benefit of peptide vaccines containing these epitopes, we have focused our recent efforts in optimizing peptide vaccinations in mouse tumor models using numerous TAA epitopes. In this focused research review, we describe how after taking lessons from the immune system's way of dealing with acute viral infections, we have designed peptide vaccination strategies capable of generating very high numbers of therapeutically effective CD8 T cells. We also discuss some of the remaining challenges to translate these findings into the clinical setting.

Hydrogen peroxide regulates osteopontin expression through activation of transcriptional and translational pathways.

Recent in vivo studies establish that osteopontin (OPN) expression is hydrogen peroxide (H2O2)-dependent. However, the mechanisms by which H2O2 increases OPN expression remain poorly defined. OPN protein expression increased in an unusual biphasic pattern in response to H2O2. To investigate whether these increases were mediated through transcriptional and/or translational regulation of OPN, smooth muscle cells stimulated with 50 µM H2O2 were used as an in vitro cell system. Early protein increases at 6 h were not preceded by increased mRNA, whereas later increases (18 h) were, suggesting multiple mechanisms of regulation by H2O2. Polyribosomal fractionation assays established that early increases (6 h) in OPN expression were due to increased translation. This increase in translation occurred through phosphorylation of 4E-BP1 at the reactive oxygen species-sensitive Ser-65, which allowed for release and activation of eukaryotic initiation factor eIF4E and subsequent OPN translation. This early increase (6 h) in OPN was blunted in cells expressing a phospho-deficient 4E-BP1 mutant. H2O2 stimulation increased rat OPN promoter activity at 8 and 18 h, and promoter truncation studies established that promoter region -2284 to -795 is crucial for H2O2-dependent OPN transcription. ChIP studies determined that H2O2-dependent transcription is mediated by the reactive oxygen species-sensitive transcription factors NF-κB and AP-1. In conclusion, H2O2 stimulates OPN expression in a unique biphasic pattern, where early increases are translational and late increases are transcriptional.

Reactive oxygen species regulate osteopontin expression in a murine model of postischemic neovascularization.

OBJECTIVE: Previous findings from our laboratory demonstrated that neovascularization was impaired in osteopontin (OPN) knockout animals. However, the mechanisms responsible for the regulation of OPN expression in the setting of ischemia remain undefined. Therefore, we sought to determine whether OPN is upregulated in response to ischemia and hypothesized that hydrogen peroxide (H(2)O(2)) is a critical component of the signaling mechanism by which OPN expression is upregulated in response to ischemia in vivo. METHODS AND RESULTS: To determine whether ischemic injury upregulates OPN, we used a murine model of hindlimb ischemia. Femoral artery ligation in C57BL/6 mice significantly increased OPN expression and H(2)O(2) production. Infusion of C57BL/6 mice with polyethylene glycol-catalase (10 000 U/kg per day) or the use of transgenic mice with smooth muscle cell-specific catalase overexpression blunted ischemia-induced OPN, suggesting ischemia-induced OPN expression is H(2)O(2)-dependent. Decreased H(2)O(2)-mediated OPN blunted reperfusion and collateral formation in vivo. In contrast, the overexpression of OPN using lentivirus restored neovascularization. CONCLUSIONS: Scavenging H(2)O(2) blocks ischemia-induced OPN expression, providing evidence that ischemia-induced OPN expression is H(2)O(2) dependent. Decreased OPN expression impaired neovascularization, whereas overexpression of OPN increased angiogenesis, supporting our hypothesis that OPN is a critical mediator of postischemic neovascularization and a potential novel therapeutic target for inducing new vessel growth.