Immunotherapies for pediatric cancer: current landscape and future perspectives.

The advent of immunotherapy has revolutionized how we manage and treat cancer. While the majority of immunotherapy-related studies performed to date have focused on adult malignancies, a handful of these therapies have also recently found success within the pediatric space. In this review, we examine the immunotherapeutic agents that have achieved the approval of the US Food and Drug Administration for treating childhood cancers, highlighting their development, mechanisms of action, and the lessons learned from the seminal clinical trials that ultimately led to their approval. We also shine a spotlight on several emerging immunotherapeutic modalities that we believe are poised to have a positive impact on the treatment of pediatric malignancies in the near future.

Characterization of MHC Class I and ß-2-Microglobulin Expression in Pediatric Solid Malignancies to Guide Selection of Immune-Based Therapeutic Trials.

BACKGROUND: Over 10,000 US children are diagnosed with cancer yearly. Though outcomes have improved by optimizing conventional therapies, recent immunotherapeutic successes in adult cancers are emerging. Cytotoxic T lymphocytes (CTLs) are the primary executioners of adaptive antitumor immunity and require antigenic presentation in the context of major histocompatibility complex (MHC) class I and the associated ß-2-microglobulin (B2M). Loss of MHC I expression is a common immune escape mechanism in adult malignancies, but pediatric cancers have not been thoroughly characterized. The essential nature of MHC I expression in CTL-mediated cell death may dictate the success of immunotherapies, which rely on eliciting an adaptive response. PROCEDURE: We queried pediatric tumor microarray databases for MHC I and B2M gene expression. We detected MHC I in pediatric tumor cell lines by flow cytometry and characterized MHC I and B2M expression in patient samples by immunohistochemistry. To determine whether therapeutic approaches might enhance MHC I expression in selected models in vitro, we tested effects of exposure to IFN-γ and histone deacetylase inhibitors. RESULTS: Pediatric tumors overall, as well as samples within select individual tumor subtypes, exhibit wide ranges of MHC I and B2M gene and protein expression. For most cell lines tested, MHC I was inducible in vitro. CONCLUSIONS: MHC I and B2M expression vary among pediatric tumor types and should be evaluated as potential biomarkers, which might identify patients most likely to benefit from MHC I dependent immunotherapies. Modulation of MHC I expression may be a promising mechanism for enhancing MHC I dependent immunotherapeutic efficacy.

Treatment of medulloblastoma with oncolytic measles viruses expressing the angiogenesis inhibitors endostatin and angiostatin.

BACKGROUND: Medulloblastoma is the most common type of pediatric brain tumor. Although numerous factors influence patient survival rates, more than 30% of all cases will ultimately be refractory to conventional therapies. Current standards of care are also associated with significant morbidities, giving impetus for the development of new treatments. We have previously shown that oncolytic measles virotherapy is effective against medulloblastoma, leading to significant prolongation of survival and even cures in mouse xenograft models of localized and metastatic disease. Because medulloblastomas are known to be highly vascularized tumors, we reasoned that the addition of angiogenesis inhibitors could further enhance the efficacy of oncolytic measles virotherapy. Toward this end, we have engineered an oncolytic measles virus that express a fusion protein of endostatin and angiostatin, two endogenous and potent inhibitors of angiogenesis. METHODS: Oncolytic measles viruses encoding human and mouse variants of a secretable endostatin/angiostatin fusion protein were designed and rescued according to established protocols. These viruses, known as MV-hE:A and MV-mE:A respectively, were then evaluated for their anti-angiogenic potential and efficacy against medulloblastoma cell lines and orthotopic mouse models of localized disease. RESULTS: Medulloblastoma cells infected by MV-E:A readily secrete endostatin and angiostatin prior to lysis. The inclusion of the endostatin/angiostatin gene did not negatively impact the measles virus' cytotoxicity against medulloblastoma cells or alter its growth kinetics. Conditioned media obtained from these infected cells was capable of inhibiting multiple angiogenic factors in vitro, significantly reducing endothelial cell tube formation, viability and migration compared to conditioned media derived from cells infected by a control measles virus. Mice that were given a single intratumoral injection of MV-E:A likewise showed reduced numbers of tumor-associated blood vessels and a trend for increased survival compared to mice treated with the control virus. CONCLUSIONS: These data suggest that oncolytic measles viruses encoding anti-angiogenic proteins may have therapeutic benefit against medulloblastoma and support ongoing efforts to target angiogenesis in medulloblastoma.

The quest for effective immunotherapies against malignant peripheral nerve sheath tumors: Is there hope?

Immune-based therapies represent a new paradigm in the treatment of multiple cancers, where they have helped achieve durable and safe clinical responses in a growing subset of patients. While a wealth of information is available concerning the use of these agents in treating the more common malignancies, little has been reported about the use of immunotherapies against malignant peripheral nerve sheath tumors (MPNSTs), a rare form of soft tissue sarcoma that arises from the myelin sheaths that protect peripheral nerves. Surgical resection has been the mainstay of therapy in MPNSTs, but the recurrence rate is as high as 65%, and chemotherapy is generally ineffective. The immune contexture of MPNSTs, replete with macrophages and a varying degree of T cell infiltration, presents multiple opportunities to design meaningful therapeutic interventions. While preliminary results with macrophage-targeting strategies and oncolytic viruses are promising, identifying the subset of patients that respond to immune-based strategies will be a milestone. As part of our effort to help advance the use of immunotherapy for MPNSTs, here we describe recent insights regarding the immune contexture of MPNSTs, discuss emerging immune-based strategies, and provide a brief overview of potential biomarkers of response.

Treatment of medulloblastoma using an oncolytic measles virus encoding the thyroidal sodium iodide symporter shows enhanced efficacy with radioiodine.

BACKGROUND: Medulloblastoma is the most common malignant brain tumor of childhood. Although the clinical outcome for medulloblastoma patients has improved significantly, children afflicted with the disease frequently suffer from debilitating side effects related to the aggressive nature of currently available therapy. Alternative means for treating medulloblastoma are desperately needed. We have previously shown that oncolytic measles virus (MV) can selectively target and destroy medulloblastoma tumor cells in localized and disseminated models of the disease. MV-NIS, an oncolytic measles virus that encodes the human thyroidal sodium iodide symporter (NIS), has the potential to deliver targeted radiotherapy to the tumor site and promote a localized bystander effect above and beyond that achieved by MV alone. METHODS: We evaluated the efficacy of MV-NIS against medulloblastoma cells in vitro and examined their ability to incorporate radioiodine at various timepoints, finding peak uptake at 48 hours post infection. The effects of MV-NIS were also evaluated in mouse xenograft models of localized and disseminated medulloblastoma. Athymic nude mice were injected with D283med-Luc medulloblastoma cells in the caudate putamen (localized disease) or right lateral ventricle (disseminated disease) and subsequently treated with MV-NIS. Subsets of these mice were given a dose of 131I at 24, 48 or 72 hours later. RESULTS: MV-NIS treatment, both by itself and in combination with 131I, elicited tumor stabilization and regression in the treated mice and significantly extended their survival times. Mice given 131I were found to concentrate radioiodine at the site of their tumor implantations. In addition, mice with localized tumors that were given 131I either 24 or 48 hours after MV-NIS treatment exhibited a significant survival advantage over mice given MV-NIS alone. CONCLUSIONS: These data suggest MV-NIS plus radioiodine may be a potentially useful therapy for the treatment of medulloblastoma.

Leveraging gene therapy to achieve long-term continuous or controllable expression of biotherapeutics.

T cells redirected to cancer cells either via a chimeric antigen receptor (CAR-T) or a bispecific molecule have been breakthrough technologies; however, CAR-T cells require individualized manufacturing and bispecifics generally require continuous infusions. We created an off-the-shelf, single-dose solution for achieving prolonged systemic serum levels of protein immunotherapeutics via adeno-associated virus (AAV) gene transfer. We demonstrate proof of principle in a CD19+ lymphoma xenograft model using a single intravenous dose of AAV expressing a secreted version of blinatumomab, which could serve as a universal alternative for CD19 CAR-T cell therapy. In addition, we created an inducible version using an exon skipping strategy and achieved repeated, on-demand expression up to at least 36 weeks after AAV injection. Our system could be considered for short-term and/or repeated expression of other transgenes of interest for noncancer applications.

Endogenous retrovirus envelope as a tumor-associated immunotherapeutic target in murine osteosarcoma.

Osteosarcoma remains one of the deadliest cancers in pediatrics and young adults. We administered two types of immunotherapies, oncolytic virotherapy and immune checkpoint inhibition, to two murine osteosarcoma models and observed divergent results. Mice bearing F420 showed no response, whereas those with K7M2 showed prolonged survival in response to combination therapy. K7M2 had higher expression of immune-related genes and higher baseline immune cell infiltrates, but there were no significant differences in tumor mutational burden or predicted MHC class I binding of nonsynonymous mutations. Instead, we found several mouse endogenous retrovirus sequences highly expressed in K7M2 compared with F420. T cell tetramer staining for one of them, gp70, was detected in mice with K7M2 but not F420, suggesting that endogenous retrovirus proteins are targets for the anti-tumor immune reaction. Given prior observations of endogenous retrovirus expression in human osteosarcomas, our findings may be translatable to human disease.

New curcumin analogues exhibit enhanced growth-suppressive activity and inhibit AKT and signal transducer and activator of transcription 3 phosphorylation in breast and prostate cancer cells.

Curcumin, the active component of turmeric, has been shown to protect against carcinogenesis and prevent tumor development in cancer. To enhance its potency, we tested the efficacy of synthetic curcumin analogues, known as FLLL11 and FLLL12, in cancer cells. We examined the impact of FLLL11 and FLLL12 on cell viability in eight different breast and prostate cancer cell lines. FLLL11 and FLLL12 (IC(50) values 0.3-5.7 and 0.3-3.8 micromol/L, respectively) were substantially more potent than curcumin (IC(50) values between 14.4-50 micromol/L). FLLL11 and FLLL12 were also found to inhibit AKT phosphorylation and downregulate the expression of HER2/neu. In addition, we demonstrate for the first time that FLLL11 and FLLL12 inhibit phosphorylation of signal transducer and activator of transcription (STAT) 3, an oncogene frequently found to be persistently active in many cancer types. The inhibition of STAT3 signaling was confirmed by the inhibition of STAT3 DNA binding and STAT3 transcriptional activity. Furthermore, FLLL11 and FLLL12 were more effective than curcumin in inhibiting cell migration and colony formation in soft agar as well as inducing apoptosis in cancer cells. These results indicate that FLLL11 and FLLL12 exhibit more potent activities than curcumin on the inhibition of STAT3, AKT, and HER-2/neu, as well as inhibit cancer cell growth and migration, and may thus have translational potential as chemopreventive or therapeutic agents for breast and prostate cancers.

Dietary agent, benzyl isothiocyanate inhibits signal transducer and activator of transcription 3 phosphorylation and collaborates with sulforaphane in the growth suppression of PANC-1 cancer cells.

The Signal Transducer and Activator of Transcription (STAT) proteins comprise a family of latent transcription factors with diverse functions. STAT3 has well established roles in cell proliferation, growth and survival, and its persistent activation has been detected with high frequency in many human cancers. As constitutive activation of STAT3 appears to be vital for the continued survival of these cancerous cells, it has emerged as an attractive target for chemotherapeutics. We examined whether the inhibitory activities of bioactive compounds from cruciferous vegetables, such as Benzyl isothiocyanate (BITC) and sulforaphane, extended to STAT3 activation in PANC-1 human pancreatic cancer cells. BITC and sulforaphane were both capable of inhibiting cell viability and inducing apoptosis in PANC-1. Sulforaphane had minimal effect on the direct inhibition of STAT3 tyrosine phosphorylation, however, suggesting its inhibitory activities are most likely STAT3-independent. Conversely, BITC was shown to inhibit the tyrosine phosphorylation of STAT3, but not the phosphorylation of ERK1/2, MAPK and p70S6 kinase. These results suggest that STAT3 may be one of the targets of BITC-mediated inhibition of cell viability in PANC-1 cancer cells. In addition, we show that BITC can prevent the induction of STAT3 activation by Interleukin-6 in MDA-MB-453 breast cancer cells. Furthermore, combinations of BITC and sulforaphane inhibited cell viability and STAT3 phosphorylation more dramatically than either agent alone. These findings suggest that the combination of the dietary agents BITC and sulforaphane has potent inhibitory activity in pancreatic cancer cells and that they may have translational potential as chemopreventative or therapeutic agents.

New structural analogues of curcumin exhibit potent growth suppressive activity in human colorectal carcinoma cells.

BACKGROUND: Colorectal carcinoma is one of the major causes of morbidity and mortality in the Western World. Novel therapeutic approaches are needed for colorectal carcinoma. Curcumin, the active component and yellow pigment of turmeric, has been reported to have several anti-cancer activities including anti-proliferation, anti-invasion, and anti-angiogenesis. Clinical trials have suggested that curcumin may serve as a potential preventive or therapeutic agent for colorectal cancer. METHODS: We compared the inhibitory effects of curcumin and novel structural analogues, GO-Y030, FLLL-11, and FLLL-12, in three independent human colorectal cancer cell lines, SW480, HT-29, and HCT116. MTT cell viability assay was used to examine the cell viability/proliferation and western blots were used to determine the level of PARP cleavages. Half-Maximal inhibitory concentrations (IC50) were calculated using Sigma Plot 9.0 software. RESULTS: Curcumin inhibited cell viability in all three of the human colorectal cancer cell lines studied with IC50 values ranging between 10.26 microM and 13.31 microM. GO-Y030, FLLL-11, and FLLL-12 were more potent than curcumin in the inhibition of cell viability in these three human colorectal cancer cell lines with IC50 values ranging between 0.51 microM and 4.48 microM. In addition, FLLL-11 and FLLL-12 exhibit low toxicity to WI-38 normal human lung fibroblasts with an IC-50 value greater than 1,000 microM. GO-Y030, FLLL-11, and FLLL-12 are also more potent than curcumin in the induction of apoptosis, as evidenced by cleaved PARP and cleaved caspase-3 in all three human colorectal cancer cell lines studied. CONCLUSION: The results indicate that the three curcumin analogues studied exhibit more potent inhibitory activity than curcumin in human colorectal cancer cells. Thus, they may have translational potential as chemopreventive or therapeutic agents for colorectal carcinoma.

Immunotherapeutic Challenges for Pediatric Cancers.

Solid tumors contain a mixture of malignant cells and non-malignant infiltrating cells that often create a chronic inflammatory and immunosuppressive microenvironment that restricts immunotherapeutic approaches. Although childhood and adult cancers share some similarities related to microenvironmental changes, pediatric cancers are unique, and adult cancer practices may not be wholly applicable to our pediatric patients. This review highlights the differences in tumorigenesis, viral infection, and immunologic response between children and adults that need to be considered when trying to apply experiences from experimental therapies in adult cancer patients to pediatric cancers.

Signal transducer and activator of transcription 3 activation is associated with bladder cancer cell growth and survival.

BACKGROUND: Constitutive activation of signal transducer and activator of transcription 3 (Stat3) signaling pathway plays an important role in several human cancers. Activation of Stat3 is dependent on the phosphorylation at the tyrosine residue 705 by upstream kinases and subsequent nuclear translocation after dimerization. It remains unclear whether oncogenic Stat3 signaling pathway is involved in the oncogenesis of bladder cancer. RESULTS: We found that elevated Stat3 phosphorylation in 19 of 100 (19%) bladder cancer tissues as well as bladder cancer cell lines, WH, UMUC-3 and 253J. To explore whether Stat3 activation is associated with cell growth and survival of bladder cancer, we targeted the Stat3 signaling pathway in bladder cancer cells using an adenovirus-mediated dominant-negative Stat3 (Y705F) and a small molecule compound, STA-21. Both prohibited cell growth and induction of apoptosis in these bladder cancer cell lines but not in normal bladder smooth muscle cell (BdSMC). The survival inhibition might be mediated through apoptotic caspase 3, 8 and 9 pathways. Moreover, down-regulation of anti-apoptotic genes (Bcl-2, Bcl-xL and survivin) and a cell cycle regulating gene (cyclin D1) was associated with the cell growth inhibition and apoptosis. CONCLUSION: These results indicated that activation of Stat3 is crucial for bladder cancer cell growth and survival. Therefore, interference of Stat3 signaling pathway emerges as a potential therapeutic approach for bladder cancer.

STAT3 can be activated through paracrine signaling in breast epithelial cells.

BACKGROUND: Many cancers, including breast cancer, have been identified with increased levels of phosphorylated or the active form of Signal Transducers and Activators of Transcription 3 (STAT3) protein. However, whether the tumor microenvironment plays a role in this activation is still poorly understood. METHODS: Conditioned media, which contains soluble factors from MDA-MB-231 and MDA-MB-468 breast cancer cells and breast cancer associated fibroblasts, was added to MCF-10A breast epithelial and MDA-MB-453 breast cancer cells. The stimulation of phosphorylated STAT3 (p-STAT3) levels by conditioned media was assayed by Western blot in the presence or absence of neutralized IL-6 antibody, or a JAK/STAT3 inhibitor, JSI-124. The stimulation of cell proliferation in MCF-10A cells by conditioned media in the presence or absence of JSI-124 was subjected to MTT analysis. IL-6, IL-10, and VEGF levels were determined by ELISA analysis. RESULTS: Our results demonstrated that conditioned media from cell lines with constitutively active STAT3 are sufficient to induce p-STAT3 levels in various recipients that do not possess elevated p-STAT3 levels. This signaling occurs through the JAK/STAT3 pathway, leading to STAT3 phosphorylation as early as 30 minutes and is persistent for at least 24 hours. ELISA analysis confirmed a correlation between elevated levels of IL-6 production and p-STAT3. Neutralization of the IL-6 ligand or gp130 was sufficient to block increased levels of p-STAT3 (Y705) in treated cells. Furthermore, soluble factors within the MDA-MB-231 conditioned media were also sufficient to stimulate an increase in IL-6 production from MCF-10A cells. CONCLUSION: These results demonstrate STAT3 phosphorylation in breast epithelial cells can be stimulated by paracrine signaling through soluble factors from both breast cancer cells and breast cancer associated fibroblasts with elevated STAT3 phosphorylation. The induction of STAT3 phosphorylation is through the IL-6/JAK pathway and appears to be associated with cell proliferation. Understanding how IL-6 and other soluble factors may lead to STAT3 activation via the tumor microenvironment will provide new therapeutic regimens for breast carcinomas and other cancers with elevated p-STAT3 levels.

Oncolytic virus and PD-1/PD-L1 blockade combination therapy.

Oncolytic viruses are lytic for many types of cancers but are attenuated or replication-defective in normal tissues. Aside from tumor lysis, oncolytic viruses can induce host immune responses against cancer cells and may thus be viewed as a form of immunotherapy. Although recent successes with checkpoint inhibitors have shown that enhancing antitumor immunity can be effective, the dynamic nature of the immunosuppressive tumor microenvironment presents significant hurdles to the broader application of these therapies. Targeting one immune-suppressive pathway may not be sufficient to eliminate tumors. Here we focus on the development of the combination of oncolytic virotherapy with checkpoint inhibitors designed to target the programmed cell death protein 1 and programmed cell death ligand 1 signaling axis. We also discuss future directions for the clinical application of this novel combination therapy.

High Mobility Group Box 1 Influences HSV1716 Spread and Acts as an Adjuvant to Chemotherapy.

High Mobility Group Box 1 (HMGB1) is a multifunctional protein that plays various roles in the processes of inflammation, cancer, and other diseases. Many reports document abundant HMGB1 release following infection with oncolytic viruses (OVs). Further, other groups including previous reports from our laboratory highlight the synergistic effects of OVs with chemotherapy drugs. Here, we show that virus-free supernatants have varying cytotoxic potential, and HMGB1 is actively secreted by two established fibroblast cell lines (NIH 3T3 and 3T6-Swiss albino) following HSV1716 infection in vitro. Further, pharmacologic inhibition or genetic knock-down of HMGB1 reveals a role for HMGB1 in viral restriction, the ability to modulate bystander cell proliferation, and drug sensitivity in 3T6 cells. These data further support the multifactorial role of HMGB1, and suggest it could be a target for modulating the efficacy of oncolytic virus therapies alone or in combination with other frontline cancer treatments.

Myelolytic Treatments Enhance Oncolytic Herpes Virotherapy in Models of Ewing Sarcoma by Modulating the Immune Microenvironment.

Ewing sarcoma is a highly aggressive cancer that promotes the infiltration and activation of pro-tumor M2-like macrophages. Oncolytic virotherapy that selectively infects and destroys cancer cells is a promising option for treating Ewing sarcoma. The effect of tumor macrophages on oncolytic virus therapy, however, is variable among solid tumors and is unknown in Ewing sarcoma. We tested the effects of macrophage reduction using liposomal clodronate (Clodrosome) and trabectedin on the antitumor efficacy of intratumoral oncolytic herpes simplex virus, rRp450, in two Ewing sarcoma xenograft models. Both agents enhanced antitumor efficacy without increasing virus replication. The most profound effects were in A673 with only a transient effect on response rates in TC71. Interestingly, A673 was more dependent than TC71 on macrophages for its tumorigenesis. We found Clodrosome and virus together induced expression of antitumorigenic genes and reduced expression of protumorigenic genes in both the tumor-associated macrophages and the overall tumor stroma. Trabectedin reduced intratumoral natural killer (NK) cells, myeloid-derived suppressor cells, and M2-like macrophages, and prevented their increase following virotherapy. Our data suggest that a combination of trabectedin and oncolytic herpes virotherapy warrants testing in the clinical setting.

TGF-ß Inhibition Improves Oncolytic Herpes Viroimmunotherapy in Murine Models of Rhabdomyosarcoma.

Oncolytic viruses are an emerging class of cancer therapeutics that couple cytotoxicity with the induction of an anti-tumor immune response. Host-virus interactions are complex and modulated by a tumor microenvironment whose immunosuppressive activities can limit the effectiveness of cancer immunotherapies. In an effort to improve this aspect of oncolytic virotherapy, we combined the oncolytic herpes virus HSV1716 with the transforming growth factor beta receptor 1 (TGF-ßR1) inhibitor A8301 to treat syngeneic models of murine rhabdomyosarcoma. Mice that received HSV1716 or A8301 alone showed little to no benefit in efficacy and survival over controls. Conversely, mice given combination therapy exhibited tumor stabilization throughout the treatment regimen, which was reflected in significantly prolonged survival times including some complete responses. In vitro cell viability and virus replication assays showed that the rhabdomyosarcoma cell lines were generally insensitive to HSV1716 and A8301. Likewise, in vivo virus replication assays showed that HSV1716 titers moderately decreased in the presence of A8301. The enhanced efficacy instead appears to be dependent on the generation of an improved anti-tumor T cell response as determined by its loss in athymic nude mice and following in vivo depletion of either CD4+ or CD8+ cells. These data suggest TGF-ß inhibition can augment the immunotherapeutic efficacy of oncolytic herpes virotherapy.

Cooperation of Oncolytic Herpes Virotherapy and PD-1 Blockade in Murine Rhabdomyosarcoma Models.

Oncolytic virotherapy is an effective immunotherapeutic approach for cancer treatment via a multistep process including direct tumor cell lysis, induction of cytotoxic or apoptosis-sensitizing cytokines and promotion of antitumor T cell responses. Solid tumors limit the effectiveness of immunotherapeutics in diverse ways such as secretion of immunosuppressive cytokines and expression of immune inhibitory ligands to inhibit antitumor T cell function. Blocking programmed cell death protein (PD)-1 signaling, which mediates T cell suppression via engagement of its inhibitory ligands, PD-L1 or PD-L2, is of particular interest due to recent successes in many types of cancer. In syngeneic murine rhabdomyosarcoma models, we found that M3-9-M (MHC I high) but not 76-9 (MHC I low) tumors respond to oncolytic herpes simplex virus-1 (oHSV-1) and PD-1 blockade combination therapy. In addition, the therapeutic outcomes in M3-9-M tumor models correlated with the increased incidence of CD4+ and CD8+ T cells but not with the CD4+CD25+Foxp3+ regulatory T cell populations in the tumor. Overall, our data suggest the combination of PD-1 blockade and oHSV-1 may be an effective treatment strategy for childhood soft tissue sarcoma.

Oncolytic Herpes Virus rRp450 Shows Efficacy in Orthotopic Xenograft Group 3/4 Medulloblastomas and Atypical Teratoid/Rhabdoid Tumors.

Pediatric brain tumors including medulloblastoma and atypical teratoid/rhabdoid tumor are associated with significant mortality and treatment-associated morbidity. While medulloblastoma tumors within molecular subgroups 3 and 4 have a propensity to metastasize, atypical teratoid/rhabdoid tumors frequently afflict a very young patient population. Adjuvant treatment options for children suffering with these tumors are not only sub-optimal but also associated with many neurocognitive obstacles. A potentially novel treatment approach is oncolytic virotherapy, a developing therapeutic platform currently in early-phase clinical trials for pediatric brain tumors and recently US Food and Drug Administration (FDA)-approved to treat melanoma in adults. We evaluated the therapeutic potential of the clinically available oncolytic herpes simplex vector rRp450 in cell lines derived from medulloblastoma and atypical teratoid/rhabdoid tumor. Cells of both tumor types were supportive of virus replication and virus-mediated cytotoxicity. Orthotopic xenograft models of medulloblastoma and atypical teratoid/rhabdoid tumors displayed significantly prolonged survival following a single, stereotactic intratumoral injection of rRp450. Furthermore, addition of the chemotherapeutic prodrug cyclophosphamide (CPA) enhanced rRp450's in vivo efficacy. In conclusion, oncolytic herpes viruses with the ability to bioactivate the prodrug CPA within the tumor microenvironment warrant further investigation as a potential therapy for pediatric brain tumors.

Aurora A kinase inhibition enhances oncolytic herpes virotherapy through cytotoxic synergy and innate cellular immune modulation.

Malignant peripheral nerve sheath tumor (MPNST) and neuroblastoma models respond to the investigational small molecule Aurora A kinase inhibitor, alisertib. We previously reported that MPNST and neuroblastomas are also susceptible to oncolytic herpes virus (oHSV) therapy. Herein, we show that combination of alisertib and HSV1716, a virus derived from HSV-1 and attenuated by deletion of RL1, exhibits significantly increased antitumor efficacy compared to either monotherapy. Alisertib and HSV1716 reduced tumor growth and increased survival in two xenograft models of MPNST and neuroblastoma. We found the enhanced antitumor effect was due to multiple mechanisms that likely each contribute to the combination effect. First, oncolytic herpes virus increased the sensitivity of uninfected cells to alisertib cytotoxicity, a process we term virus-induced therapeutic adjuvant (VITA). Second, alisertib increased peak virus production and slowed virus clearance from tumors, both likely a consequence of it preventing virus-mediated increase of intratumoral NK cells. We also found that alisertib inhibited virus-induced accumulation of intratumoral myeloid derived suppressor cells, which normally are protumorigenic. Our data suggest that clinical trials of the combination of oHSV and alisertib are warranted in patients with neuroblastoma or MPNST.