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Background Oncolytic viral efficacy may be limited by the penetration of the virus into tumors. This may be enhanced by intraoperative application of virus immediately after surgical resection. Methods Oncolytic vaccinia virus GLV-1h68 was delivered in silk-elastin-like protein polymer (SELP) in vitro and in vivo in anaplastic thyroid carcinoma cell line 8505c in nude mice. Results GLV-1h68 in SELP infected and lysed anaplastic thyroid cancer cells in vitro equally as effectively as in phosphate-buffered saline (PBS), and at 1 week retains a thousand fold greater infectious plaque-forming units. In surgical resection models of residual tumor, GLV-1h68 in SELP improves tumor control and shows increased viral β-galactosidase expression as compared to PBS. Conclusion The use of SELP matrix for intraoperative oncolytic viral delivery protects infectious viral particles from degradation, facilitates sustained viral delivery and transgene expression, and improves tumor control. Such optimization of methods of oncolytic viral delivery may enhance therapeutic outcomes. © 2014 Wiley Periodicals, Inc. Head Neck, 2015

Salivary gland carcinomas are relatively rare malignant tumors, accounting for less than 5% of all cancers of the head and neck.1 They encompass a wide spectrum of histologic abnormalities with varied biologic behavior. Initial therapy of localized disease consists of complete surgical excision. The risk of recurrence and metastasis is significantly higher in patients with locally advanced salivary carcinomas. Individuals with high-grade salivary carcinomas have a 5-year survival of roughly 40%; those with low-and intermediate-grade tumors have a 5-year survival of 85% to 90%.2,3 For these patients, complete surgical resection followed by postoperative radiation therapy is recommended.4 Unfortunately, patients with recurrent and/or unresectable disease have few effective treatment options. Clinical trials exploring the role of chemotherapy in the management of salivary gland carcinomas failed to show survival benefit.1 Chemotherapy, therefore, is generally reserved as a palliative measure for patients with symptoms and/or rapid disease progression. Clearly, novel therapies are needed to improve outcomes for patients with salivary carcinomas. Oncolytic viruses have emerged as versatile therapeutic agents that can selectively infect, replicate within, and ultimately lyse a host cancer cell. A variety of viruses have been reported5–7 to possess on-colytic antitumoral activity, including herpes simplex type 1, adenovirus, reovirus, vesicular stomatitis virus, measles virus, poliovirus, West Nile virus, and New-castle disease virus. Vaccinia virus, which is a double-stranded DNA member of the genus Orthopoxvirus, has many unique characteristics that make it an excellent tool in cancer treatment. In addition to its natural tropism for tumor tissues and its abilities for efficient entry, replication, and lysis, vaccinia virus has a remarkable safety record in its historical widespread human application as a vaccine for smallpox. The vaccinia replication cycle occurs exclusively in the cytoplasm, which eliminates the possibility of chromosomal integration.7 Furthermore, there are many Food and Drug Administration–approved antiviral agents available to limit viral spread and control the unlikely possibility of viral toxicity.8 Consequently, several recombinant vaccinia viruses have been constructed and studied in preclinical and clinical phase 1 settings.8,9 Recombinant, replication-competent vaccinia virus (GLV-1h68) has been reported10–16 as effective in breast cancer, thyroid cancer, mesothelioma, pancreatic cancer, prostate cancer, human hepatocellular carcinoma, and head and neck cancer models. The aim of this study was to assess the usefulness of GLV-1h68 as a therapeutic agent against salivary gland carcinoma in vitro and in vivo. Murine flank and orthotopic parotid tumor models were used.

Background Approximately one fourth of bone and soft-tissue sarcomas recur after prior treatment. GLV-1h68 is a recombinant, replication-competent vaccinia virus that has been shown to have oncolytic effects against many human cancer types. We sought to determine whether GLV-1h68 could selectively target and lyse a panel of human bone and soft-tissue sarcoma cell lines in vitro and in vivo . Methods GLV-1h68 was tested in a panel of four cell lines including: fibrosarcoma HT-1080, osteosarcoma U-2OS, fibrohistiocytoma M-805, and rhabdomyosarcoma HTB-82. Gene expression, infectivity, viral proliferation, and cytotoxicity were characterized in vitro . HT-1080 xenograft flank tumors grown in vivo were injected intratumorally with a single dose of GLV-1h68. Results All four cell lines supported robust viral transgene expression in vitro . At a multiplicity of infection (MOI) of five, GLV-1h68 was cytotoxic to three cell lines, resulting in >80% cytotoxicity over 7 d. In vivo , a single injection of GLV-1h68 into HT-1080 xenografts exhibited localized intratumoral luciferase activity peaking at d 2–4, with gradual resolution over 8 d and no evidence of spread to normal tissues. Treated animals exhibited near-complete tumor regression over a 28-d period without observed toxicity. Conclusion GLV-1h68 has potent direct oncolytic effects against human sarcoma in vitro and in vivo . Recombinant vaccinia oncolytic virotherapy could provide a new platform for the treatment of patients with bone and soft tissue sarcomas. Future clinical trials investigating oncolytic vaccinia as a therapy for sarcomas are warranted.

Replication-competent oncolytic viruses have shown great promise as a potential cancer treatment. This study aimed to determine whether a novel vaccinia virus, GLV-1h151, with genetic modifications enhancing cancer specificity and enabling virus detection, is effective against a range of human cancers and is safe when administered in preclinical models. GLV-1h151 was modified with deletion of thymidine kinase enhancing specificity and insertion of the green fluorescent protein (GFP) gene. The virus was tested in several human cancer cell lines for cytotoxicity including breast, lung, pancreatic, and colorectal. Virus replication was assessed via visualization of GFP expression and bioluminescence, and viral plaque assays. Finally, GLV-1h151 was administered systemically or intratumorally in mice with pancreatic cancer xenografts (PANC-1) to assess virus biodistribution, toxicity, and effect on tumor growth. GLV-1h151 effectively infected, replicated in, and killed several cancer cell types. Detection and visualization of virus replication was successful via fluorescence imaging of GFP expression, which was dose dependent. When administered intravenously or intratumorally in vivo, GLV-1h151 regressed tumor growth (P

Background. Newcastle disease virus (NDV) is a paramyxovirus that is pathogenic in birds but causes only mild flu- like symptoms in human beings. NDV(F3aa)-GFP is a genetically modified, fusogenic NDV. We assessed the utility of NDV(F3aa)- GFP in treating head and neck squamous cell carcinoma. Methods and Results. At a multiplicity of infection (MOI) of 1, NDV(F3aa)-GFP infection of 3 cell lines supported strong GFP expression by 36 hours. Four cell lines were highly sensitivite to vi- ral cytotoxicity, with >75% of cells lysed by day 6 at MOI 0.1, and 2 other cell lines were partially susceptible. Murine SCC25 flank tumors exhibited robust GFP expression after a single intratumoral viral injection and showed near-complete tumor regression over 34 days. There were no adverse effects attributable to therapy. Conclusions. We demonstrate that a fusogenic NDV exerts potent oncolytic effects against human head and neck cancer and support its continued investigation for clinical application. V C 2010 Wiley Periodicals, Inc. Head Neck 33: 1394-1399

Background Sinonasal squamous cell carcinoma (SSCC) and nasal inverted papilloma (NIP) represent the predominant type of malignant and benign tumors in sinonasal tract, respectively. CD4+CD25+Foxp3+ natural regulatory T (Treg) cells might play critical role(s) in the suppression of anti-tumor immune response and thus shed light on tumor progression from benign to malignant. Objective This study aimed to evaluate the frequency and suppressive capacity of Treg cells in SSCC compared to NIP and further to explore the underlying mechanisms. Patients and Methods Frequencies of Treg, Th1 and Th2 cells were evaluated by flow cytometry in tissue homogenate and peripheral blood from 31 SSCC patients, 32 NIP patients and 35 normal controls. Treg cells were tested for regulatory function by co-culture with effector T cells. CCR4 and its ligands, CCL22 and CCL17, were analyzed by flow cytometry and Luminex, respectively. The chemoattractant properties of CCR4/CCL22 and CCR4/CCL17 for Treg cells were assessed using the Boyden chamber technique, to elucidate the potential mechanisms of Treg recruitment in tumor microenvironment. Treg cells induction via TGF-β was assessed with transwells after local CD4+Foxp3+ T cells were assessed by immunohistochemistry and TGF-β concentration was measured by Luminex. Results Tumor-infiltrating Treg cells increased significantly from normal to NIP to SSCC (P ≤ 0.001 for normal vs. NIP and P = 0.004 for NIP vs. SSCC). Significantly elevated frequency and enhanced suppression capacity of circulating Treg cells in SSCC were detected compared to NIP and healthy controls, concomitant with Th1 decrease and Th2 increase. Apparently increased CCL22 attracted CCR4-expressing Treg cells to tumor microenvironment in SSCC, compared to NIP. SSCC produced significantly more TGF-β than NIP and thus possessed greater potential for Treg cell induction. Conclusion Frequency and suppressive capacity of Treg cells enhanced with progression of malignancy from NIP to SSCC. Circulating Treg cells were recruited to tumor tissue via CCR4/CCL22 signalling, whereas tumor-synthesised TGF-β contributed to induction of peripheral Treg cells.