Systemic oncolytic herpes virus therapy of poorly immunogenic prostate cancer metastatic to lung.

PURPOSE: Our goal was to evaluate whether systemic administration of NV1042, an interleukin-12 (IL-12)-expressing oncolytic herpes simplex virus, and its noncytokine parental vector NV1023 are effective against preexisting metastatic prostate cancer in an immunocompetent mice model. EXPERIMENTAL DESIGN: Metastatic TRAMP-C2 lung tumors established in C57Bl/6 or nude mice were treated on day 21 with four i.v. administrations of NV1042 or NV1023 and sacrificed on day 42 to assess virus efficacy and the potential mechanism of efficacy. RESULTS: NV1042 or NV1023 treatment was similarly effective in eliminating extrapleural and hemorrhagic tumors present in mock-treated mice. However, NV1042 was further effective compared with NV1023 in controlling the growth of lung tumors (as determined by mean surface tumor nodule number, lung weights, and surface tumor burden) and in extending survival. NV1042-treated mice exhibited a transient increase of serum IL-12 1 day posttreatment, whereas IL-12 levels in tumor bearing lungs persisted a further 2 days at least. Only splenocytes from NV1042-treated mice secreted IFN-gamma in response to TRAMP-C2 stimulation and displayed natural killer activity. The IL-12-mediated enhancement observed with NV1042 in the syngeneic model was abrogated in athymic mice treated in a similar manner, thus indicating a role for T cells in the augmented efficacy of NV1042 virus. CONCLUSIONS: Systemic administration of the IL-12-expressing NV1042 virus is more effective than its noncytokine parent, NV1023, against preestablished metastatic lung tumors. Given the clinical safety profile of NV1020, the parental vector of NV1023, and NV1042's enhanced efficacy and ability to activate the host immune system, NV1042 merits clinical consideration for treating metastatic prostate cancers.     

Systemic therapy of spontaneous prostate cancer in transgenic mice with oncolytic herpes simplex viruses

Oncolytic viruses are an innovative therapeutic strategy for cancer, wherein viral replication and cytotoxicity are selective for tumor cells. Here we show the efficacy of systemically administered oncolytic viruses for the treatment of spontaneously arising tumors, specifically the use of oncolytic herpes simplex viruses (HSV) administered i.v. to treat spontaneously developing primary and metastatic prostate cancer in the transgenic TRAMP mouse, which recapitulates human prostate cancer progression. Four administrations of systemically delivered NV1023 virus, an HSV-1/HSV-2 oncolytic recombinant, to TRAMP mice at 12 or 18 weeks of age (presence of prostate adenocarcinoma or metastatic disease, respectively) inhibited primary tumor growth and metastases to lymph nodes. Expression of interleukin 12 (IL-12) from NV1042 virus, a derivative of NV1023, was additionally effective, significantly reducing the frequency of development of prostate cancer and lung metastases, even when the mice were treated after the onset of metastasis at 18 weeks of age. NV1042-infected cells, as detected by 5-bromo-4-chloro-3-indolyl-beta-d-galactopyranoside staining for Lac Z expressed by the virus, were present in prostate tumors 1 week after the final virus injection and viral DNA was detected at 2 weeks after final virus injection by real-time PCR in primary and metastatic tumors but not in liver or blood. No toxicity was observed in any of the treated mice. The efficacy of the IL-12-expressing NV1042 virus in this aggressive prostate cancer model using a clinically relevant treatment paradigm merits its consideration for clinical studies.     

Angiogenesis inhibition by an oncolytic herpes virus expressing interleukin 12.

PURPOSE: Oncolytic herpes simplex viruses (HSVs) may have significant antitumor effects resulting from the direct lysis of cancer cells. HSVs may also be used to express inserted transgenes to exploit additional therapeutic strategies. The ability of an interleukin (IL)-12-expressing HSV to treat squamous cell carcinoma (SCC) by inhibition of tumor angiogenesis is investigated in this study. EXPERIMENTAL DESIGN: A replication-competent, attenuated, oncolytic HSV carrying the murine IL-12 gene (NV1042), its non-cytokine-carrying analog (NV1023), or saline was used to treat established murine SCC flank tumors by intratumoral injection. The expression of secondary antiangiogenic mediators was measured. Angiogenesis inhibition was assessed by in vivo Matrigel plug assays, flank tumor subdermal vascularity, and in vitro endothelial cell tubule formation assay. RESULTS: Intratumoral injections of NV1042 (2 x 10(7) plaque-forming units) into murine SCC VII flank tumors resulted in smaller tumor volumes as compared with NV1023 or saline. IL-12 and IFN-gamma expression in tumors was 440 and 2.2 pg/mg, respectively, at 24 h after NV1042 injection, but both IL-12 and IFN-gamma were undetectable (<0.2 pg/mg) after NV1023 or saline injections. Expression of two antiangiogenesis mediators, monokine induced by IFN-gamma and IFN-inducible protein 10, was elevated after NV1042 treatment. Matrigel plug assays of NV1042-transfected SCC VII tumor cells demonstrated significantly decreased hemoglobin content and microvessel density as compared with NV1023 and PBS. Excised murine flank tumors treated with NV1042 had decreased subdermal vascularity as compared with NV1023 and PBS. Both splenocytes and IL-12 expression by NV1042 were required for in vitro inhibition of endothelial tubule formation. CONCLUSIONS: IL-12 expression by an oncolytic herpes virus enhances therapy of SCC through antiangiogenic mechanisms. Strategies combining HSV oncolysis with angiogenesis inhibition merit further investigation for potential clinical application.     

Neoadjuvant treatment of hepatic malignancy: an oncolytic herpes simplex virus expressing IL-12 effectively treats the parent tumor and protects against recurrence-after resection

The objective of the study was to evaluate the utility of NV1042, a replication competent, oncolytic herpes simplex virus (HSV) containing the interleukin-12 (IL-12) gene, as primary treatment for hepatic tumors and to further assess its ability to reduce tumor recurrence following resection. Resection is the most effective therapy for hepatic malignancies, but is not possible in the majority of the patients. Furthermore, recurrence is common after resection, most often in the remnant liver and likely because of microscopic residual disease in the setting of postoperative host cellular immune dysfunction. We hypothesize that, unlike other gene transfer approaches, direct injection of liver tumors with replication competent, oncolytic HSV expressing IL-12 will not only provide effective control of the parent tumor, but will also elicit an immune response directed at residual tumor cells, thus decreasing the risk of cancer recurrence after resection. Solitary Morris hepatomas, established in Buffalo rat livers, were injected directly with 10(7) particles of NV1042, NV1023, an oncolytic HSV identical to NV1042 but without the IL-12 gene, or with saline. Following tumor injection, the parent tumors were resected and measured and the animals were challenged with an intraportal injection of 10(5) tumor cells, recreating the clinical scenario of residual microscopic cancer. In vitro cytotoxicity against Morris hepatoma cells was similar for both viruses at a multiplicity of infection of 1 (MOI, ratio of viral particles to target cells), with >90% tumor cell kill by day 6. NV1042 induced high-level expression of IL-12 in vitro, peaking after 4 days in culture. Furthermore, a single intratumoral injection of NV1042, but not NV1023, induced marked IL-12 and interferon-gamma (IFN-gamma) expression. Both viruses induced a significant local immune response as evidenced by an increase in the number of intratumoral CD4(+) and CD8(+) lymphocytes, although the peak of CD8(+) infiltration was later with NV1042 compared with NV1023. NV1042 and NV1023 reduced parent tumor volume by 74% (P<.003) and 52% (P<.03), respectively, compared to control animals. Treatment of established tumors with NV1042, but not with NV1023, significantly reduced the number of hepatic tumors after resection of the parent tumor and rechallenge (16.8+/-11 (median=4) vs. 65.9+/-15 (median=66) in control animals, P<.025). In conclusion, oncolytic HSV therapy combined with local immune stimulation with IL-12 offers effective control of parent hepatic tumors and also protects against microscopic residual disease after resection. The ease of use of this combined modality approach, which appears to be superior to either approach alone, suggests that it may have clinical relevance, both as primary treatment for patients with unresectable tumors and also as a neoadjuvant strategy for reducing recurrence after resection.     

Cytokine-secreting herpes viral mutants effectively treat tumor in a murine metastatic colorectal liver model by oncolytic and T-cell-dependent mechanisms

In this model of hepatic micrometastases, the antitumor efficacy and role of the T-cell and natural killer (NK) cell populations were studied for oncolytic herpes simplex virus type-1 (HSV-1) viral mutants containing the granulocyte-monocyte colony stimulating factor (GM-CSF (NV1034)) or interluken-12 (IL-12 (NV1042)) cytokine genes. These were compared to saline and control virus (NV1023) in vitro and in vivo. HSV-1 mutants were assessed for cytotoxicity, replication and cytokine expression in CT-26 cells. A syngeneic micrometastatic liver model was then established in naive and immune cell-depleted animals to assess the antitumor efficacy of these viruses. In vitro cytotoxicity and viral replication were similar for each virus, resulting in greater than 80 and 98% cytotoxicity at multiplicity of infection of 1 and 10, respectively. Peak viral titers were 25- to 50-fold higher than initial titer and were not significantly different between viruses. In vivo, all three viruses reduced metastases relative to control, but cytokine-secreting viruses did so with greater efficacy compared to NV1023. This effect was abrogated by T-cell depletion, but not NK-cell depletion. Single-agent therapy with oncolytic viral agents containing GM-CSF or IL-12 is effective in a murine model of liver metastases and likely involves direct viral oncolysis and actions of specific immune effector cells.     

Oncolytic herpes simplex virus vector therapy of breast cancer in C3(1)/SV40 T-antigen transgenic mice

Oncolytic herpes simplex virus vectors are a promising strategy for cancer therapy, as direct cytotoxic agents, inducers of antitumor immune responses, and as expressers of anticancer genes. Progress is dependent upon representative preclinical models to evaluate therapy. In this study, two families of oncolytic herpes simplex virus vectors (G207 and NV1020 series) that have been in clinical trials were examined for the treatment of breast cancer, using the C3(1)/T-Ag transgenic mouse model. Female mice spontaneously develop mammary carcinomas, and the C3(1)/T-Ag-derived tumor cell line M6c forms implantable tumors. Both in vitro and in vivo, G47Delta, derived from G207 by deletion of ICP47 and the US11 promoter, was more efficacious than G207. Whereas NV1023, derived from NV1020 by deletion of ICP47 and insertion of LacZ, was as cytotoxic to M6c cells in vitro as G47Delta, it did not inhibit the growth of s.c. M6c tumors but did extend the survival of intracerebral tumor bearing mice. In contrast, NV1042, NV1023 expressing interleukin 12, inhibited s.c. M6c tumor growth to a similar extent as G47Delta, but was less effective than NV1023 in intracerebral tumors. In the spontaneously arising mammary tumor model, when only the first arising tumor per mouse was treated, G47Delta inhibited the growth of a subset of tumors, and when all tumors were treated, G47Delta significantly delayed tumor progression. When the first mammary tumor was treated and the remaining mammary glands removed, NV1042 was more efficacious than G47Delta at inhibiting the growth and progression of injected tumors.     

Sensitivity of squamous cell carcinoma lymph node metastases to herpes oncolytic therapy.

PURPOSE: Cancer metastases may have phenotypic and genetic differences from their primary cancers of origin. Engineered, replication-competent, attenuated viruses based on herpes simplex virus-1 (HSV-1) have shown potent oncolytic effects in treating primary tumors in animal tumor models, but their efficacy in treating lymph node metastases is poorly understood. We compared the efficacy of an attenuated oncolytic HSV-1 (NV1023) in treating a series of murine squamous carcinoma cell lines derived from serial implantation and harvest from metastatic lymph nodes. EXPERIMENTAL DESIGN and RESULTS: The auricles of C3H/HeJ mice were implanted with SCCVII. Cervical nodal metastases were isolated, expanded in vitro, and reimplanted into new mice. A series of cell lines (LN1-LN7) were generated through seven serial passages. Cells from higher LN passages showed consistent trends toward increased migratory and invasive ability, increased cell surface nectin-1 (an HSV-1 receptor) expression, and increased glycoprotein D binding. Exposure to NV1023 showed increased viral entry, replication, and cytotoxicity with higher LN passages. Intratumoral injection of NV1023 in a murine flank tumor model caused significantly greater tumor regression and increased viral infection of LN7 compared with SCCVII. CONCLUSIONS: These results show that lymph node metastases may undergo selection for characteristics, including increased nectin-1 expression, that make them more sensitive targets for herpes oncolytic therapy. These findings support the clinical application of these agents for the treatment of lymph node metastases.     

Calcium depletion enhances nectin-1 expression and herpes oncolytic therapy of squamous cell carcinoma

Attenuated, replication-competent, oncolytic herpes simplex virus type 1 (HSV-1) are effective at infecting and lysing many human malignancies in preclinical studies. Nectin-1 is a cell-surface receptor for HSV-1 envelope glycoprotein D (gD) that also forms a component of intercellular adherens junctions (AJs). We sought to determine if the disruption of AJs in squamous cell carcinoma (SCC) through calcium depletion could be utilized to increase nectin-1 exposure and enhance HSV therapy. NV1023 is a single copy gamma(1)34.5-deleted, lacZ-expressing, oncolytic HSV-1. Calcium depletion caused cell separation and increased nectin-1 expression for three SCC cell lines growing at confluence. NV1023 viral entry, soluble gD protein binding and NV1023 cytotoxicity were all significantly enhanced for these cell lines at low calcium conditions. The increase in NV1023 entry at low calcium conditions was abrogated by nectin-1 antibody blockade. Murine SCC flank tumors treated with ethylenediaminetetraacetic acid (EDTA) showed increased nectin-1 expression and increased susceptibility to NV1023 infection. Combined NV1023 and EDTA intratumoral injections demonstrated significantly enhanced tumor regression as compared to NV1023 alone. These findings establish, as proof-of-principle, that herpes viral receptor expression may be modulated on cancer cells to enhance oncolytic therapy. This strategy might have future application toward improving therapy with a variety of herpes vectors.     

Triple combination of oncolytic herpes simplex virus-1 vectors armed with interleukin-12, interleukin-18, or soluble B7-1 results in enhanced antitumor efficacy.

Conditionally replicating herpes simplex virus-1 (HSV-1) vectors are promising therapeutic agents for cancer. Insertion of therapeutic transgenes into the viral genome should confer desired anticancer functions in addition to oncolytic activities. Herein, using bacterial artificial chromosome and two recombinase-mediated recombinations, we simultaneously created four "armed" oncolytic HSV-1, designated vHsv-B7.1-Ig, vHsv-interleukin (IL)-12, vHsv-IL-18, and vHsv-null, which express murine soluble B7.1 (B7.1-Ig), murine IL-12, murine IL-18, and no transgene, respectively. These vHsv vectors possess deletions in the gamma34.5 genes and contain the green fluorescent protein gene as a histochemical marker and the immunostimulatory transgene inserted in the deleted ICP6 locus. The vHsv showed similar replicative capabilities in vitro. The in vivo efficacy was tested in A/J mice harboring s.c. tumors of syngeneic and poorly immunogenic Neuro2a neuroblastoma. The triple combination of vHsv-B7.1-Ig, vHsv-IL-12, and vHsv-IL-18 exhibited the highest efficacy among all single vHsv or combinations of two viruses. Combining 1 x 10(5) plaque-forming units each of the three armed viruses showed stronger antitumor activities than any single armed virus at 3 x 10(5) plaque-forming units in inoculated tumors as well as in noninoculated remote tumors. Studies using athymic mice indicated that this enhancement of antitumor efficacy was likely mediated by T-cell immune responses. The combined use of multiple oncolytic HSV-1 armed with different immunostimulatory genes may be a useful strategy for cancer therapy.     

Nerve-sparing therapy with oncolytic herpes virus for cancers with neural invasion.

PURPOSE: The invasion of cancer cells along nerves is an ominous pathologic finding associated with poor outcomes for a variety of tumors, including pancreatic and head and neck carcinomas. Peripheral nerves may serve as a conduit for these cancers to track into the central nervous system. Cancer progression within nerves and surgical resection of infiltrated nerves result in a permanent loss of neural function, potentially causing cosmetic and functional morbidity. Herpes simplex viruses (HSV) have utility for gene transfer into nerves and as oncolytic agents. We studied the use of an attenuated HSV, NV1023, as treatment for cancers with neural invasion. EXPERIMENTAL DESIGN AND RESULTS: NV1023 injection into the sciatic nerves of nude mice had no toxic effect on nerve function, whereas similar doses of wild-type HSV-1 (F' strain) caused complete nerve paralysis within 4 days and 100% mortality at day 6. NV1023 showed effective cytotoxicity in vitro on three neurotrophic human carcinoma cell lines, including pancreatic (MiaPaCa2), squamous cell (QLL2), and adenoid cystic (ACC3) carcinomas. A model of neural invasion was established by implanting human carcinoma cells in the sciatic nerves of nude mice. All control group mice developed left hind limb paralysis 5 to 7 weeks after tumor injection, whereas animals treated with NV1023 maintained intact nerve function and showed significant tumor regression (P < 0.0001). CONCLUSIONS: These results show that NV1023 oncolytic therapy may effectively treat cancers with neural invasion and preserve neural function. These findings hold significant clinical implications for patients with cancer neural invasion.     

Enhanced nectin-1 expression and herpes oncolytic sensitivity in highly migratory and invasive carcinoma.

PURPOSE: Although a variety of malignant tumors are susceptible to therapy with oncolytic herpes simplex viruses, the determinants of tumor sensitivity to these viruses are poorly understood. Nectin-1 is a cell surface adhesion molecule that is a component of intercellular adherens junctions and also functions as a herpes viral receptor. Because highly invasive cells may have decreased intercellular adhesion, we sought to determine if such cells might also have altered availability of cell surface nectin-1 to act as a herpes receptor. EXPERIMENTAL DESIGN and RESULTS: A series of squamous cell carcinoma lines of increasing migratory and invasive potential, termed MG1-MG14, were selected by serial passages of murine SCC7 through Matrigel invasion chambers. Available cell surface nectin-1 was enhanced on the MG11 and MG14 cell lines in comparison to SCC7 as measured by cellular ELISA and immunofluorescence microscopy. A replication-competent, oncolytic herpes virus (NV1023) showed an increased ability to enter MG11 and MG14 cells as compared with SCC7 cells. Furthermore, MG11 and MG14 supported increased herpes viral replication and cytotoxicity over SCC7. For all three of the cell lines, viral entry assays revealed that the actively migrating cells were significantly more susceptible to herpes infection than the nonmigrating cells. CONCLUSIONS: These results show that malignant cells with highly migratory and invasive properties may exhibit increased cell surface nectin-1 availability, which may serve as a herpes viral receptor to enhance the efficacy of herpes oncolytic therapy. This finding has implications regarding patient selection for future clinical trials using these promising therapeutic vectors.     

Versatile prostate cancer treatment with inducible caspase and interleukin-12

To establish optimized conditions for immunity against prostate cancer, we compared the efficacy of multiple approaches in autochthonous and s.c. transgenic adenocarcinoma of the mouse prostate (TRAMP)-based models. Mice immunized with interleukin (IL)-12-containing apoptotic, but not necrotic TRAMP-C2 cell-based, vaccines were resistant to TRAMP-C2 tumor challenge and re-challenge, independently of the route of vaccination (s.c. or i.p.). Administration of gamma-irradiated TRAMP-C2 cells preinfected with adenovirus containing both B7-1 and IL-12 genes, unlike adenovirus containing B7-1 alone, considerably protected C57BL/6 mice from TRAMP-C2 tumor growth and extended the life span of TRAMP mice. Vaccines that included dendritic cells, instead of IL-12, were equally efficient. Whereas injections of ligand-inducible caspase-1- and IL-12-containing adenoviruses cured small s.c. TRAMP-C2 tumors, nanopump-regulated delivery of viruses led to elimination of much larger tumors. The antitumor immune responses involved CD4+-, CD8+-, and natural killer cells and were strengthened by increasing the number of vaccinations. Intraprostatic administration of inducible caspase-1- and IL-12-containing adenoviruses resulted in local cell death and improved survival of adenocarcinoma-bearing TRAMP mice. Thus, tumor cell apoptosis induced by caspase in situ and accompanied by IL-12 is efficient against prostate cancer in a preclinical model.     

Treatment of aggressive thyroid cancer with an oncolytic herpes virus

Although many thyroid cancers carry a favorable prognosis, there is a subgroup of patients with more aggressive histologies. Current therapies offer no significant survival benefit to patients with anaplastic thyroid carcinomas, which are considered fatal. Oncolytic herpes simplex viruses (HSVs) have potent antitumor effects against a variety of human malignancies. We assessed the activity of a replication-competent, attenuated, oncolytic HSV (NV1023) against 7 different thyroid cancers, including one papillary (NPA-187), one follicular (WRO82-1), one medullary (DRO81-1) and 4 anaplastic (DRO90-1, ARO, KAT-4C and KAT-18) cell lines. Only the follicular WRO82-1 line was resistant to NV1023 infection and cell lysis at a concentration of 5 viral pfu per cell (MOI 5). All other cell lines at MOI 5 demonstrated >95% infection in vitro at day 2 by X-gal staining and >88% cell death at day 4 by cytotoxicity assays. Even at MOI 0.1, 4 of these lines displayed complete cell death by day 7. Viral proliferation assays revealed that all of the nonfollicular cell lines supported logarithmic viral replication. Flank tumors of NPA-187, DRO81-1, DRO90-1 and ARO in athymic nude mice were treated with NV1023 (2 x 10(7) pfu). All NPA-187 tumors completely regressed following a single dose. DRO81-1 tumors demonstrated partial response with a single dose and significant improvement with 3 serial doses. ARO and DRO90-1 tumors showed a significant response following either single injection (54 +/- 22 and 292 +/- 138 mm3, respectively) or 3 serial injections (33 +/- 14 and 241 +/- 68 mm3, respectively) compared to saline injections (472 +/- 193 and 1,257 +/- 204 mm3, respectively) at day 20. These data suggest that herpes oncolytic therapy may be effective for the treatment of aggressive thyroid carcinomas and merits further investigation.     

Triple gene-deleted oncolytic herpes simplex virus vector double-armed with interleukin 18 and soluble B7-1 constructed by bacterial artificial chromosome-mediated system

Conditionally replicating herpes simplex virus type 1 (HSV-1) vectors are promising therapeutic agents for cancer. Certain antitumor functions may be added to oncolytic activities of recombinant HSV-1 vectors by inserting transgenes into the viral genome. Because conventional homologous recombination techniques had required time-consuming processes to create "armed" oncolytic HSV-1 vectors, we established an innovative construction system using bacterial artificial chromosome and two recombinase systems (Cre/loxP and FLPe/FRT). Using G47Delta, a safe and efficacious oncolytic HSV-1 with triple gene mutations, as the backbone, this system allowed a rapid generation of multiple vectors with desired transgenes inserted in the deleted ICP6 locus. Four oncolytic HSV-1 vectors, expressing murine interleukin 18 (mIL-18), soluble murine B7-1 [B7-1-immunoglobulin (B7-1-Ig)], both, or none, were created simultaneously within 3 months. In vitro, all newly created recombinant vectors exhibited virus yields and cytopathic effects similar to the parental G47Delta. In two immunocompetent mouse tumor models, TRAMP-C2 prostate cancer and Neuro2a neuroblastoma, the vector expressing both mIL-18 and B7-1-Ig showed a significant enhancement of antitumor efficacy via T-cell-mediated immune responses. The results show that "arming" with multiple transgenes can improve the efficacy of oncolytic HSV-1 vectors. The use of our system may facilitate the development and testing of various armed oncolytic HSV-1 vectors.     

Comparison of safety,delivery, and efficacy of two oncolytic herpes viruses (G207 and NV1020) for peritoneal cancer

G207 and NV1020 are two replication-competent, multimutant oncolytic herpes simplex viruses evaluated in the current studies for their anticancer effects in the treatment of gastric cancer. Deletion of both gamma(1)34.5 genes and inactivation of ICP6 (ribonucleotide reductase) allows G207 to selectively replicate within tumor cells. NV1020 is another attenuated recombinant herpes virus with deletions of the HSV joint region, with deletion of only one copy of the gamma(1)34.5 gene, and with the ICP6 gene intact. In vitro, both G207 and NV1020 effectively infected, replicated, and killed human gastric cancer cells, with NV1020 being more effective at lower concentrations of virus. In a murine xenograft model of peritoneally disseminated gastric cancer, both NV1020 and G207 reduced tumor burden when given intraperitoneally (i.p.) at higher doses. When viral doses were lowered or when advanced tumor was treated, i.p. NV1020 was superior to i.p. G207. In vitro viral replication and cytotoxicity predicted the in vivo antitumor response. Intravenous delivery of either G207 or NV1020 failed to reduce tumor burden, demonstrating the importance of regional therapy as treatment for compartmentalized malignancy. Both agents were safe for use in animals, and immunohistochemistry performed on mouse tissue revealed selective viral targeting of tumor. Oncolytic therapy using genetically engineered HSVs represents a promising strategy for peritoneal malignancies.     

Synergy of a herpes oncolytic virus and paclitaxel for anaplastic thyroid cancer.

PURPOSE: Novel therapeutic regimens are needed to improve the dismal outcomes of patients with anaplastic thyroid cancer (ATC). Oncolytic herpes simplex virus have shown promising activity against human ATC. We studied the application of oncolytic herpes simplex virus (G207 and NV1023) in combination with currently used chemotherapeutic drugs (paclitaxel and doxorubicin) for the treatment of ATC. EXPERIMENTAL DESIGN AND RESULTS: All four agents showed dose-response cytotoxicity in vitro for the human ATC cell lines KAT4 and DRO90-1. G207, combined with paclitaxel, showed synergistic cytotoxicity. Chou-Talalay combination indices ranged from 0.56 to 0.66 for KAT4, and 0.68 to 0.74 for DRO90-1 at higher affected fractions. Paclitaxel did not enhance G207 viral entry and early gene expression or G207 viral replication. Paclitaxel combined with G207 compared with single-agent treatment or controls showed significantly increased microtubule acetylation, mitotic arrest, aberrant chromatid separation, inhibition of metaphase to anaphase progression, and apoptosis. A single i.t. injection of G207 combined with biweekly i.p. paclitaxel injections in athymic nude mice bearing KAT4 flank tumors showed significantly reduced mean tumor volume (74 +/- 38 mm(3)) compared with G207 alone (388 +/- 109 mm(3)), paclitaxel alone (439 +/- 137 mm(3)), and control (520 +/- 160 mm(3)) groups at 16 days. There was no morbidity in vivo attributable to therapy. CONCLUSIONS: Mechanisms of paclitaxel antitumoral activity, including microtubule acetylation, mitotic block, and apoptosis, were enhanced by G207, which also has direct oncolytic effects. Combination of G207 and paclitaxel therapy is synergistic in treating ATC and holds promise for patients with this fatal disease.     

Secretable chaperone Grp170 enhances therapeutic activity of a novel tumor suppressor, mda-7/IL-24

Melanoma differentiation-associated gene-7 (mda-7)/interleukin-24 (IL-24) is a cancer-specific, apoptosis-inducing gene with broad-spectrum antitumor activity, making it an ideal candidate for a novel cancer gene therapy. A systemic and sustained antitumor immune response generated at the time of initial molecular-targeted therapy would provide additional clinical benefits in cancer patients, resulting in improved prevention of tumor recurrence. In this study, we explored the therapeutic efficacy of intratumoral delivery of a nonreplicating adenoviral vector encoding mda-7/IL-24 (Ad.mda-7) and a secretable form of endoplasmic reticulum resident chaperone grp170 (Ad.sgrp170), a potent immunostimulatory adjuvant and antigen carrier. Intratumoral administration of Ad.mda-7 in combination with Ad.sgrp170 was more effective in controlling growth of TRAMP-C2 prostate tumor compared with either Ad.mda-7 or Ad.sgrp170 treatment. Generation of systemic antitumor immunity was shown by enhanced protection against subsequent tumor challenge and improved control of distant tumors. The combined treatments enhanced antigen and tumor-specific T-cell response, as indicated by increased IFN-gamma production and cytolytic activity. Antibody depletion suggests that CD8(+) T cells may be involved in the antitumor effect of the dual molecule-targeted therapies. Therefore, introducing immunostimulatory chaperone grp170 in situ strongly promotes the "immunogenic" cell death when delivered to the mda-7/IL-24-induced apoptotic tumor cells, indicating that an improved anticancer efficacy may be achieved by concurrently targeting both tumor and immune compartments. Given multiple undefined antigens present endogenously within prostate cancer, these data provide a rationale for combining sgrp170-based vaccine strategy with mda-7/IL-24-targeted cancer therapy to induce durable systemic immunity.     

Triple-mutated oncolytic herpes virus for treating both fast- and slow-growing tumors

Oncolytic virus therapy has emerged as a promising treatment option against cancer. To date, oncolytic viruses have been developed for malignant tumors, but the need for this new therapeutic modality also exists for benign and slow-growing tumors. G47∆ is an oncolytic herpes simplex virus type 1 (HSV-1) with an enhanced replication capability highly selective to tumor cells due to genetically engineered, triple mutations in the γ34.5, ICP6 and α47 genes. To create a powerful, but safe oncolytic HSV-1 that replicates efficiently in tumors regardless of growth speed, we used a bacterial artificial chromosome system that allows a desired promoter to regulate the expression of the ICP6 gene in the G47∆ backbone. Restoration of the ICP6 function in a tumor-specific manner using the hTERT promoter led to a highly capable oncolytic HSV-1. T-hTERT was more efficacious in the slow-growing OS-RC-2 and DU145 tumors than the control viruses, while retaining a high efficacy in the fast-growing U87MG tumors. The safety features are also retained, as T-hTERT proved safe when inoculated into the brain of HSV-1 sensitive A/J mice. This new technology should facilitate the use of oncolytic HSV-1 for all tumors irrespective of growth speed.     

Relaxin-expressing, fiber chimeric oncolytic adenovirus prolongs survival of tumor-bearing mice

Selective replication of oncolytic viruses in tumor cells provides a promising approach for the treatment of human cancers. One of the limitations observed with oncolytic viruses currently used in the treatment of solid tumors is the inefficient spread of virus throughout the tumor mass following intratumoral injection. Data are presented showing that oncolytic adenoviruses expressing the relaxin gene and containing an Ad5/Ad35 chimeric fiber showed significantly enhanced transduction and increased virus spread throughout the tumor when compared with non-relaxin-expressing, Ad5-based viruses. The increased spread of such viruses throughout tumors correlated well with improved antitumor efficacy and overall survival in two highly metastatic tumor models. Furthermore, nonreplicating viruses expressing relaxin did not increase metastases, suggesting that high level expression of relaxin will not enhance metastatic spread of tumors. In summary, the data show that relaxin may play a role in rearranging matrix components within tumors, which helps recombinant oncolytic adenoviruses to spread effectively throughout the tumor mass and thereby increase the extent of viral replication within the tumor. Expressing relaxin from Ad5/Ad35 fiber chimeric adenoviruses may prove a potent and novel approach to treating patients with cancer.     

The potential of oncolytic virus therapy for pancreatic cancer

The objective of this paper was to review a new category of gene therapy using oncolytic viruses for the treatment of pancreatic cancer. The eligibility and feasibility of oncolytic virus therapy as a novel therapeutic agent against pancreatic cancer are discussed as well as basic research for clinical trials, including a historical perspective and the current status of these novel agents. Even combination therapy, such as surgery with radiation and chemotherapy, has not significantly improved the survival rate of pancreatic cancer. Recently, a clinical trial (phase I and II) using an oncolytic adenovirus, ONYX-015, was completed in patients with pancreatic cancer. The phase II trial yielded beneficial results (tumor reduction or stabilization) in about 50% of the patients. A phase I study of the efficacy of oncolytic herpes viruses, G207, OncoVEX GM-CSF, and 1716 against a variety of tumors has been completed, and G207 is in phase II trials for use against brain tumors. In addition, a phase I trial using the herpesvirus showed good tolerance at all dosages. We discuss the basic scientific principles and current results of the above clinical trials with respect to these oncolytic viruses, and then compare the relative advantages and disadvantages of adenoviruses and herpesviruses as oncolytic agents. We also review the published literature on newly developed oncolytic viruses. The concept of oncolytic therapy has been studied for a century. Recent technological developments have made these oncolytic viruses more tumor-specific by exploiting the tumor cell environments. In addition, these viruses have been reported to increase the immunosusceptibility of the tumor cells, and have been designed to express other genes to increase the susceptibility of tumor cells to other therapeutic agents. Oncolytic virus therapy certainly appears to be a feasible treatment for pancreatic cancer.