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Komane MD, Kayoka-Kabongo PN, Rutkowska DA. The Use of Plant Viral Nanoparticles in Cancer Biotherapy-A Review. Viruses 2025; 17:218. [PMID: 40006973 PMCID: PMC11860677 DOI: 10.3390/v17020218] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2024] [Revised: 01/24/2025] [Accepted: 01/29/2025] [Indexed: 02/27/2025] Open
Abstract
Cancer is a major global health problem that poses significant challenges. Conventional cancer therapies often have severe side effects, necessitating the development of novel therapeutic approaches that are more effective and less toxic. The utilization of plant viral nanoparticles is one of the more promising strategies for cancer biotherapy. Plant viral nanoparticles exhibit advantageous properties, including safety, high stability, rapid production and scalability, biocompatibility and biodegradability, structural uniformity, inherent immunogenicity, ease of modification and high update efficacy as well as lower cost implications, making them attractive vehicles for health applications. Various studies have demonstrated the efficacy of plant viral nanoparticles in targeted therapeutic drug/molecule delivery, tumor imaging and immunotherapy, highlighting their potential as a versatile platform for cancer biotherapy. The drawbacks of plant viral nanoparticles include their perceived ability to induce a hypersensitive/allergic immune response, non-well-defined regulatory approval processes as well as the reluctance of pharmaceutical companies to adapt their manufacturing processes to facilitate plant-based expression. This review discusses applications of plant virus-derived nanoparticles in cancer therapeutics and prospects for translating these findings into clinical practice.
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Affiliation(s)
- Mamorake Donty Komane
- Department of Agriculture and Animal Health, College of Agriculture and Environmental Sciences, University of South Africa, Science Campus, Private Bag X6 Florida 1710, Pretoria 0002, South Africa; (M.D.K.); (P.N.K.-K.)
| | - Prudence Ngalula Kayoka-Kabongo
- Department of Agriculture and Animal Health, College of Agriculture and Environmental Sciences, University of South Africa, Science Campus, Private Bag X6 Florida 1710, Pretoria 0002, South Africa; (M.D.K.); (P.N.K.-K.)
| | - Daria Anna Rutkowska
- Advanced Agriculture and Food Cluster, Council for Scientific and Industrial Research, P.O. Box 395, Pretoria 0001, South Africa
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Kawashita Y, Deb NJ, Garg M, Kabarriti R, Alfieri A, Takahashi M, Roy-Chowdhury J, Guha C. An autologous in situ tumor vaccination approach for hepatocellular carcinoma. 1. Flt3 ligand gene transfer increases antitumor effects of a radio-inducible suicide gene therapy in an ectopic tumor model. Radiat Res 2014; 182:191-200. [PMID: 24972258 DOI: 10.1667/rr13594.1] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/03/2022]
Abstract
Hepatocellular carcinoma (HCC) often presents as a diffuse or multifocal tumor making it difficult to control by surgery or radiation. Radio-inducible herpes simplex virus thymidine kinase (HSV-TK) gene therapy has been shown to enhance local tumor control after radiation therapy (RT), while limiting the expression of the transgene in the irradiated tumor tissues. To prevent liver tumor recurrence and control systemic disease while limiting the potential bystander toxicity of HSV-TK therapy, we proposed to stimulate endogenous dendritic cell (DC) proliferation with systemic adenovirus Flt3 ligand (Adeno-Flt3L) gene therapy, followed by primary tumor radiation therapy combined with a radio-inducible HSV-TK gene therapy. We hypothesized that adenovirus-expressing Flt3L gene therapy will stimulate DC proliferation, allowing the upregulated DCs to locally harness tumor antigens released from HSV-TK/RT-treated HCC cells, thereby converting irradiated tumors to an autologous in situ tumor vaccine in mice with primary liver tumors. To test this hypothesis, an expression vector of HSV-TK was constructed under the control of a radio-inducible promoter early-growth response (Egr-TK) and a recombinant adenovirus-expressing human Flt3L was constructed. The Adeno-Flt3L [10(9) plaque forming units (pfu)] was administered intravenously on days 1 and 8 after radiation therapy. The murine hepatoma cell line (BNL1ME) was stably transfected by Egr-TK or Egr-Null (encoding no therapeutic gene). Palpable tumors in BALB/c mice were treated with a localized dose of 25 Gy of radiation followed by ganciclovir (GCV, 100 mg/kg, 14 days). Four treatment cohorts were compared: Egr-Null/GCV + RT + Adeno-LacZ; Egr-Null/GCV + RT + Adeno-Flt3L; Egr-TK/GCV + RT + Adeno-LacZ; and Egr-TK/GCV + RT + Adeno-Flt3L. There was no primary tumor regression in the Egr-Null tumors after radiation therapy alone. In contrast, Egr-TK tumors had nearly complete tumor regression for 3 weeks after radiation therapy (P < 0.01), however, long-term follow-up demonstrated primary tumor recurrence and death secondary to pulmonary metastasis. Flt3L expression was confirmed by serum bioassay (mean = 88 ng/mL) in these animals and Western blotting of tissue culture medium in Adeno-Flt3L-infected BaF/huFlt3L cells. Radiation therapy with Adeno-Flt3L gene therapy effectively retarded primary tumor growth when compared to radiation therapy alone. The trimodality therapy (Egr-TK/GCV + RT + Adeno-Flt3L) was the most efficacious with 40% complete tumor regression (>100 days) and <20% pulmonary metastases, indicating the development of sustained antitumor immune response. These studies provide a rationale for triple modality therapies with radiation-inducible HSV-TK gene therapy and Adeno-Flt3L when used in combination with primary tumor radiation therapy for improved local and systemic control of HCC.
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Affiliation(s)
- Yujo Kawashita
- a Department of Surgery, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan
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Sen D, Balakrishnan B, Jayandharan GR. Cellular unfolded protein response against viruses used in gene therapy. Front Microbiol 2014; 5:250. [PMID: 24904562 PMCID: PMC4033601 DOI: 10.3389/fmicb.2014.00250] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/24/2014] [Accepted: 05/07/2014] [Indexed: 01/21/2023] Open
Abstract
Viruses are excellent vehicles for gene therapy due to their natural ability to infect and deliver the cargo to specific tissues with high efficiency. Although such vectors are usually "gutted" and are replication defective, they are subjected to clearance by the host cells by immune recognition and destruction. Unfolded protein response (UPR) is a naturally evolved cyto-protective signaling pathway which is triggered due to endoplasmic reticulum (ER) stress caused by accumulation of unfolded/misfolded proteins in its lumen. The UPR signaling consists of three signaling pathways, namely PKR-like ER kinase, activating transcription factor 6, and inositol-requiring protein-1. Once activated, UPR triggers the production of ER molecular chaperones and stress response proteins to help reduce the protein load within the ER. This occurs by degradation of the misfolded proteins and ensues in the arrest of protein translation machinery. If the burden of protein load in ER is beyond its processing capacity, UPR can activate pro-apoptotic pathways or autophagy leading to cell death. Viruses are naturally evolved in hijacking the host cellular translation machinery to generate a large amount of proteins. This phenomenon disrupts ER homeostasis and leads to ER stress. Alternatively, in the case of gutted vectors used in gene therapy, the excess load of recombinant vectors administered and encountered by the cell can trigger UPR. Thus, in the context of gene therapy, UPR becomes a major roadblock that can potentially trigger inflammatory responses against the vectors and reduce the efficiency of gene transfer.
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Affiliation(s)
- Dwaipayan Sen
- Department of Hematology, Christian Medical College Vellore, India
| | | | - Giridhara R Jayandharan
- Department of Hematology, Christian Medical College Vellore, India ; Centre for Stem Cell Research, Christian Medical College Vellore, India
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Kantor B, Bailey RM, Wimberly K, Kalburgi SN, Gray SJ. Methods for gene transfer to the central nervous system. ADVANCES IN GENETICS 2014; 87:125-97. [PMID: 25311922 DOI: 10.1016/b978-0-12-800149-3.00003-2] [Citation(s) in RCA: 73] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Gene transfer is an increasingly utilized approach for research and clinical applications involving the central nervous system (CNS). Vectors for gene transfer can be as simple as an unmodified plasmid, but more commonly involve complex modifications to viruses to make them suitable gene delivery vehicles. This chapter will explain how tools for CNS gene transfer have been derived from naturally occurring viruses. The current capabilities of plasmid, retroviral, adeno-associated virus, adenovirus, and herpes simplex virus vectors for CNS gene delivery will be described. These include both focal and global CNS gene transfer strategies, with short- or long-term gene expression. As is described in this chapter, an important aspect of any vector is the cis-acting regulatory elements incorporated into the vector genome that control when, where, and how the transgene is expressed.
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Affiliation(s)
- Boris Kantor
- Department of Pharmacology, Physiology, and Neuroscience, University of South Carolina, Columbia, SC, USA
| | - Rachel M Bailey
- Gene Therapy Center, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
| | - Keon Wimberly
- Gene Therapy Center, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
| | - Sahana N Kalburgi
- Gene Therapy Center, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
| | - Steven J Gray
- Gene Therapy Center, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA; Department of Ophthalmology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
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Stancevic B, Varda-Bloom N, Cheng J, Fuller JD, Rotolo JA, García-Barros M, Feldman R, Rao S, Weichselbaum RR, Harats D, Haimovitz-Friedman A, Fuks Z, Sadelain M, Kolesnick R. Adenoviral transduction of human acid sphingomyelinase into neo-angiogenic endothelium radiosensitizes tumor cure. PLoS One 2013; 8:e69025. [PMID: 23936314 PMCID: PMC3732255 DOI: 10.1371/journal.pone.0069025] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/28/2013] [Accepted: 06/03/2013] [Indexed: 12/31/2022] Open
Abstract
These studies define a new mechanism-based approach to radiosensitize tumor cure by single dose radiotherapy (SDRT). Published evidence indicates that SDRT induces acute microvascular endothelial apoptosis initiated via acid sphingomyelinase (ASMase) translocation to the external plasma membrane. Ensuing microvascular damage regulates radiation lethality of tumor stem cell clonogens to effect tumor cure. Based on this biology, we engineered an ASMase-producing vector consisting of a modified pre-proendothelin-1 promoter, PPE1(3x), and a hypoxia-inducible dual-binding HIF-2α-Ets-1 enhancer element upstream of the asmase gene, inserted into a replication-deficient adenovirus yielding the vector Ad5H2E-PPE1(3x)-ASMase. This vector confers ASMase over-expression in cycling angiogenic endothelium in vitro and within tumors in vivo, with no detectable enhancement in endothelium of normal tissues that exhibit a minute fraction of cycling cells or in non-endothelial tumor or normal tissue cells. Intravenous pretreatment with Ad5H2E-PPE1(3x)-ASMase markedly increases SDRT cure of inherently radiosensitive MCA/129 fibrosarcomas, and converts radiation-incurable B16 melanomas into biopsy-proven tumor cures. In contrast, Ad5H2E-PPE1(3x)-ASMase treatment did not impact radiation damage to small intestinal crypts as non-dividing small intestinal microvessels did not overexpress ASMase and were not radiosensitized. We posit that combination of genetic up-regulation of tumor microvascular ASMase and SDRT provides therapeutic options for currently radiation-incurable human tumors.
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Affiliation(s)
- Branka Stancevic
- Laboratory of Signal Transduction, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
| | - Nira Varda-Bloom
- Laboratory of Signal Transduction, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
| | - Jin Cheng
- Laboratory of Signal Transduction, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
| | - John D. Fuller
- Laboratory of Signal Transduction, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
| | - Jimmy A. Rotolo
- Department of Radiation Oncology, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
| | - Mónica García-Barros
- Laboratory of Signal Transduction, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
| | - Regina Feldman
- Department of Radiation Oncology, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
| | - Shyam Rao
- Department of Radiation Oncology, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
| | - Ralph R. Weichselbaum
- Department of Radiation and Cellular Oncology, University of Chicago, and the Ludwig Center for Metastasis Research, Chicago, Illinois, United States of America
| | | | - Adriana Haimovitz-Friedman
- Department of Radiation Oncology, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
| | - Zvi Fuks
- Department of Radiation Oncology, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
| | - Michel Sadelain
- Center for Cell Engineering, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
| | - Richard Kolesnick
- Laboratory of Signal Transduction, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America
- * E-mail:
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Wainwright DA, Nigam P, Thaci B, Dey M, Lesniak MS. Recent developments on immunotherapy for brain cancer. Expert Opin Emerg Drugs 2012; 17:181-202. [PMID: 22533851 DOI: 10.1517/14728214.2012.679929] [Citation(s) in RCA: 52] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/12/2023]
Abstract
INTRODUCTION Brain tumors are a unique class of cancers since they are anatomically shielded from normal immunosurveillance by the blood-brain barrier, lack a normal lymphatic drainage system and reside in a potently immunosuppressive environment. Of the primary brain cancers, glioblastoma multiforme (GBM) is the most common and aggressive in adults. Although treatment options include surgery, radiation and chemotherapy, the average lifespan of GBM patients remains at only 14.6 months post-diagnosis. AREAS COVERED A review of key cellular and molecular immune system mediators in the context of brain tumors including TGF-β, cytotoxic T cells, Tregs, CTLA-4, PD-1 and IDO is discussed. In addition, prognostic factors, currently utilized immunotherapeutic strategies, ongoing clinical trials and a discussion of new or potential immunotherapies for brain tumor patients are considered. EXPERT OPINION Current drugs that improve the quality of life and overall survival in patients with brain tumors, especially for GBM, are poorly effective. This disease requires a reanalysis of currently accepted treatment strategies, as well as newly designed approaches. Here, we review the fundamental aspects of immunosuppression in brain tumors, new and promising immunotherapeutic drugs as well as combinatorial strategies that focus on the simultaneous inhibition of immunosuppressive hubs, both in immune and brain tumor cells, which is critical to consider for achieving future success for the treatment of this devastating disease.
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Castro MG, Candolfi M, Kroeger K, King GD, Curtin JF, Yagiz K, Mineharu Y, Assi H, Wibowo M, Ghulam Muhammad AKM, Foulad D, Puntel M, Lowenstein PR. Gene therapy and targeted toxins for glioma. Curr Gene Ther 2011; 11:155-80. [PMID: 21453286 DOI: 10.2174/156652311795684722] [Citation(s) in RCA: 63] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2011] [Accepted: 03/08/2011] [Indexed: 12/12/2022]
Abstract
The most common primary brain tumor in adults is glioblastoma. These tumors are highly invasive and aggressive with a mean survival time of 15-18 months from diagnosis to death. Current treatment modalities are unable to significantly prolong survival in patients diagnosed with glioblastoma. As such, glioma is an attractive target for developing novel therapeutic approaches utilizing gene therapy. This review will examine the available preclinical models for glioma including xenographs, syngeneic and genetic models. Several promising therapeutic targets are currently being pursued in pre-clinical investigations. These targets will be reviewed by mechanism of action, i.e., conditional cytotoxic, targeted toxins, oncolytic viruses, tumor suppressors/oncogenes, and immune stimulatory approaches. Preclinical gene therapy paradigms aim to determine which strategies will provide rapid tumor regression and long-term protection from recurrence. While a wide range of potential targets are being investigated preclinically, only the most efficacious are further transitioned into clinical trial paradigms. Clinical trials reported to date are summarized including results from conditionally cytotoxic, targeted toxins, oncolytic viruses and oncogene targeting approaches. Clinical trial results have not been as robust as preclinical models predicted; this could be due to the limitations of the GBM models employed. Once this is addressed, and we develop effective gene therapies in models that better replicate the clinical scenario, gene therapy will provide a powerful approach to treat and manage brain tumors.
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Affiliation(s)
- Maria G Castro
- Gene Therapeutics Research Institute, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA.
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Uyama A, Kondoh T, Nariyama N, Umetani K, Fukumoto M, Shinohara K, Kohmura E. A narrow microbeam is more effective for tumor growth suppression than a wide microbeam: an in vivo study using implanted human glioma cells. JOURNAL OF SYNCHROTRON RADIATION 2011; 18:671-678. [PMID: 21685685 PMCID: PMC3286866 DOI: 10.1107/s090904951101185x] [Citation(s) in RCA: 28] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 10/26/2010] [Accepted: 03/30/2011] [Indexed: 05/30/2023]
Abstract
The tumoricidal mechanisms of microbeam radiation therapy, and the more recently proposed minibeam radiation therapy, for the treatment of brain tumors are as yet unclear. Moreover, from among the various parameters of beam geometry the impact of changing the beam width is unknown. In this study, suppression of tumor growth in human glioma cells implanted in mice was evaluated experimentally using microbeams of two different widths: a conventional narrow beam (20 µm width, 100 µm center-to-center distance) and a wide beam (100 µm width, 500 µm center-to-center distance). The tumor growth ratio was compared and acute cell death was studied histologically. With cross-planar irradiation, tumor growth was significantly suppressed between days 4 and 28 after 20 µm microbeam irradiation, whereas tumor growth was suppressed, and not significantly so, only between days 4 and 18 after 100 µm microbeam irradiation. Immunohistochemistry using TUNEL staining showed no increase in TUNEL-positive cells with either microbeam at 24 and 72 h post-irradiation. The 20 µm microbeam was found to be more tumoricidal than the 100 µm microbeam, and the effect was not related to apoptotic cell death. The underlying mechanism may be functional tissue deterioration rather than direct cellular damage in the beam path.
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Affiliation(s)
- Atsushi Uyama
- Department of Neurosurgery, Kobe University Graduate School of Medicine, Kobe, Hyogo, Japan.
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van Putten EH, Dirven CM, van den Bent MJ, Lamfers ML. Sitimagene ceradenovec: a gene-based drug for the treatment of operable high-grade glioma. Future Oncol 2011; 6:1691-710. [PMID: 21142657 DOI: 10.2217/fon.10.134] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023] Open
Abstract
The field of gene therapy for malignant glioma has made important advances since the first gene transfer studies were performed 20 years ago. Multiple Phase I/II trials and two Phase III trials have been performed and have demonstrated the feasibility and safety of intratumoral vector delivery in the brain. Sitimagene ceradenovec is an adenoviral vector encoding the herpes simplex thymidine kinase gene, developed by Ark Therapeutics Group plc (UK and Finland) for the treatment of patients with operable high-grade glioma. In preclinical and Phase I/II clinical studies, sitimagene ceradenovec exhibited a significant increase in survival. Although the preliminary results of a Phase III clinical study demonstrated a significant positive effect of sitimagene ceradenovec treatment on time to reintervention or death when compared with standard care treatment (hazard ratio: 1.43; 95% CI: 1.06-1.93; p < 0.05), the European Committee for Medicinal Products for Human Use did not consider the data to provide sufficient evidence of clinical benefit. Further clinical evaluation, powered to demonstrate a benefit on a robust end point, is required. This article focuses on sitimagene ceradenovec and provides an overview of the developments in the field of gene therapy for malignant glioma.
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Affiliation(s)
- Erik Hp van Putten
- Department of Neurosurgery, Erasmus University Medical Center, PO Box 2040, 3000 CA Rotterdam, The Netherlands.
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White E, Bienemann A, Megraw L, Bunnun C, Gill S. Evaluation and optimization of the administration of a selectively replicating herpes simplex viral vector to the brain by convection-enhanced delivery. Cancer Gene Ther 2011; 18:358-69. [PMID: 21372854 DOI: 10.1038/cgt.2011.2] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
The direct intraparenchymal administration of oncolytic viral vectors by convection-enhanced delivery (CED) represents a promising new treatment strategy for malignant gliomas. However, there is no evidence to suggest that oncolytic viruses as large as herpes simplex virus-1 (HSV-1) can be administered by CED, as this has not been systematically examined in an animal model. In this study, the administration of a herpes simplex viral vector, HSV1, has been evaluated in detail in the gray and white matter of both rat and pig models, using high flow-rate infusions, co-infusing heparin or preinfusing the tissue with an isotonic albumin solution. Rat HSV-1 infusions at both slow (0.5 μl min(-1)) and high infusion rates (2.5 μl min(-1)) led to extensive tissue damage and negligible cell transduction. Co-infusion with heparin led to extensive hemorrhage. Preinfusion of tissue with an isotonic albumin solution facilitated widespread vector distribution and cell transduction in white matter only. Using this approach in pig brain led to widespread vector distribution with extensive transduction of astrocytes and activated microglia. In rat brain, enhanced green fluorescent protein expression peaked 48 h after vector administration and was associated with a vigorous immune response. These findings indicate that direct infusions of HSV-1-based viral vectors into the brain lead to minimal vector distribution, negligible cell transduction and extensive damage. Tissue preinfusion with an isotonic solution prior to vector administration represents an effective technique for achieving widespread HSV-1 distribution.
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Affiliation(s)
- E White
- Department of Neurosurgery, Frenchay Hospital, Bristol, UK
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Marconi P, Argnani R, Epstein AL, Manservigi R. HSV as a vector in vaccine development and gene therapy. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2010; 655:118-44. [PMID: 20047039 DOI: 10.1007/978-1-4419-1132-2_10] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/23/2022]
Abstract
The very deep knowledge acquired on the genetics and molecular biology of herpes simplex virus (HSV), major human pathogen whose lifestyle is based on a long-term dual interaction with the infected host characterized by the existence of lytic and latent infections, has allowed the development of potential vectors for several applications in human healthcare. These include delivery and expression of human genes to cells of the nervous system, selective destruction of cancer cells, prophylaxis against infection with HSV or other infectious diseases and targeted infection of specific tissues or organs. Three different classes of vectors can be derived from HSV-1: replication-competent attenuated vectors, replication-incompetent recombinant vectors and defective helper-dependent vectors known as amplicons. This chapter highlights the current knowledge concerning design, construction and recent applications, as well as the potential and current limitations of the three different classes of HSV-1-based vectors.
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Affiliation(s)
- Peggy Marconi
- Department of Experimental and Diagnostic Medicine-Section of Microbiology, University of Ferrara, Via Luigi Borsari 46, Ferrara, 44100, Italy.
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Manservigi R, Argnani R, Marconi P. HSV Recombinant Vectors for Gene Therapy. Open Virol J 2010; 4:123-56. [PMID: 20835362 DOI: 10.2174/1874357901004030123] [Citation(s) in RCA: 29] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2009] [Revised: 03/13/2010] [Accepted: 03/31/2010] [Indexed: 12/16/2022] Open
Abstract
The very deep knowledge acquired on the genetics and molecular biology of herpes simplex virus (HSV), has allowed the development of potential replication-competent and replication-defective vectors for several applications in human healthcare. These include delivery and expression of human genes to cells of the nervous systems, selective destruction of cancer cells, prophylaxis against infection with HSV or other infectious diseases, and targeted infection to specific tissues or organs. Replication-defective recombinant vectors are non-toxic gene transfer tools that preserve most of the neurotropic features of wild type HSV-1, particularly the ability to express genes after having established latent infections, and are thus proficient candidates for therapeutic gene transfer settings in neurons. A replication-defective HSV vector for the treatment of pain has recently entered in phase 1 clinical trial. Replication-competent (oncolytic) vectors are becoming a suitable and powerful tool to eradicate brain tumours due to their ability to replicate and spread only within the tumour mass, and have reached phase II/III clinical trials in some cases. The progress in understanding the host immune response induced by the vector is also improving the use of HSV as a vaccine vector against both HSV infection and other pathogens. This review briefly summarizes the obstacle encountered in the delivery of HSV vectors and examines the various strategies developed or proposed to overcome such challenges.
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Affiliation(s)
- Roberto Manservigi
- Department of Experimental and Diagnostic Medicine - Section of Microbiology, University of Ferrara, Via Luigi Borsari 46, 44100 Ferrara, Italy
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13
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Abstract
The very deep knowledge acquired on the genetics and molecular biology of herpes simplex virus (HSV), has allowed the development of potential replication-competent and replication-defective vectors for several applications in human healthcare. These include delivery and expression of human genes to cells of the nervous systems, selective destruction of cancer cells, prophylaxis against infection with HSV or other infectious diseases, and targeted infection to specific tissues or organs. Replication-defective recombinant vectors are non-toxic gene transfer tools that preserve most of the neurotropic features of wild type HSV-1, particularly the ability to express genes after having established latent infections, and are thus proficient candidates for therapeutic gene transfer settings in neurons. A replication-defective HSV vector for the treatment of pain has recently entered in phase 1 clinical trial. Replication-competent (oncolytic) vectors are becoming a suitable and powerful tool to eradicate brain tumours due to their ability to replicate and spread only within the tumour mass, and have reached phase II/III clinical trials in some cases. The progress in understanding the host immune response induced by the vector is also improving the use of HSV as a vaccine vector against both HSV infection and other pathogens. This review briefly summarizes the obstacle encountered in the delivery of HSV vectors and examines the various strategies developed or proposed to overcome such challenges.
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Affiliation(s)
- Roberto Manservigi
- Department of Experimental and Diagnostic Medicine - Section of Microbiology, University of Ferrara, Via Luigi Borsari 46, 44100 Ferrara, Italy
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Hong CS, Fellows W, Niranjan A, Alber S, Watkins S, Cohen JB, Glorioso JC, Grandi P. Ectopic matrix metalloproteinase-9 expression in human brain tumor cells enhances oncolytic HSV vector infection. Gene Ther 2010; 17:1200-5. [PMID: 20463757 DOI: 10.1038/gt.2010.66] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Oncolytic herpes simplex virus (oHSV) vectors have shown promise in the treatment of patients with recurrent brain tumors although few complete responses have accrued. Impediments to effective therapy include limited vector distribution on delivery, a consequence of injected virion particle trapping in the tumor extracellular matrix (ECM). To enhance virus delivery and spread, we investigated the use of the matrix metalloproteinase-9 (MMP-9) as a means to degrade collagen type IV, a major component of the ECM and basement membranes of gliomas that is absent in normal brain tissue. SK-N-AS neuroblastoma cells were transduced for constitutive, elevated expression of MMP-9, which did not enhance tumor cell migration in vitro or tumor progression in a murine xenograft brain tumor model. MMP-9 expression improved the distribution and infection of oHSV vectors in spheroid model in vitro. Furthermore, MMP9 induced a vector infection over larger areas of brain tumors in vivo. These results suggest that vector delivery and distribution in vivo can be improved by compromising the ECM, potentially enhancing oncolytic efficacy.
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Affiliation(s)
- C-S Hong
- Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
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Liang B, He ML, Chan CY, Chen YC, Li XP, Li Y, Zheng D, Lin MC, Kung HF, Shuai XT, Peng Y. The use of folate-PEG-grafted-hybranched-PEI nonviral vector for the inhibition of glioma growth in the rat. Biomaterials 2009; 30:4014-20. [PMID: 19427690 DOI: 10.1016/j.biomaterials.2009.04.011] [Citation(s) in RCA: 90] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2009] [Accepted: 04/13/2009] [Indexed: 12/23/2022]
Abstract
Combined treatment using nonviral agent-mediated enzyme/prodrug therapy and immunotherapy had been proposed as a powerful alternative method of cancer therapy. The present study was aimed to evaluate the cytotoxicity in vitro and the therapeutic efficacy in vivo when the cytosine deaminase/5-fluorocytosine (CD/5-FC) and TNF-related apoptosis-inducing ligand (TRAIL) genes were jointly used against rat C6 glioma cells. The potency of the FA-PEG-PEI used as a nonviral vector was tested in the FR-expressed C6 glioma cells and Wistar rats. The C6 glioma cells and animal model were treated by the combined application of FA-PEG-PEI/pCD/5-FC and FA-PEG-PEI/pTRAIL. The antitumor effect was evaluated by survival assays and tumor volume. This study revealed a significant increase of cytotoxicity in vitro following the combined application of FA-PEG-PEI/pCD/5-FC and FA-PEG-PEI/pTRAIL treatments in C6 glioma cells. Animal studies showed a significant growth inhibition of the C6 glioma xenografts using the combined treatment. These results demonstrated that the combined treatment generated additive cytotoxic effect in C6 glioma cells in both in vitro and in vivo conditions, and indicated that such treatment method using both enzyme/prodrug therapy and TRAIL immunotherapy might be a promising therapeutic strategy in treating glioma.
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Affiliation(s)
- Bing Liang
- Department of Neurology, The Second Affiliated Hospital, Sun Yat-sen University, No. 107 West Road of Riverside, Guangzhou 510120, China
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16
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Hadjipanayis CG, Fellows-Mayle W, Deluca NA. Therapeutic efficacy of a herpes simplex virus with radiation or temozolomide for intracranial glioblastoma after convection-enhanced delivery. Mol Ther 2008; 16:1783-8. [PMID: 18728637 DOI: 10.1038/mt.2008.185] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/16/2023] Open
Abstract
The herpes simplex virus-1 (HSV-1)-infected cell protein 0 (ICP0) is an E3 ubiquitin ligase implicated in cell cycle arrest and DNA repair inhibition. Convection-enhanced delivery (CED) of either the replication-defective, ICP0-producing HSV-1 mutant, d106, or the recombinant d109, devoid of all viral genome expression, was performed to determine the in vivo efficacy of ICP0 in combination with ionizing radiation (IR) or systemic temozolomide (TMZ) in the treatment of glioblastoma multiforme (GBM). Intracranial U87-MG xenografts were established in athymic nude mice. Animal survival was determined after mice underwent intracranial CED of either the replication-defective d106 or d109 viruses, or Hanks' balanced salt solution (HBSS), before a single session of whole-brain irradiation or TMZ treatment. Median survival for animals that underwent treatment with HBSS alone, d109 alone, d106 alone, HBSS + IR, HBSS + TMZ, d109 + IR, d106 + IR, and d106 + TMZ was 28, 35, 41, 39, 44, 39, 68 (P < 0.01), and 66 days (P < 0.01), respectively. Intracerebral d106 CED resulted in a significant increase in athymic nude mouse survival when combined with IR or TMZ. d106 CED allows for distribution of HSV-1 in human GBM xenografts and persistent viral infection.
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Affiliation(s)
- Costas G Hadjipanayis
- Department of Neurological Surgery, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA.
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17
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Yamini B, Yu X, Pytel P, Galanopoulos N, Rawlani V, Veerapong J, Bickenbach K, Weichselbaum RR. Adenovirally delivered tumor necrosis factor-alpha improves the antiglioma efficacy of concomitant radiation and temozolomide therapy. Clin Cancer Res 2007; 13:6217-23. [PMID: 17947489 DOI: 10.1158/1078-0432.ccr-07-1421] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
PURPOSE Treatment of malignant glioma involves concomitant temozolomide and ionizing radiation (IR). Nevertheless, overall patient survival remains poor. This study was designed to evaluate if addition of Ad.Egr-tumor necrosis factor (TNF), a replication defective adenovector encoding a cDNA for TNF-alpha, to temozolomide and IR can improve overall antiglioma effect. EXPERIMENTAL DESIGN The efficacy of combination treatment with Ad.Egr-TNF, IR, and temozolomide was assessed in two glioma xenograft models. Animal toxicity and brain histopathology after treatment were also examined. In addition, in an attempt to explain the antitumor interaction between these treatments, the activation status of the transcription factor nuclear factor-kappaB was examined. RESULTS Triple therapy (Ad.Egr-TNF, IR, and temozolomide) leads to significantly increased survival in mice bearing glioma xenografts compared with dual treatment. Fifty percent of animals treated with the triple regimen survive for >130 days. Pathologic examination shows that triple therapy leads to a complete response with formation of a collagenous scar. No significant change in myelination pattern is noted after triple therapy, compared with any double treatment. Treatment of intracranial glioma bearing mice with Ad.Egr-TNF and IR leads to cachexia and poor feeding that does not improve, whereas triple therapy results in less toxicity, which improves over 21 days. Both Ad.Egr-TNF and IR activate nuclear factor-kappaB, and temozolomide inhibits this activity in an inhibitor of kappaBalpha (IkappaBalpha)-independent manner. CONCLUSION This work shows that the addition of adenoviral TNF-alpha gene delivery to temozolomide and IR significantly improves antiglioma efficacy and illustrates a potential new treatment regimen for use in patients with malignant glioma.
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Affiliation(s)
- Bakhtiar Yamini
- Section of Neurosurgery, Department of Surgery, Pritzker School of Medicine, The University of Chicago, Chicago, Illinois 60637, USA.
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Advani SJ, Weichselbaum RR, Chmura SJ. Enhancing Radiotherapy With Genetically Engineered Viruses. J Clin Oncol 2007; 25:4090-5. [PMID: 17827458 DOI: 10.1200/jco.2007.12.2739] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Concurrent radiotherapy and chemotherapy have been used to treat a variety of tumors to improve local control and overall survival. Gene therapy strategies represent a novel means to further improve the therapeutic ratio of ionizing radiation. Cancer gene therapy strategies in clinical trials include the use of replication-defective shuttle vectors to deliver exogenous genes and replication-competent oncolytic viruses. This review focuses on these approaches in the context of radiotherapy and radiochemotherapy. In the shuttle vector approach, exogenous gene products that enhance ionizing radiation–mediated tumor cell destruction have been selected. Moreover, the expression of exogenous genes encoding therapeutic proteins can be regulated through the use of ionizing radiation–enhanced promoters. Also, genetically engineered attenuated replication-competent viruses have been investigated in clinical trials. Preclinical data indicate that ionizing radiation interacts with replication-competent oncolytic viruses to enhance viral replication and tumor destruction. Here, we review the background preclinical and current clinical data utilizing gene therapy with radiotherapy.
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Affiliation(s)
- Sunil J Advani
- Department of Radiation and Cellular Oncology, Center for Molecular Medicine, University of Chicago, Chicago, IL, USA
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19
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Haghikia A, Ladage K, Lafênetre P, Haghikia A, Hinkerohe D, Smikalla D, Haase CG, Dermietzel R, Faustmann PM. Intracellular application of TNF-alpha impairs cell to cell communication via gap junctions in glioma cells. J Neurooncol 2007; 86:143-52. [PMID: 17690839 DOI: 10.1007/s11060-007-9462-8] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2007] [Accepted: 07/13/2007] [Indexed: 12/23/2022]
Abstract
Human gliomas are the most common class of brain neoplasm. In order to better characterize their response to inflammation, we evaluated the influence of tumor necrosis factor alpha (TNF-alpha) on the coupling behaviour and the membrane resting potential (MRP) of glioma cells (F98 glioma cell line) compared to primary astrocytes. In contrast to cultured primary astrocytes which exhibited a profound inhibition of gap junction mediated intercellular communication (GJIC), extracellular exposure of TNF-alpha to F98 glioma cells gained no effect on the functional coupling. Whereas, intracellular application of TNF-alpha into the glioma cells elicited similar effects as those found in primary astrocytes indicating a compromised accessibility of the TNF-alpha receptor in F98 cells. Western blotting, immunocytochemical staining and real time RT PCR analysis revealed a differential expression and distribution of TNF-alpha receptor 1 (TNFR1) in the glioma cells. Connexin 43 (Cx43) is the major astrocytic gap junction protein which when phosphorylated has been shown to reveal altered gating properties. Here we show that TNF-alpha increases the level of phosphorylated Cx43 in primary astrocytes but not in the F98 glioma cells. Our observations could account for the decreased regulatory effects of TNF-alpha on GJIC of F98 glioma cells.
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Affiliation(s)
- Aiden Haghikia
- Department of Neurology, St. Josef-Hospital Bochum, Ruhr-University Bochum, Gudrunstrasse 56, Bochum, Germany.
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20
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Berto E, Bozac A, Volpi I, Lanzoni I, Vasquez F, Melara N, Manservigi R, Marconi P. Antitumor effects of non-replicative herpes simplex vectors expressing antiangiogenic proteins and thymidine kinase on Lewis lung carcinoma establishment and growth. Cancer Gene Ther 2007; 14:791-801. [PMID: 17557110 DOI: 10.1038/sj.cgt.7701058] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
There is growing evidence that combinations of antiangiogenic proteins with other antineoplastic treatments such as chemo- or radiotherapy and suicide genes-mediated tumor cytotoxicity lead to synergistic effects. In the present work, we tested the activity of two non-replicative herpes simplex virus (HSV)-1-based vectors, encoding human endostatin::angiostatin or endostatin::kringle5 fusion proteins in combination with HSV-1 thymidine kinase (TK) molecule, on endothelial cells (ECs) and Lewis lung carcinoma (LLC) cells. We observed a significant reduction of the in vitro growth, migration and tube formation by primary ECs upon direct infection with the two recombinant vectors or cultivation with conditioned media obtained from the vector-infected LLC cells. Moreover, direct cytotoxic effect of HSV-1 TK on both LLC and ECs was demonstrated. We then tested the vectors in vivo in two experimental settings, that is, LLC tumor growth or establishment, in C57BL/6 mice. The treatment of pre-established subcutaneous tumors with the recombinant vectors with ganciclovir (GCV) induced a significant reduction of tumor growth rate, while the in vitro infection of LLC cells with the antiangiogenic vectors before their implantation in mice flanks, either in presence or absence of GCV, completely abolished the tumor establishment.
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Affiliation(s)
- E Berto
- Department of Experimental and Diagnostic Medicine, Section of Microbiology, University of Ferrara, Ferrara, Italy
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21
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Oh BC, Liu CY, Wang MY, Pagnini PG, Yu C, Apuzzo MLJ. Stereotactic Radiosurgery: Adjacent Tissue Injury and Response after High-Dose Single Fraction Radiation—Part II: Strategies for Therapeutic Enhancement, Brain Injury Mitigation, and Brain Injury Repair. Neurosurgery 2007; 60:799-814; discussion 799-814. [PMID: 17460515 DOI: 10.1227/01.neu.0000255454.28225.5c] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022] Open
Abstract
Abstract
IN THE FIRST part of this series, we reviewed the histological, radiographic, and molecular data gathered regarding the brain parenchymal response to radiosurgery and suggested future studies that could enhance our understanding of the topic. With this article, we begin by addressing methods of potentiating the effect of radiosurgery on target lesions of the central nervous system. Much of the work on potentiating the effects of cranial radiation has been performed in the field of whole-brain radiotherapy. Data from Phase III trials evaluating the efficacy of various agents as radiosensitizers or radioenhancers in whole-brain radiotherapy are reviewed, and trials for investigating certain agents as enhancers of radiosurgery are suggested. The roles of gene therapy and nanotechnology in enhancing the therapeutic efficacy of radiosurgery are then addressed. Focus is then shifted to a discussion of strategies of protecting healthy tissue from the potentially deleterious aspects of the brain's response to radiosurgery that were presented in the first article of this series. Finally, comments are made regarding the role of neural progenitor or stem cells in the repair of radiation-induced brain injury after radiosurgery. The importance of both the role of the extracellular matrix and properly directed axonal regrowth leading to appropriate target reinnervation is highlighted.
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22
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Colen CB, Seraji-Bozorgzad N, Marples B, Galloway MP, Sloan AE, Mathupala SP. Metabolic remodeling of malignant gliomas for enhanced sensitization during radiotherapy: an in vitro study. Neurosurgery 2007; 59:1313-23; discussion 1323-4. [PMID: 17277695 PMCID: PMC3385862 DOI: 10.1227/01.neu.0000249218.65332.bf] [Citation(s) in RCA: 57] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022] Open
Abstract
OBJECTIVE To investigate a novel method to enhance radiosensitivity of gliomas via modification of metabolite flux immediately before radiotherapy. Malignant gliomas are highly glycolytic and produce copious amounts of lactic acid, which is effluxed to the tumor microenvironment via lactate transporters. We hypothesized that inhibition of lactic acid efflux would alter glioma metabolite profiles, including those that are radioprotective. H magnetic resonance spectroscopy (MRS) was used to quantify key metabolites, including those most effective for induction of low-dose radiation-induced cell death. METHODS We inhibited lactate transport in U87-MG gliomas with alpha-cyano-4-hydroxycinnamic acid (ACCA). Flow cytometry was used to assess induction of cell death in treated cells. Cells were analyzed by MRS after ACCA treatment. Control and treated cells were subjected to low-dose irradiation, and the surviving fractions of cells were determined by clonogenic assays. RESULTS MRS revealed changes to intracellular lactate on treatment with ACCA. Significant decreases in the metabolites taurine, glutamate, glutathione, alanine, and glycine were observed, along with inversion of the choline/phosphocholine profile. On exposure to low-dose radiation, ACCA-pretreated U-87MG cells underwent rapid morphological changes, which were followed by apoptotic cell death. CONCLUSION Inhibition of lactate efflux in malignant gliomas results in alterations of glycolytic metabolism, including decreased levels of the antioxidants taurine and glutathione and enhanced radiosensitivity of ACCA-treated cells. Thus, in situ application of lactate transport inhibitors such as ACCA as a novel adjunctive therapeutic strategy against glial tumors may greatly enhance the level of radiation-induced cell killing during a combined radio- and chemotherapeutic regimen.
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Affiliation(s)
- Chaim B Colen
- Department of Neurological Surgery, Wayne State University School of Medicine, Detroit, Michigan 48201, USA
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23
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Huang Q, Pu P, Xia Z, You Y. Exogenous wt-p53 enhances the antitumor effect of HSV-TK/GCV on C6 glioma cells. J Neurooncol 2006; 82:239-48. [PMID: 17102907 DOI: 10.1007/s11060-006-9279-x] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2006] [Accepted: 10/02/2006] [Indexed: 10/23/2022]
Abstract
OBJECTIVE To study on the antitumor effect of combining wt-p53 gene with suicide gene therapy (HSV-tk+GCV) for malignant gliomas. METHODS AdCMV-p53 was transfected into C6 glioma cells at MOI of (Multiplicity of infection) 0(G100), 10(TPG1), 100(TPG2), then AdCMV-tk was transducted to C6 glioma cells of G100, TPG1 and TPG2, respectively, at MOI of 100. The C6 glioma cells tranfected with both AdCMV-p53 and AdCMV-tk were exposed to various concentration of GCV. The cell survival rate was measured by MTT assay in vitro. Rat glioma model was established by injecting 5 x 10(5) C6 glioma cells into right caudate nucleus of SD rats. AdCMV-p53 and AdCMV-tk were injected into glioma on day 5 and 6, respectively. On day 7, ganciclovir (GCV) was administrated intraperitoneally at 15 mg/kg/day for 14 days. The survival time of all rats was observed. The growth of intracerebral tumors was monitored dynamically by enhanced MRI. Cell apoptosis was evaluated by TUNEL method. Expression of HSV-tk gene was identified by in situ hybridization and expression of exogenous p53 gene was detected with Western blotting. RESULTS In vitro, wt-p53 significantly enhanced antitumor effect of HSV-tk/GCV. The concentration of GCV for ID50 of TPG2 cells (0.001 microg/ml GCV) was 10 times lower than that for the cells of tk-GCV group (MOI = 100), while the concentration of GCV for ID100 of TPG2 (0.01 microg/ml GCV) and TPG1(0.1 microg/ml GCV) was 100 and 10 times lower than that for the cells of tk-GCV group (MOI = 100), respectively. Apoptosis of C6 glioma cells also could be induced by transfection with wt-p53 gene slightly. For in vivo study, the survival time of tumor-bearing rats treated with HSV-TK/GCV or wt-p53 combined with HSV-TK/GCV was significantly prolonged and the intracerebral tumors were regressed and disappeared earlier in the combined gene therapy group than those in the HSV-TK/GCV therapy group as shown in enhanced MRI. However, only half dose of GCV for the rats treated with both wt-p53 and HSV-TK/GCV was needed to obtain the same efficacy as those rats treated with HSV-TK/GCV alone. These results indicate that the transfection of wt-p53 potentiates the effect of HSV-TK/GCV therapy. CONCLUSIONS The combination of HSV-tk/GCV system with wt-p53 gene transduction is optimal for clinical therapeutic trials of suicide gene therapy for malignant gliomas.
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Affiliation(s)
- Qiang Huang
- Department of Neurosurgery, Tianjin Medical University General Hospital, 154 Anshan Road, Tianjin, 300052, People's Republic of China.
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24
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Jiang C, Ataai M, Ozuer A, Krisky D, Wechuck J, Pornsuwan S, Pourarian F, Glorioso JC. Inactivation of herpes simplex type 1 gene vector on immobilized metal affinity chromatography: oxidative damage by hydroxyl free radicals and its prevention. Biotechnol Bioeng 2006; 95:48-57. [PMID: 16673413 DOI: 10.1002/bit.20943] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
Metal catalyzed oxidation (MCO), which typically involves oxygen free radical generation, is an important pathway that leads to the deterioration of many biological molecules in solution. The occurrence of MCO in immobilized metal affinity chromatography (IMAC) systems and its potential for inactivating biological products has not been well recognized. In this study, we report the inactivation of herpes simplex virus type 1 (HSV-1) gene therapy vector on immobilized cobalt affinity chromatography. We observed that purification of KgBHAT, an HSV-1 mutant bearing cobalt affinity tags (HAT) on the surface, on an IDA-Co2+ column using crude supernatant as starting material resulted in signification loss in virus infectivity (<5% recovery). Electron spin resonance (ESR) revealed that the virus inactivation was caused by hydroxyl free radicals generated from the interactions between cellular impurities and the metal ions on the column. Inclusion of 20 mM ascorbate, a free radical scavenger, in the chromatography mobile phase effectively scavenged the hydroxyl radicals and dramatically augmented the infectivity recovery to 70%. This finding is the first demonstration of oxygen free radical-mediated biological inactivation in an actual IMAC purification and the way on how to effectively prevent it.
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Affiliation(s)
- Canping Jiang
- Department of Chemical Engineering, University of Pittsburgh, Pittsburgh, Pennsylvania 15261, USA
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25
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King GD, Curtin JF, Candolfi M, Kroeger K, Lowenstein PR, Castro MG. Gene therapy and targeted toxins for glioma. Curr Gene Ther 2006; 5:535-57. [PMID: 16457645 PMCID: PMC1629033 DOI: 10.2174/156652305774964631] [Citation(s) in RCA: 41] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
The most common primary brain tumor in adults is glioblastoma. These tumors are highly invasive and aggressive with a mean survival time of nine to twelve months from diagnosis to death. Current treatment modalities are unable to significantly prolong survival in patients diagnosed with glioblastoma. As such, glioma is an attractive target for developing novel therapeutic approaches utilizing gene therapy. This review will examine the available preclinical models for glioma including xenographs, syngeneic and genetic models. Several promising therapeutic targets are currently being pursued in pre-clinical investigations. These targets will be reviewed by mechanism of action, i.e., conditional cytotoxic, targeted toxins, oncolytic viruses, tumor suppressors/oncogenes, and immune stimulatory approaches. Preclinical gene therapy paradigms aim to determine which strategies will provide rapid tumor regression and long-term protection from recurrence. While a wide range of potential targets are being investigated preclinically, only the most efficacious are further transitioned into clinical trial paradigms. Clinical trials reported to date are summarized including results from conditionally cytotoxic, targeted toxins, oncolytic viruses and oncogene targeting approaches. Clinical trial results have not been as robust as preclinical models predicted, this could be due to the limitations of the GBM models employed. Once this is addressed, and we develop effective gene therapies in models that better replicate the clinical scenario, gene therapy will provide a powerful approach to treat and manage brain tumors.
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Affiliation(s)
- Gwendalyn D King
- Gene Therapeutics Research Institute, Cedars-Sinai Medical Center, and Department of Molecular and Medical Pharmacology, David Geffen School of Medicine, UCLA, 8700 Beverly Boulevard, Los Angeles, CA 90048, USA
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Abstract
Cancer remains a serious threat to human health, causing over 500 000 deaths each year in US alone, exceeded only by heart diseases. Many new technologies are being developed to fight cancer, among which are gene therapies and oncolytic virotherapies. Herpes simplex virus type 1 (HSV-1) is a neurotropic DNA virus with many favorable properties both as a delivery vector for cancer therapeutic genes and as a backbone for oncolytic viruses. Herpes simplex virus type 1 is highly infectious, so HSV-1 vectors are efficient vehicles for the delivery of exogenous genetic materials to cells. The inherent cytotoxicity of this virus, if harnessed and made to be selective by genetic manipulations, makes this virus a good candidate for developing viral oncolytic approach. Furthermore, its large genome size, ability to infect cells with a high degree of efficiency, and the presence of an inherent replication controlling mechanism, the thymidine kinase gene, add to its potential capabilities. This review briefly summarizes the biology of HSV-1, examines various strategies that have been used to genetically modify the virus, and discusses preclinical as well as clinical results of the HSV-1-derived vectors in cancer treatment.
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Affiliation(s)
- Y Shen
- Mary Crowley Medical Research Center, Dallas, TX 75201, USA
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27
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Berto E, Bozac A, Marconi P. Development and application of replication-incompetent HSV-1-based vectors. Gene Ther 2006; 12 Suppl 1:S98-102. [PMID: 16231061 DOI: 10.1038/sj.gt.3302623] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Abstract
The replication-incompetent HSV-1-based vectors are herpesviruses in which genes that are 'essential' for viral replication have been either mutated or deleted. These deletions have substantially reduced their cytotoxicity by preventing early and late viral gene expression and, together with other deletions involving 'nonessential' genes, have also created space to introduce distinct and independently regulated expression cassettes for different transgenes. Therapeutic effects in gene therapy applications requiring simultaneous and synergic expression of multiple gene products are easily achievable with these vectors. A number of different HSV-1-based nonreplicative vectors for specific gene therapy applications have been developed so far. They have been tested in different gene therapy animal models of neuropathies (Parkinson's disease, chronic pain, spinal cord injury pain) and lysosomal storage disorders. Many replication-incompetent HSV-1-based vectors have also been used either as potential anti-herpes vaccines, as well as vaccine vectors for other pathogens in murine and simian models. Anticancer gene therapy approaches have also been successfully set up; gene therapy to other targets by using these vectors is feasible.
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Affiliation(s)
- E Berto
- Department of Experimental and Diagnostic Medicine, Section of Microbiology, University of Ferrara, Ferrara, Italy
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28
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Hadjipanayis CG, DeLuca NA. Inhibition of DNA repair by a herpes simplex virus vector enhances the radiosensitivity of human glioblastoma cells. Cancer Res 2005; 65:5310-6. [PMID: 15958578 DOI: 10.1158/0008-5472.can-04-3793] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
Expression of the herpes simplex virus (HSV) protein, ICP0, from the viral genome, rendered two radioresistant human glioblastoma multiforme cell lines more sensitive to the effects of ionizing radiation. Using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and clonogenic survival assays, U87-MG and T98 cell survival was more greatly decreased as a function of ionizing radiation dose when ICP0 was preexpressed in cells compared with when ICP0 was not expressed. Consistent with previous results, we found that the catalytic subunit of DNA-dependent protein kinase was degraded as a function of ICP0 in both cell types. This most likely resulted in the inhibition of DNA repair as inferred by the persistence of gammaH2AX foci or DNA double-strand breaks. Enhanced apoptosis was also found to occur following irradiation of U87-MG cells preinfected with the ICP0-producing HSV-1 mutant, d106. Our results suggest that expression of ICP0 in human glioblastoma multiforme cells inhibits the repair of DNA double-strand breaks after ionizing radiation treatment, decreasing the survival of these cells in part by induction of apoptosis.
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Affiliation(s)
- Costas G Hadjipanayis
- Department of Neurological Surgery, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261, USA
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29
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Moriuchi S, Glorioso JC, Maruno M, Izumoto S, Wolfe D, Huang S, Cohen JB, Yoshimine T. Combination gene therapy for glioblastoma involving herpes simplex virus vector-mediated codelivery of mutant IκBα and HSV thymidine kinase. Cancer Gene Ther 2005; 12:487-96. [PMID: 15692608 DOI: 10.1038/sj.cgt.7700816] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/21/2023]
Abstract
To improve the effectiveness of herpes simplex virus (HSV) thymidine kinase/ganciclovir (HSV-tk/GCV) suicide gene therapy, the replication-defective HSV vector TOIkappaB expressing both HSV-TK and a mutant form of the NF-kappaB inhibitor IkappaBalpha (IkappaBalphaM) was developed. TOIkappaB was constructed by recombining the IkappaBalphaM gene into the U(L)41 locus of a replication-defective lacZ expression vector, TOZ.1. Expression of IkappaBalphaM was confirmed by Western blotting, and the ability of the mutant protein to inhibit NF-kappaB nuclear translocation was examined by electrophoretic mobility shift assay. In human glioblastoma U-87MG cells, the p50/p50 dimer of NF-kappaB was already translocated to the nucleus without receptor-dependent signaling by TNF-alpha. Following infection with TOIkappaB, nuclear translocation of NF-kappaB in U-87MG cells was significantly inhibited and caspase-3 activity increased compared with TOZ.1-infected cells. The cytotoxicity of TOIkappaB for U-87MG cells was investigated by colorimetric MTT assay. At an MOI of 3, TOIkappaB infection killed 85% of the cells compared to 20% killed by TOZ.1 infection. In the presence of GCV, these numbers increased to 95-100% for TOIkappaB and 80-85% for TOZ.1. TOIkappaB neurotoxicity measured on cultured murine neurons was relatively low and similar to that of TOZ.1. The survival of nude mice implanted into the brain with U-87MG tumor cells was markedly prolonged by intratumoral TOIkappaB injection and GCV administration. Survival of TOIkappaB+GCV group was significantly longer (P<.02, Wilcoxon test) than for the control groups (TOZ.1 or TOIkappaB only, PBS or PBS+GCV). These results suggest that IkappaBalphaM expression may be a safe enhancement of replication-defective HSV-based suicide gene therapy in vitro and in vivo.
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Affiliation(s)
- Shusuke Moriuchi
- Department of Neurosurgery, Osaka Graduate University Medical School, 2-2 Yamada-oka, Suita city, Osaka 565, Japan.
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Wolfe D, Niranjan A, Trichel A, Wiley C, Ozuer A, Kanal E, Kondziolka D, Krisky D, Goss J, Deluca N, Murphey-Corb M, Glorioso JC. Safety and biodistribution studies of an HSV multigene vector following intracranial delivery to non-human primates. Gene Ther 2005; 11:1675-84. [PMID: 15306839 PMCID: PMC1449743 DOI: 10.1038/sj.gt.3302336] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Malignant glioma is a fatal human cancer in which surgery, chemo- and radiation therapies are ineffective. Therapeutic gene transfer used in combination with current treatment methods may augment their effectiveness with improved clinical outcome. We have shown that NUREL-C2, a replication-defective multigene HSV-based vector, is effective in treating animal models of glioma. Here, we report safety and biodistribution studies of NUREL-C2 using rhesus macaques as a model host. Increasing total doses (1 x 10(7) to 1 x 10(9) plaque forming units (PFU)) of NUREL-C2 were delivered into the cortex with concomitant delivery of ganciclovir (GCV). The animals were evaluated for changes in behavior, alterations in blood cell counts and chemistry. The results showed that animal behavior was generally unchanged, although the chronic intermediate dose animal became slightly ataxic on day 12 postinjection, a condition resolved by treatment with aspirin. The blood chemistries were unremarkable for all doses. At 4 days following vector injections, magnetic resonance imaging showed inflammatory changes at sites of vector injections concomitant with HSV-TK and TNFalpha expression. The inflammatory response was reduced at 14 days, resolving by 1 month postinjection, a time point when transgene expression also became undetectable. Immunohistochemical staining following animal killing showed the presence of a diffuse low-grade gliosis with infiltrating macrophages localized to the injection site, which also resolved by 1 month postinoculation. Viral antigens were not detected and injected animals did not develop HSV-neutralizing antibodies. Biodistribution studies revealed that vector genomes remained at the site of injection and were not detected in other tissues including contralateral brain. We concluded that intracranial delivery of 1 x 10(9) PFU NUREL-C2, the highest anticipated patient dose, was well tolerated and should be suitable for safety testing in humans.
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Affiliation(s)
- D Wolfe
- Department of Molecular Genetics and Biochemistry, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
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Glorioso JC, Fink DJ. Herpes vector-mediated gene transfer in treatment of diseases of the nervous system. Annu Rev Microbiol 2004; 58:253-71. [PMID: 15487938 DOI: 10.1146/annurev.micro.58.030603.123709] [Citation(s) in RCA: 90] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Vectors constructed from recombinant herpes simplex virus (HSV) have special utility for gene transfer to the nervous system. Nonreplicating vectors created by deletion of essential immediate early genes can be propagated to high titers on complementing cell lines that provide the missing gene product(s) in trans. Direct inoculation of these vectors into neural parenchyma is effective in rodent models of brain tumor, Parkinson disease, spinal cord injury, and spinal root trauma. Subcutaneous inoculation of the HSV vectors can be used to transduce neurons of the dorsal root ganglion to provide a therapeutic effect in models of polyneuropathy and chronic regional pain. In human trials, direct injection of replication-competent HSV into brain tumors has proven safe. Human trials of nonreplicating HSV gene transfer by direct inoculation for treatment of glioblastoma and HSV gene transfer by subcutaneous inoculation for the treatment of chronic intractable pain should commence soon.
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Affiliation(s)
- Joseph C Glorioso
- Department of Molecular Genetics and Biochemistry, University of Pittsburgh, Pittsburgh, Pennsylvania 15261, USA.
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32
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Jiang C, Wechuck JB, Goins WF, Krisky DM, Wolfe D, Ataai MM, Glorioso JC. Immobilized cobalt affinity chromatography provides a novel, efficient method for herpes simplex virus type 1 gene vector purification. J Virol 2004; 78:8994-9006. [PMID: 15308696 PMCID: PMC506967 DOI: 10.1128/jvi.78.17.8994-9006.2004] [Citation(s) in RCA: 30] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Herpes simplex virus type 1 (HSV-1) is a promising vector for gene therapy applications, particularly at peripheral nerves, the natural site of virus latency. Many gene vectors require large particle numbers for even early-phase clinical trials, emphasizing the need for high-yield, scalable manufacturing processes that result in virus preparations that are nearly free of cellular DNA and protein contaminants. HSV-1 is an enveloped virus that requires the development of gentle purification methods. Ideally, such methods should avoid centrifugation and may employ selective purification processes that rely on the recognition of a unique envelope surface chemistry. Here we describe a novel method that fulfills these criteria. An immobilized metal affinity chromatography (IMAC) method was developed for the selective purification of vectors engineered to display a high-affinity binding peptide. Feasibility studies involving various transition metal ions (Cu2+, Zn2+, Ni2+, and Co2+) showed that cobalt had the most desirable features, which include a low level of interaction with either the normal virus envelope or contaminating DNA and proteins. The introduction of a cobalt-specific recognition element into the virus envelope may provide a suitable target for cobalt-dependent purification. To test this possibility, we engineered a peptide with affinity for immobilized cobalt in frame in the heparan sulfate binding domain of HSV-1 glycoprotein B, which is known to be exposed on the surface of the virion particle and recombined into the viral genome. By optimizing the IMAC loading conditions and reducing cobalt ion leakage, we recovered 78% of the tagged HSV-1 recombinant virus, with a >96% reduction in contaminating proteins and DNA.
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Affiliation(s)
- Canping Jiang
- Department of Chemical Engineering, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
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Niranjan A, Gobbel GT, Kondziolka D, Flickinger JC, Lunsford LD. Experimental Radiobiological Investigations into Radiosurgery: Present Understanding and Future Directions. Neurosurgery 2004; 55:495-504; discussion 504-5. [PMID: 15335417 DOI: 10.1227/01.neu.0000134283.69965.a7] [Citation(s) in RCA: 34] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2003] [Accepted: 02/13/2004] [Indexed: 11/19/2022] Open
Abstract
Abstract
LARS LEKSELL BEGAN radiobiological investigations to study the effect of high-dose focused radiation on the central nervous system more than 5 decades ago. Although the effects of radiosurgery on the brain tumor microenvironment are still under investigation, radiosurgery has become a preferred management modality for many intracranial tumors and vascular malformations. The effects and the pathogenesis of biological effects after radiosurgery may be unique. The need for basic research concerning the radiobiological effects of high-dose, single-fraction, ionizing radiation on nervous system tissue is crucial. Information from those studies would be useful in devising strategies to avoid, prevent, or ameliorate damage to normal tissue without compromising treatment efficacy. The development of future applications of radiosurgery will depend on an increase in our understanding of the radiobiology of radiosurgery, which in turn will affect the efficacy of treatment. This article analyzes the current state of radiosurgery research with regard to the nature of central nervous system effects, the techniques developed to increase therapeutic efficacy, investigations into the use of radiosurgery for functional disorders, radiosurgery as a tool for investigations into basic central nervous system biology, and the additional areas that require further investigation.
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Affiliation(s)
- Ajay Niranjan
- Department of Neurological Surgery, University of Pittsburgh, Center for Image-Guided Neurosurgery, University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania, USA
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34
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Lundstrom K, Boulikas T. Viral and non-viral vectors in gene therapy: technology development and clinical trials. Technol Cancer Res Treat 2004; 2:471-86. [PMID: 14529313 DOI: 10.1177/153303460300200513] [Citation(s) in RCA: 84] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022] Open
Abstract
Gene therapy as part of modern molecular medicine holds great promise for the treatment of both acute and chronic diseases and has the potential to bring a revolutionary era to cancer treatment. Gene therapy has been named the medicine of the future. For the past 10 years various viral and non-viral vectors have been engineered for improved gene and drug delivery. Although various diseases have been targeted, cancer therapy has been addressed to a large extent because of the straight forward approach. Delivery of toxic or immunostimulatory genes by viral and non-viral vectors has been investigated and encouraging results have been obtained in animal models. A large number of clinical trials have been conducted with some highly promising outcome. We propose that combinations of viruses with liposomes or polymers will solve the problem of systemic viral delivery and tumor targeting, bringing a revolution in molecular medicine and in applications of gene therapy in humans.
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Affiliation(s)
- Kenneth Lundstrom
- Regulon Inc./BioXtal, Chemin des Croisettes 22, CH-1066, Epalinges, Swizerland.
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Abstract
Developing and conducting gene therapy clinical trials poses unique challenges which must be addressed to satisfy regulatory requirements and, most importantly, to protect human subjects. Experimental products used for gene transfer studies, such as viral vectors, are often complex and cannot be sterilized or completely characterized to the extent of a typical pharmaceutical. Thus, quality and characterization must be built into the production process. Extensive preclinical studies must be performed to determine the feasibility of the approach, the safety of the product, and the appropriate dose range to evaluate in humans. Once a clinical trial is initiated, subjects must be followed carefully for short- and long-term toxicity especially since preclinical studies may not adequately predict the toxicity profile of these novel, complicated products. Results of early phase studies in gene therapy have often sent the investigators back to the laboratory to improve the delivery vector or identify a more potent or less toxic gene. This circular developmental process is expected for the early stages of a new technology such as gene therapy. Although these hurdles appear extensive, they can be overcome, as evidenced by the initiation of more than 500 clinical gene therapy trials in the United States to date, and are imperative for the maintenance of high-quality studies and public trust. This article describes the step-by-step process for developing a gene therapy trial incorporating specific examples relevant to neuro-oncology.
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Affiliation(s)
- Laura K Aguilar
- Harvard Gene Therapy Initiative, Harvard Medical School, Boston, MA, USA.
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36
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Levy EI, Kim SH, Bendok BR, Boulos AS, Xavier AR, Yahia AM, Qureshi AI, Guterman LR, Hopkins LN. Interventional Neuroradiologic Therapy. Stroke 2004. [DOI: 10.1016/b0-44-306600-0/50087-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/18/2023]
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Zhang JH, Wan MX, Yuan JY, Pan BR. Do there exist synergistic antitumor effects by coexpression of herpes simplex virus thymidine kinase with cytokine genes on human gastric cancer cell line SGC7901? World J Gastroenterol 2004; 10:147-51. [PMID: 14695787 PMCID: PMC4717068 DOI: 10.3748/wjg.v10.i1.147] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
AIM: To evaluate the synergistic antitumor effects of herpes simplex virus thymidine kinase (HSV-TK) together with tumor necrosis factor alpha (TNF-α) or interleukin-2 (IL-2) gene expression on gastric cancer cell line SGC7901.
METHODS: Recombinant vectors pL(TT)SN and pL(TI)SN, which express TK-IRES-TNF-α and TK-IRES-IL-2 genes separately, as well as the control plasmids pL(TK)SN and pLXSN were employed to transfect PA317 cells respectively to generate the viruses that can stably express the objective genes through G418 selection. The gastric cancer cells were then transfected by the retroviral serum from the package cells and maintained in culture to determine the cell growth and apoptosis. The cytotoxic effects of HSV-TK together with TNF-α or IL-2 gene expression on the transfected cancer cells were evaluated by the cell viability and bystander effects in the presence of GCV supplemented in the cultural medium.
RESULTS: Expression of recombinant proteins including TNF-α and IL-2 by stable transfectants was confirmed by Western blotting. The percentage of cell apoptosis in the SGC/0, SGC/TK-TNF-α, SGC/TK-IL-2 and SGC/TK clone was 2.3%, 12.3%, 11.1% and 10.9% respectively at 24 h post-transfection. Cell growth status among all the experimental groups as judged by cell absorbance (A) at 570nm did not exhibit any significant difference (P > 0.05); although it was noted to be slightly lower in the SGC/TT group. Cell survival rate in SGC/TI, SGC/TT and SGC/TK group was significantly decreased in a dose-dependent manner of GCV compared with that of the SGC/0 group (P < 0.05-0.01). Among all studied cells, the SGC/TT was shown most sensitive to GCV with a half lethal dose of 0.5 mg·L-1. In contrast, the survival rate of SGC/0 cells was not affected by the presence of GCV with the doses less than 10 mg·L-1. The half lethal dose of GCV for SGC/0 cells was more than 100 mg·L-1. Marked bystander effect induced by SGC/TI, SGC/TT and SGC/TK cells was confirmed by the fact that 20% of these stable transfectants could kill 50% of the co-cultured cells, in which the most prominent bystander effect was found in the circumstance of SGC/TT presence. However, no significant difference of these variables was found among SGC/TI, SGC/TT and SGC/TK cells (P > 0.05).
CONCLUSION: The synergistic antitumor effects produced by the co-expression of HSV-TK with TNF-α or IL-2 genes were not present in the transfected SGC7901 cells. The mechanism underlying these phenomena was not known.
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Affiliation(s)
- Jian-Hua Zhang
- Department of Biomedical Engineering, School of Life Science and Technology, Xi'an Jiaotong University, 28 West Xianning Road, Xi'an 710049, Shaanxi Province, China.
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38
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Niranjan A, Wolfe D, Tamura M, Soares MK, Krisky DM, Lunsford LD, Li S, Fellows-Mayle W, DeLuca NA, Cohen JB, Glorioso JC. Treatment of rat gliosarcoma brain tumors by HSV-based multigene therapy combined with radiosurgery. Mol Ther 2003; 8:530-42. [PMID: 14529825 DOI: 10.1016/s1525-0016(03)00232-6] [Citation(s) in RCA: 41] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022] Open
Abstract
Our laboratory has employed replication-defective herpes simplex virus type 1 gene transfer vectors for treatment of animal models of human malignant glioblastoma. The base vectors were defective for the immediate early (IE) genes ICP4, ICP27, and ICP22 but expressed the IE gene ICP0, which can arrest tumor cell division, and an IE thymidine kinase (alpha-tk) gene construct that mediates suicide gene therapy (SGT) in the presence of ganciclovir (GCV). Previously, we reported that SGT using ICP0/alpha-tk vectors in nude mouse models of glioblastoma was improved by coexpression of the gap-junction-forming protein connexin43 (Cx43) or human tumor necrosis factor alpha (TNF alpha). We also showed that further gains in therapeutic outcome could be achieved by combining TNF alpha-enhanced SGT with gamma-knife radiosurgery (GKR). To expand these observations, we have first repeated these studies in immunocompetent rats with brain tumors derived from implanted 9L gliosarcoma cells and second compared the most efficient vector from this study with a new recombinant vector, NUREL-C2, which expressed both TNF alpha and Cx43 along with ICP0 and alpha-tk. Results from the first part indicated that our ICP0/alpha-tk/TNF alpha vector in combination with GKR provides an effective therapy although this treatment was not statistically better than GKR combined with the ICP0/alpha-tk/Cx43 vector. Our observations in the second part suggested that NUREL-C2 may be more effective than the ICP0/alpha-tk/TNF alpha vector in combination treatments with GCV (P = 0.08) or GCV plus GKR (P = 0.10). GKR significantly enhanced the efficacy of NUREL-C2/GCV treatment (P = 0.02) as well as other virus/GCV treatments (P < or = 0.05). Conversely, the efficacy of GKR was significantly improved by both the ICP0/alpha-tk/TNF alpha vector and NUREL-C2 in combination with GCV (P = 0.02 and P < 0.01, respectively). Together these results indicate that NUREL-C2 may be an attractive candidate for Phase I gene-therapy safety studies in patients with recurrent malignant glioblastoma.
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Affiliation(s)
- Ajay Niranjan
- Department of Neurological Surgery, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261, USA
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Ren H, Boulikas T, Lundstrom K, Söling A, Warnke PC, Rainov NG. Immunogene therapy of recurrent glioblastoma multiforme with a liposomally encapsulated replication-incompetent Semliki forest virus vector carrying the human interleukin-12 gene--a phase I/II clinical protocol. J Neurooncol 2003; 64:147-54. [PMID: 12952295 DOI: 10.1007/bf02700029] [Citation(s) in RCA: 83] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
Glioblastoma multiforme (GBM) is an incurable brain tumor resistant to standard treatment modalities such as surgery, radiation, and chemotherapy. Since recurrent GBM tends to develop predominantly within the infiltrative rim surrounding the primary tumor focus, novel therapy strategies need in addition to focal tumor destruction to target this somewhat diffuse area. This is a phase I/II clinical study in adult patients with recurrent GBM which is aimed at evaluating biological safety, maximum tolerated dose, and antitumor efficacy of a genetically modified replication-disabled Semliki forest virus vector (SFV) carrying the human interleukin 12 (IL-12) gene and encapsulated in cationic liposomes (LSFV-IL12). The vector will be administered in doses of 1 x 10(7)-1 x 10(9) infectious particles by continuous intratumoral infusion, thus exploiting the advantages of convection-enhanced drug delivery in the brain. The present protocol is also designed to investigate systemic and local immune response and to identify factors predicting tumor response to LSFV-IL12 therapy, such as volume of extracellular space of the tumor, volume of contrast enhancing lesion, and immune status of the patients. SFV, an insect alphavirus, infects mitotic and non-mitotic cells and triggers apoptosis in tumor cells within 48-72 h. Preclinical work with the LSFV-IL12 vector in breast and prostate cancer animal models demonstrated its biosafety and some antitumor efficacy. An ongoing phase I clinical study in patients with melanoma and renal cell carcinoma seems also to confirm the biosafety of intravenously administered vectors. This protocol will be the first study of SFV-IL12 therapy of human recurrent GBM.
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Affiliation(s)
- H Ren
- Department of Neurological Science, University of Liverpool, Liverpool, UK
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40
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Curi MA, Skelly CL, Meyerson SL, Baldwin ZK, Balasubramanian V, Advani SJ, Glagov S, Roizman B, Weichselbaum RR, Schwartz LB. Sustained inhibition of experimental neointimal hyperplasia with a genetically modified herpes simplex virus. J Vasc Surg 2003; 37:1294-300. [PMID: 12764278 DOI: 10.1016/s0741-5214(02)75333-0] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/27/2022]
Abstract
OBJECTIVE Reported herein is a potential strategy for sustained smooth muscle cell (SMC) inhibition with a virulence-attenuated herpes simplex virus (HSV). Experiments were conducted in vitro to demonstrate selective SMC cytotoxicity and in vivo to demonstrate reduced neointimal hyperplasia (NIH) in a clinically relevant animal model. METHODS In vitro: Cultured human umbilical artery smooth muscle cells (UASMC) and venous endothelial cells (HUVEC) were exposed to varying multiplicities of infection (MOI) of a gamma(1)34.5-deleted HSV-1 virus (R849). Cell survival was assessed at 48 and 72 hours with a colorimetric MTT viability assay. In vivo: New Zealand White rabbit external jugular veins (n = 21) were exposed to R849 (2.5 x 10(6) pfu/mL) or culture medium at 110 to 120 mm Hg for 10 minutes, then fashioned as vein patches on carotid arteries. Carotid arteries were ligated distally to decrease blood flow and stimulate a hyperplastic response (ultra-low shear stress model). After 2, 4, 12, and 24 weeks, patched segments were perfusion-fixed with glutaraldehyde and morphometrically examined for NIH formation. RESULTS In vitro: At 48 hours, R849 exhibited preferential cytotoxicity to UASMC compared with HUVEC, with 11% +/- 10% of UASMCs and 49% +/- 8% of HUVECs surviving after infection with MOI = 25 (P <.05). Higher MOI resulted in poor survival of both cell lines. In vivo: Blood flow was similarly reduced in all animals both at surgery (0.9 +/- 0.1 mL/min vs 1.6 +/- 0.3 mL/min) and at harvest (2.7 +/- 0.4 mL/min vs 2.5 +/- 0.5 mL/min). R849-infected patches exhibited markedly less NIH than control patches did at 2 weeks (162 +/- 14 microm vs 49 +/- 6 microm; P <.05), 4 weeks (190 +/- 27 microm vs 67 +/- 8 microm; P <.05), and 12 weeks (233 +/- 18 microm vs 113 +/- 2 microm; P <.05). CONCLUSION The virulence-attenuated HSV strain R849 demonstrates selective cytotoxicity for SMC and is capable of sustained inhibition of NIH in an experimental model of vein graft failure.
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Affiliation(s)
- Michael A Curi
- Section of Vascular Surgery, Department of Surgery, University of Chicago MC 5028, 5841 S Maryland Ave, Chicago, IL 60637, USA
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41
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Gerosa M, Nicolato A, Foroni R. The role of gamma knife radiosurgery in the treatment of primary and metastatic brain tumors. Curr Opin Oncol 2003; 15:188-96. [PMID: 12778010 DOI: 10.1097/00001622-200305000-00002] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
With the widespread diffusion of stereotactic radiosurgical procedures, GKR treatments have gained considerable momentum as a major therapeutic option for patients harboring primary or metastatic brain tumors. Present results in high grade gliomas indicate a potential palliative role of this technique. The overall low radiosensitivity of these oncotypes and their infiltrative nature-with the resulting problems in properly defining the tumor target-are still a major obstacle to further development of the approach. In this regard, useful contributions are expected from advances in molecular neurobiology and functional neuroimaging as shown by preliminary investigations with MR spectroscopy. Surgery maintains a dominant role in the therapeutic armamentarium for low grade gliomas. However, in unfavorable cases (unresectable tumors, recurrences), GKR seems to be an effective alternative to conventional radiochemotherapy. In grade 2 astrocytomas and specifically in grade 1 pilocytic forms, short-to-mid-term reported studies have documented encouraging 70 to 93% local tumor control rates, with minimal cerebral toxicity. Finally, during the last decade, GKR has become a primary treatment choice for patients harboring small-to-medium-size brain metastases, with reasonable life expectancy and no impending intracranial hypertension. Focal tumor responses are consistently elevated, even in the most radioresistant oncotypes (melanoma, renal carcinoma); median and actuarial survival rates are far better than with conventional radiation treatments and are comparable to those observed in accurately selected surgical-radiation series.
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Affiliation(s)
- Massimo Gerosa
- Department of Neurosurgery, University Hospital, Piazzale Stefani 1, 37126 Verona, Italy.
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42
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Roberge D, Souhami L. Stereotactic radiosurgery in the management of intracranial gliomas. Technol Cancer Res Treat 2003; 2:117-25. [PMID: 12680792 DOI: 10.1177/153303460300200207] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2022] Open
Abstract
Glial neoplasms are the most common primary intracranial malignancies. Treatment of high-grade gliomas has been frustrating, with less than 5% of patients surviving 5 years after a diagnosis of glioblastoma multiforme (GBM). Stereotactic radiosurgery (SRS) and fractionated strereotactic radiotherapy (F-SRT) provide means to either escalate the dose in primary treatment or to palliate recurrences. Because of their lower alpha/beta ratios and more focal nature, low-grade gliomas (LGG) are more attractive targets for stereotactically focused radiation. Results of available phase I-II data are reviewed for both low and high-grade gliomas. In the case of high-grade gliomas disappointing preliminary phase III data from RTOG 93-05 are discussed. Toxicity of SRS is discussed. Acute treatment toxicity of significance is unusual and generally self-limited. Occasionally an exacerbation of existing symptoms occurs. Late complications attributable to SRS are usually defined as necrosis within the treatment volume. The rate of necrosis can be hard to define in high-grade gliomas as tumor cells are often present in surgical specimens. New strategies in the application of stereotactic radiation are touched upon, these include: changes in planning and fractionation, concurrent use of chemotherapy, use of radiation modifiers and biologic agents. After reviewing the current data for high-grade gliomas, it appears that any apparent improvement in outcome seen in phase I-II trials is attributable to patient selection. The best evidence available does not support the use of SRS for primary high-grade gliomas. The somewhat limited experience in LGG also indicates a lack of benefit for patients treated with stereotactic radiosurgery or F-SRT. For a very select group of patients with small recurrent lesions, F-SRT may represent a safe, reasonable treatment.
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Affiliation(s)
- David Roberge
- Division of Radiation Oncology, McGill University, Montreal General Hospital, 1650 Cedar Ave., Montreal, QC, Canada H3G 1A4
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43
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Aghi M, Chiocca EA. Genetically engineered herpes simplex viral vectors in the treatment of brain tumors: a review. Cancer Invest 2003; 21:278-92. [PMID: 12743992 DOI: 10.1081/cnv-120016423] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/03/2022]
Affiliation(s)
- Manish Aghi
- Molecular Neuro-Oncology Laboratories, Neurosurgical Service, Massachusetts, General Hospital, Harvard Medical School, Boston, Massachusetts, USA
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44
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Castro MG, Cowen R, Williamson IK, David A, Jimenez-Dalmaroni MJ, Yuan X, Bigliari A, Williams JC, Hu J, Lowenstein PR. Current and future strategies for the treatment of malignant brain tumors. Pharmacol Ther 2003; 98:71-108. [PMID: 12667889 DOI: 10.1016/s0163-7258(03)00014-7] [Citation(s) in RCA: 116] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
Glioblastoma (GB) is the most common subtype of primary brain tumor in adults. These tumors are highly invasive, very aggressive, and often infiltrate critical neurological areas within the brain. The mean survival time after diagnosis of GB has remained unchanged during the last few decades, in spite of advances in surgical techniques, radiotherapy, and also chemotherapy; patients' survival ranges from 9 to 12 months after initial diagnosis. In the same time frame, with our increasing understanding and knowledge of the physiopathology of several cancers, meaningful advances have been made in the treatment and control of several cancers, such as breast, prostate, and hematopoietic malignancies. Although a number of the genetic lesions present in GB have been elucidated and our understanding of the progressions of this cancer has increased dramatically over the last few years, it has not yet been possible to harness this information towards developing effective cures. In this review, we will focus on the classical ways in which GB is currently being treated, and will introduce a novel therapeutic modality, i.e., gene therapy, which we believe will be used in combination with classical treatment strategies to prolong the life-span of patients and to ultimately be able to control and/or cure these brain tumors. We will discuss the use of several vector systems that are needed to introduce the therapeutic genes within either the tumor mass, if these are not resectable, or the tumor bed, after successful tumor resection. We also discuss different therapeutic modalities that could be exploited using gene therapy, i.e., conditional cytotoxic approach, direct cytotoxicity, immunotherapy, inhibition of angiogenesis, and the use of pro-apoptotic genes. The advantages and disadvantages of each of the current vector systems available to transfer genes into the CNS are also discussed. With the advances in molecular techniques, both towards the elucidation of the physiopathology of GB and the development of novel, more efficient and less toxic vectors to deliver putative therapeutic genes into the CNS, it should be possible to develop new rationale and effective therapeutic approaches to treat this devastating cancer.
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Affiliation(s)
- M G Castro
- Gene Therapeutics Research Institute, Cedars-Sinai Medical Center, Research Pavilion, 8700 Beverly Boulevard, Suite 5090, Los Angeles, CA 90048, USA.
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45
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Abstract
Gene therapy includes the application of various viral vectors, which represent most types and families of viruses, suitable for infection of mammalian host cells. Both hereditary diseases and acquired illnesses, such as cancer, can be targeted. Because of the various properties of each viral vector, the definition of their application range depends on factors such as packaging capacity, host range, cell- or tissue-specific targeting, replication competency, genome integration and duration of transgene expression. Recent engineering of modified viral vectors has contributed to improved gene delivery efficacy.
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Affiliation(s)
- Kenneth Lundstrom
- Regulon Inc./BioXtal., Chemin des Croisettes 22, CH-1066 Epalinges, Switzerland.
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46
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Abstract
Malignant glioma formation is associated with characteristic genetic alterations, although epigenetic mechanisms may contribute in tumorigenesis. Until recently, our knowledge has mainly been based on chromosomal and molecular studies performed in the last two decades. This has increased tremendously with the advent of new technologies, in particular expression arrays for simultaneous analysis of thousands of genes. Consequently, gene therapy of gliomas may aim at molecular interference with 'gain of function' genes (oncogenes) or replacement of 'loss of function' genes (tumor suppressor genes). Such approaches require transgene expression in whole tumor cell populations (if not other mechanisms come into play) which cannot be achieved with current vector systems. Hence other strategies have been pursued which may be independent of genes actually involved in tumorigenesis. Microbial genes (e.g. herpes simplex virus thymidine kinase) may be transferred into the tumors allowing for prodrug activation (e.g. ganciclovir). Furthermore, cytokines or other immunomodulatory genes may be used for vaccination purposes which frequently involves ex vivo transfection of autologous tumor cells with such genes. These approaches proved promising in preclinical studies performed in cell culture and different inbred rodent models. A considerable number of clinical trials have been initiated based on these approaches. Although most therapeutic strategies proved safe, clinical responses fell short of expectations raised by preclinical results. This, to a large extent, has to be attributed to a lag in the development of efficient vector systems. Although much effort has been put into this area of research, neuro-oncologists are still in await of a vector system allowing for selective and efficient tumor cell transduction. This has led to increased interest in distinct but related strategies, e.g. oncolytic viruses or direct intra-tumoral delivery of anti-sense oligonucleotides.
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Affiliation(s)
- W Hamel
- Klinik für Neurochirurgie, Universitätsklinikum Hamburg-Eppendorf, Hamburg, Germany
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47
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Tasciotti E, Zoppè M, Giacca M. Transcellular transfer of active HSV-1 thymidine kinase mediated by an 11-amino-acid peptide from HIV-1 Tat. Cancer Gene Ther 2003; 10:64-74. [PMID: 12489030 DOI: 10.1038/sj.cgt.7700526] [Citation(s) in RCA: 37] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2002] [Indexed: 11/08/2022]
Abstract
Suicide gene therapy using herpes simplex virus type-1 (HSV-1) thymidine kinase (TK) is a widely exploited approach for gene therapy of cancer and other hyperproliferative disorders. Despite its popularity, clinical success has been so far hampered mostly by the relative inefficiency of TK gene transfer and its limited bystander effect. Here we report that fusion of TK to an 11-amino-acid peptide from the basic domain of the HIV-1 Tat protein (Tat11) imparts cell membrane translocating ability to the enzyme and significantly increases its cytotoxic efficacy. In cells expressing Tat11-TK, this protein is found extracellularly, associated with cell surface heparan sulfate proteoglycans, and is released into the cell culture medium. Based on its interaction with HSPGs, the protein is then internalized by neighboring, nonexpressing cells, which become susceptible to cell death when treated with the nucleoside analogue acyclovir. As a consequence, co-cultures of wild-type cells with cells expressing Tat11-TK show increased sensitivity to ACV through a mechanism involving apoptosis. Modification of TK by fusion with Tat11 might constitute an important step for the optimization of TK suicide gene strategy for gene therapy of cellular proliferation.
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Affiliation(s)
- Ennio Tasciotti
- Molecular Medicine Laboratory, International Center for Genetic Engineering and Biotechnology (ICGEB), Trieste, Italy
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48
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Abstract
Herpes simplex virus (HSV) is a neurotropic DNA virus with many favorable properties as a gene delivery vector. HSV is highly infectious, so HSV vectors are efficient vehicles for the delivery of exogenous genetic material to cells. Viral replication is readily disrupted by null mutations in immediate early genes that in vitro can be complemented in trans, enabling straightforward production of high-titre pure preparations of non-pathogenic vector. The genome is large (152 Kb) and many of the viral genes are dispensable for replication in vitro, allowing their replacement with large or multiple transgenes. Latent infection with wild-type virus results in episomal viral persistence in sensory neuronal nuclei for the duration of the host lifetime. Transduction with replication-defective vectors causes a latent-like infection in both neural and non-neural tissue; the vectors are non-pathogenic, unable to reactivate and persist long-term. The latency active promoter complex can be exploited in vector design to achieve long-term stable transgene expression in the nervous system. HSV vectors transduce a broad range of tissues because of the wide expression pattern of the cellular receptors recognized by the virus. Increasing understanding of the processes involved in cellular entry has allowed preliminary steps to be taken towards targeting the tropism of HSV vectors. Using replication-defective HSV vectors, highly encouraging results have emerged from recent pre-clinical studies on models of neurological disease, including glioma, peripheral neuropathy, chronic pain and neurodegeneration. Consequently, HSV vectors encoding appropriate transgenes to tackle these pathogenic processes are poised to enter clinical trials.
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Affiliation(s)
- Edward A Burton
- Department of Clinical Neurology, University of Oxford, United Kingdom
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49
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Ehtesham M, Samoto K, Kabos P, Acosta FL, Gutierrez MAR, Black KL, Yu JS. Treatment of intracranial glioma with in situ interferon-gamma and tumor necrosis factor-alpha gene transfer. Cancer Gene Ther 2002; 9:925-34. [PMID: 12386831 DOI: 10.1038/sj.cgt.7700516] [Citation(s) in RCA: 55] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/08/2002] [Indexed: 02/06/2023]
Abstract
Interferon-gamma (IFNgamma) and tumor necrosis factor-alpha (TNFalpha) are potent immunostimulatory cytokines with demonstrated tumoricidal effects in a variety of cancers. With the aim of investigating their ability to generate antitumor immune responses in malignant brain tumors, we describe the use of in situ adenoviral-mediated IFNgamma and TNFalpha gene transfer in glioma-bearing rodents. Survival was prolonged in mice treated with AdmIFNgamma or AdTNFalpha compared to AdLacZ- and saline-inoculated controls, and AdmIFNgamma- or AdTNFalpha-treated animals revealed significantly smaller tumors. These effects were accompanied by significant up-regulation of tumor MHC-I expression in AdmIFNgamma-inoculated animals, and of MHC-II in AdTNFalpha-treated tumors. Significantly enhanced intratumoral infiltration with CD4(+) and CD8(+) T cells was visible in animals treated with AdmIFNgamma, AdTNFalpha, or a combination of AdmIFNgamma and AdTNFalpha. In addition, AdTNFalpha therapy down-regulated the expression of endothelial Fas ligand, a cell membrane protein implicated as a contributor to immune privilege in cancer. These findings demonstrate the effectiveness of local IFNgamma and TNFalpha gene transfer as a treatment strategy for glioma and illustrate possible physiological pathways responsible for the therapeutic benefit observed.
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Affiliation(s)
- Moneeb Ehtesham
- Maxine Dunitz Neurosurgical Institute, Cedars-Sinai Medical Center, Los Angeles, California 90048, USA
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50
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Moriuchi S, Wolfe D, Tamura M, Yoshimine T, Miura F, Cohen JB, Glorioso JC. Double suicide gene therapy using a replication defective herpes simplex virus vector reveals reciprocal interference in a malignant glioma model. Gene Ther 2002; 9:584-91. [PMID: 11973634 DOI: 10.1038/sj.gt.3301693] [Citation(s) in RCA: 49] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2001] [Accepted: 02/15/2002] [Indexed: 01/12/2023]
Abstract
Herpes simplex virus thymidine kinase (HSV-TK) and Escherichia coli cytosine deaminase (CD) are non-mammalian enzymes capable of converting innocuous prodrugs into cytotoxic metabolites. Both enzymes have been utilized independently, as well as together in 'suicide' gene therapy protocols to eliminate tumor cells in vitro and in vivo. We have used a set of replication defective HSV vectors expressing either or both enzymes to compare the efficacies of single and double suicide gene therapies in the 9L gliosarcoma model in vitro and in vivo. In cell culture experiments at high and low multiplicities of infection, combined expression of the two genes by vector TOCD/TK along with exposure to the matching prodrugs (ganciclovir and 5-fluorocytosine) showed increased cytotoxicity compared with exposure to either prodrug alone. However, the two gene combination was inferior to single gene treatments, suggesting that HSVtk and CD are mutually counteractive in the prodrug-dependent killing of glioma cells. In animal experiments, survival was not significantly prolonged by administration of both prodrugs to TOCD/TK-treated animals, while each single gene/prodrug pair resulted in increased survival. These results indicate that single suicide gene systems employing HSVtk or CD may be preferable over combinations of the two.
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Affiliation(s)
- S Moriuchi
- Department of Molecular Genetics and Biochemistry, University of Pittsburgh Medical School, Pittsburgh, Pittsburgh, PA 15261, USA
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