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1.
Cancer Gene Ther ; 16(4): 373-81, 2009 Apr.
Article in English | MEDLINE | ID: mdl-18974777

ABSTRACT

TNFerade is a radioinducible adenoviral vector expressing tumor necrosis factor-alpha (TNF-alpha) (Ad.Egr-TNF) currently in a phase III trial for inoperable pancreatic cancer. We studied B16-F1 melanoma tumors in TNF receptor wild-type (C57BL/6) and deficient (TNFR1,2-/- and TNFR1-/-) mice. Ad.Egr-TNF+IR inhibited tumor growth compared with IR in C57BL/6 but not in receptor-deficient mice. Tumors resistant to TNF-alpha were also sensitive to Ad.Egr-TNF+IR in C57BL/6 mice. Ad.Egr-TNF+IR produced an increase in tumor-associated endothelial cell apoptosis not observed in receptor-deficient animals. Also, B16-F1 tumors in mice with germline deletions of TNFR1,2, TNFR1 or TNF-alpha, or in mice receiving anti-TNF-alpha exhibited radiosensitivity. These results show that tumor-associated endothelium is the principal target for Ad.Egr-TNF radiosensitization and implicate TNF-alpha signaling in tumor radiosensitivity.


Subject(s)
Genetic Therapy/methods , Melanoma, Experimental/therapy , Radiation-Sensitizing Agents , Tumor Necrosis Factor-alpha/metabolism , X-Ray Therapy , Animals , Apoptosis/drug effects , Cell Line, Tumor , Endothelial Cells/drug effects , Endothelial Cells/physiology , Etanercept , Humans , Immunoglobulin G/pharmacology , Immunosuppressive Agents/pharmacology , Mice , Neoplasm Transplantation , Receptors, Tumor Necrosis Factor , Receptors, Tumor Necrosis Factor, Type I/deficiency , Receptors, Tumor Necrosis Factor, Type II/deficiency , Tumor Necrosis Factor-alpha/antagonists & inhibitors
2.
Cancer Gene Ther ; 15(3): 133-9, 2008 Mar.
Article in English | MEDLINE | ID: mdl-18157146

ABSTRACT

We report the anticarcinogenic, anti-aging polyphenol resveratrol activates the radio- and chemo-inducible cancer gene therapy vector Ad.Egr.TNF, a replication-deficient adenovirus that expresses human tumor necrosis factor alpha (TNF-alpha) under control of the Egr-1 promoter. Like ionizing radiation or chemotherapeutic agents previously shown to activate Ad.Egr.TNF, resveratrol also induces Egr-1 expression from its chromosomal locus with a possible role for Egr-1 promoter CC(A+T)richGG sequences in the expression of TNF-alpha. Resveratrol induction of TNF-alpha in Ad.Egr.TNF-infected tumor xenografts demonstrated antitumor response in human and rat tumor models comparable to that of radio- or chemotherapy-induced TNF-alpha. Although sirtuins are known targets of resveratrol, in vitro inhibition of SIRT1 activity did not abrogate resveratrol induction of Egr-1 expression. This suggests that SIRT1 is not essential to mediate resveratrol induction of Egr-1. Nevertheless, control of transgene expression via resveratrol activation of Egr-1 may extend use of Ad.Egr.TNF to patients intolerant of radiation or cytotoxic therapy and offer a novel tool for development of other inducible gene therapies.


Subject(s)
Adenoviridae/genetics , Genetic Therapy/methods , Stilbenes/pharmacology , Tumor Necrosis Factor-alpha/genetics , Xenograft Model Antitumor Assays/methods , Acetylation/drug effects , Animals , Blotting, Western , Cell Line, Tumor , Early Growth Response Protein 1/genetics , Early Growth Response Protein 1/metabolism , Enzyme-Linked Immunosorbent Assay , Etoposide/pharmacology , Female , Gene Expression Regulation, Neoplastic/drug effects , Humans , Mice , Mice, Nude , Rats , Resveratrol , Sirtuins/metabolism , Tumor Necrosis Factor-alpha/metabolism , Tumor Suppressor Protein p53/metabolism
3.
Gene Ther ; 10(3): 206-12, 2003 Feb.
Article in English | MEDLINE | ID: mdl-12571627

ABSTRACT

The efficacy of replication-deficient adenoviral vectors in gene therapy is confined to the number of tumor cells the vector infects. To focus and enhance the therapeutic efficacy, we employed a conditionally replication-competent adenoviral vector with a tissue-specific promoter, DF3/MUC1, in a human esophageal adenocarcinoma model. Our results demonstrate that Ad.DF3.E1A.CMV.TNF (Ad.DF3.TNF) specifically replicates in Bic-1 (DF3-producing cells) and mediates an enhanced biologic effect due to increased TNF-alpha in the same DF3-producing cells. We also show that the increased TNF-alpha interacts with ionizing radiation to produce greater tumor regression and a greater delay in tumor regrowth in Bic-1 (DF3-producing cells) compared to Seg-1 (DF3 non-producers). Tumor cell targeting using conditionally replication-competent adenoviral vectors with tumor-specific promoters to drive viral replication and deliver TNF-alpha provides a novel approach to enhancing tumor radiosensitivity.


Subject(s)
Adenocarcinoma/therapy , Esophageal Neoplasms/therapy , Genetic Therapy/methods , Mucin-1/genetics , Peptide Fragments/genetics , Adenocarcinoma/immunology , Adenocarcinoma/radiotherapy , Adenoviridae/genetics , Animals , Combined Modality Therapy , Esophageal Neoplasms/immunology , Esophageal Neoplasms/radiotherapy , Genetic Vectors/administration & dosage , Green Fluorescent Proteins , Humans , Luminescent Proteins/genetics , Mice , Mice, Nude , Models, Animal , Mucin-1/analysis , Peptide Fragments/analysis , Promoter Regions, Genetic , Random Allocation , Transplantation, Heterologous , Tumor Cells, Cultured , Tumor Necrosis Factor-alpha/biosynthesis , Virus Replication
4.
Int J Oncol ; 19(4): 833-5, 2001 Oct.
Article in English | MEDLINE | ID: mdl-11562763

ABSTRACT

We examined the interaction between Alimta and ionizing radiation (IR) as a potential strategy to enhance the therapeutic ratio of combined-modality cancer treatment. Mice bearing human esophageal adenocarcinoma xenografts (Seg-1) or squamous cell carcinoma xenografts (SQ-20B) were treated with Alimta and IR employing a fractionated treatment schedule. Treatment with Alimta alone slowed the growth of Seg-1 but not SQ-20B tumors compared with control tumors. In Seg-1 xenografts combined treatment with Alimta and IR produced significant tumor growth inhibition compared with Alimta alone or IR alone. In SQ-20B xenografts, treatment with Alimta did not enhance IR-mediated tumor growth inhibition suggesting that sensitivity to Alimta is necessary for an interactive cytotoxic effect with IR. The present data suggest the potential clinical efficacy of combining Alimta administration with radiotherapy for Alimta-sensitive cells and indicate that further testing needs to be conducted to optimize the dosing schedule to enhance the interaction between the therapeutic agents.


Subject(s)
Antineoplastic Agents/therapeutic use , Folic Acid Antagonists/therapeutic use , Glutamates/therapeutic use , Guanine/analogs & derivatives , Guanine/therapeutic use , Neoplasms/drug therapy , Neoplasms/radiotherapy , Adenocarcinoma/drug therapy , Adenocarcinoma/pathology , Adenocarcinoma/radiotherapy , Animals , Carcinoma, Squamous Cell/drug therapy , Carcinoma, Squamous Cell/pathology , Carcinoma, Squamous Cell/radiotherapy , Combined Modality Therapy , Dose-Response Relationship, Radiation , Esophageal Neoplasms/drug therapy , Esophageal Neoplasms/pathology , Esophageal Neoplasms/radiotherapy , Female , Head and Neck Neoplasms/drug therapy , Head and Neck Neoplasms/pathology , Head and Neck Neoplasms/radiotherapy , Humans , Mice , Mice, Nude , Neoplasm Transplantation , Neoplasms/pathology , Pemetrexed , Radiation, Ionizing , Transplantation, Heterologous , Tumor Cells, Cultured
5.
Gene Ther ; 8(24): 1840-6, 2001 Dec.
Article in English | MEDLINE | ID: mdl-11821937

ABSTRACT

Neointimal hyperplasia resulting from vascular smooth muscle cell (SMC) proliferation and luminal migration is the major cause of autologous vein graft failure following vascular coronary or peripheral bypass surgery. Strategies to attenuate SMC proliferation by the delivery of oligonucleotides or genes controlling cell division rely on the use of high concentrations of vectors, and require pre-emptive disruption of the endothelial cell layer. We report a genetically engineered herpes simplex virus (HSV-1) mutant that, in an in vivo rabbit model system, infects all vascular layers without prior injury to the endothelium; expresses a reporter gene driven by a viral promoter with high efficiency for at least 4 weeks; exhibits no systemic toxicity; can be eliminated at will by administration of the antiviral drug acyclovir; and significantly reduces SMC proliferation and restenosis in vein grafts in immunocompetent hosts.


Subject(s)
Genetic Therapy/methods , Genetic Vectors/administration & dosage , Graft Occlusion, Vascular/prevention & control , Herpesvirus 1, Human/genetics , Tunica Intima/pathology , Animals , Humans , Hyperplasia/prevention & control , Jugular Veins , Models, Animal , Muscle, Smooth, Vascular , Mutation , Organ Culture Techniques/methods , Rabbits , Recurrence , Saphenous Vein , Transfection/methods
6.
Cancer J ; 6(5): 287-93, 2000.
Article in English | MEDLINE | ID: mdl-11079167

ABSTRACT

PURPOSE: The purpose of this study was to evaluate whether endostatin, an antiangiogenic cleavage fragment of collagen XVIII, enhances the antitumor effects of ionizing radiation (IR). Endostatin was injected to coincide with fractionated radiotherapy. METHODS: Xenografts of radioresistant SQ-20B tumor cells were established in athymic nude mice. Lewis lung carcinoma cells were injected into C57BI/6 mice. Mice bearing SQ-20B xenografts were injected intraperitoneally with 2.5 mg/kg/day of murine recombinant endostatin 5 times per week for 2 weeks 3 hours before IR treatment (50 Gy total dose). Mice bearing Lewis lung carcinoma tumors were injected intraperitoneally with endostatin (2.5 mg/kg/day) four times; the first injection was given 24 hours before the first IR dose (15 Gy) and then 3 hours before IR (15 Gy/day) for 3 consecutive days. Microvascular density was assessed on tumor tissue sections by use of CD31 immunohistochemistry and light microscopy. Endothelial cell survival analyses were employed to evaluate endostatin effects on human aortic endothelial cells and human umbilical vein endothelial cells. Endothelial cell apoptosis was examined by use of FACS analysis and DAPI microscopy. RESULTS: In SQ-20B xenografts, combined treatment with endostatin and IR produced tumor growth inhibition that was most pronounced at the nadir of regression (day 21). By day 35, tumors receiving combined treatment with endostatin and IR were 47% smaller than tumors treated with endostatin alone. Interactive cytotoxic treatment effects between endostatin and IR were also demonstrated in mice bearing Lewis lung carcinoma tumors. Significant tumor growth inhibition was observed in the endostatin/IR group at days 11 and 13 compared with IR alone. Histologic analyses demonstrated a reduction in microvascular density after combined treatment with endostatin and IR compared with endostatin treatment alone. Survival analyses confirmed interactive cytotoxicity between endostatin and IR in both human aortic endothelial cells and human umbilical vein endothelial cells but not in SQ-20B tumor cells. Combined treatment with endostatin and IR produced an increase in cow pulmonary artery endothelial apoptosis compared with either treatment alone. DISCUSSION: The tumor regression observed after combined treatment with endostatin and IR suggests additive antitumor effects in both human and murine tumors. Importantly, the concentrations of endostatin employed produced little tumor regression when endostatin was employed as a single agent. The results from the clonogenic and apoptosis assays support the hypothesis that the endothelial compartment is the target for the endostatin/IR interaction.


Subject(s)
Antineoplastic Agents/therapeutic use , Collagen/therapeutic use , Neoplasms/drug therapy , Neoplasms/radiotherapy , Peptide Fragments/therapeutic use , Radiation, Ionizing , Animals , Apoptosis , Carcinoma, Lewis Lung/drug therapy , Cell Separation , Cells, Cultured , Cloning, Molecular , Collagen Type XVIII , Combined Modality Therapy , Dose-Response Relationship, Drug , Endostatins , Endothelium, Vascular/cytology , Endothelium, Vascular/drug effects , Escherichia coli/metabolism , Female , Flow Cytometry , Humans , Immunohistochemistry , Mice , Mice, Inbred C57BL , Microcirculation/drug effects , Microcirculation/radiation effects , Neoplasm Transplantation , Neoplasms/metabolism , Pichia/metabolism , Platelet Endothelial Cell Adhesion Molecule-1/biosynthesis , Recombinant Proteins/metabolism , Time Factors , Tumor Cells, Cultured , Umbilical Veins/cytology , Umbilical Veins/drug effects
7.
Mol Cell Biol Res Commun ; 3(5): 277-82, 2000 May.
Article in English | MEDLINE | ID: mdl-10964751

ABSTRACT

Angiostatin is an inhibitor of tumor angiogenesis that induces regression of experimental tumors and enhances the antitumor effects of radiation therapy. We report that the cytotoxic effects of angiostatin are restricted to the proliferating endothelial cell population. In addition, angiostatin and ionizing radiation (IR) interact by inducing death of dividing endothelial cells. We also show that angiostatin and IR interact to inhibit endothelial cell migration. These findings demonstrate that angiostatin targets the proliferating tumor vasculature and provide a mechanistic basis for the cytotoxic interaction of angiostatin and IR.


Subject(s)
Antineoplastic Agents/pharmacology , Cell Death/drug effects , Cell Division/drug effects , Endothelium, Vascular/cytology , Peptide Fragments/pharmacology , Plasminogen/pharmacology , Angiogenesis Inhibitors/metabolism , Angiogenesis Inhibitors/pharmacology , Angiostatins , Animals , Antineoplastic Agents/metabolism , Aorta/cytology , Cattle , Cell Migration Inhibition , Cells, Cultured , Endothelium, Vascular/physiology , Endothelium, Vascular/radiation effects , Humans , Mitosis , Peptide Fragments/genetics , Peptide Fragments/metabolism , Plasminogen/genetics , Plasminogen/metabolism , Radiation, Ionizing , Recombinant Proteins/metabolism , Umbilical Veins/cytology
8.
J Surg Res ; 89(2): 155-62, 2000 Apr.
Article in English | MEDLINE | ID: mdl-10729244

ABSTRACT

BACKGROUND: Although ionizing radiation (IR) has been demonstrated to attenuate vessel wall restenosis and intimal hyperplasia (IH), dose-related mural injury and atrophy are possible deleterious side effects. We tested the hypothesis that a radiosensitizing strategy may improve IR-induced inhibition of in vivo vascular smooth muscle cells (VSMCs) without influencing apoptotic cell death. METHODS: In 28 New Zealand White rabbits, the right common carotid artery (CCA) was injured and subjected to low-flow conditions to promote IH. The CCA was transfected with an adenoviral vector incorporating the cytosine deaminase (CD) gene (1 x 10(9) PFU/ml). 5-Fluorocytosine (5-FC), a prodrug that is converted to the radiosensitizing agent 5-fluorouracil (5-FU) by CD, was thereafter administered intravenously. The CCA was exposed to 5 Gy IR at 24 h. Intimal/medial (I/M) area and thickness ratios were determined in the harvested CCAs at 14 days. VSMC proliferative and apoptotic indices were assessed with immunohistochemistry. RESULTS: A 50% reduction in I/M area was found in rabbits treated with IR and IR + CD/5-FC (0.19 +/- 0.03 and 0.18 +/- 0.02) when compared with untreated controls (UC) (0.37 +/- 0.06) (P = 0.005). This finding was substantiated by attenuation of I/M thickness in the IR groups [0.47 +/- 0.13 (IR), 0.41 +/- 0.11 (IR + CD/5-FC), 0.61 +/- 0.17 (UC)] (P = 0.007). The number of proliferating VSMCs was notably smaller when IR was combined with CD/5-FC (4.17 +/- 1.16 vs 2.97 +/- 1.09 log transformed cells/mm(2), P < 0.07). Apoptosis was similar in all groups. CONCLUSIONS: Both IR alone and IR combined with a radiosensitizing agent are effective in attenuating experimental IH. However, combination therapy is synergistic and achieves greater inhibition of VSMC proliferation and may involve selective killing of radioresistant S-phase VSMCs. IR + CD/5-FC represents a novel therapeutic strategy that offers potential for long-term control of IH.


Subject(s)
Genetic Therapy , Tunica Intima/pathology , Tunica Intima/radiation effects , Animals , Apoptosis/drug effects , Apoptosis/radiation effects , Carotid Artery, Common/drug effects , Carotid Artery, Common/pathology , Carotid Artery, Common/physiopathology , Cell Division/drug effects , Cell Division/radiation effects , Cytosine Deaminase , Flucytosine/pharmacology , Hemodynamics/drug effects , Hemodynamics/radiation effects , Hyperplasia/pathology , Male , Muscle, Smooth, Vascular/drug effects , Muscle, Smooth, Vascular/pathology , Muscle, Smooth, Vascular/physiopathology , Nucleoside Deaminases/genetics , Prodrugs/pharmacology , Rabbits , Tunica Intima/drug effects
9.
Clin Cancer Res ; 6(2): 737-42, 2000 Feb.
Article in English | MEDLINE | ID: mdl-10690561

ABSTRACT

Although clonogenic or divisional death is the main mechanism by which DNA-damaging agents demonstrate antitumor activity, recent data indicate that strategies specifically designed to trigger apoptosis may also prove to be useful antitumor agents. Protein kinase C (PKC) isoenzymes are involved in the regulation of cell proliferation, differentiation, and survival. Whereas pharmacological inhibition of PKC activity triggers apoptosis in most mammalian cells, cell line and tissue differences in sensitivities to these inhibitors remain. Whereas PKC inhibitors have potential as antitumor agents, issue of kinase specificity and solubility have remained obstacles to their clinical use. In this report, we investigated the antitumor activity of the PKC inhibitor chelerythrine chloride (chelerythrine), a selective inhibitor of group A and B PKC isoforms. Chelerythrine exhibited cytotoxic activity against nine human tumor cell lines tested in vitro. On the basis of the finding that radioresistant and chemoresistant squamous cell carcinoma lines (HNSCC) undergo apoptosis rapidly after treatment with chelerythrine in vitro, we assessed the effects of this agent on p53-deficient SQ-20B HNSCC cells in vivo. The results demonstrate that chelerythrine treatment of nude mice bearing SQ-20B is associated with significant tumor growth delay. Significantly, treatment with chelerythrine resulted in minimal toxicity. These findings demonstrate a potential for chelerythrine as an antitumor drug against squamous cell carcinoma.


Subject(s)
Antineoplastic Agents/toxicity , Carcinoma, Squamous Cell/pathology , Head and Neck Neoplasms/pathology , Phenanthridines/toxicity , Alkaloids , Animals , Antineoplastic Agents/therapeutic use , Apoptosis/drug effects , Benzophenanthridines , Body Weight/drug effects , Carcinoma, Squamous Cell/drug therapy , Cell Division/drug effects , Enzyme Inhibitors/toxicity , Head and Neck Neoplasms/drug therapy , Humans , Mice , Mice, Nude , Phenanthridines/therapeutic use , Protein Kinase C/antagonists & inhibitors , Transplantation, Heterologous , Tumor Cells, Cultured
10.
Cancer Res ; 60(24): 6958-63, 2000 Dec 15.
Article in English | MEDLINE | ID: mdl-11156396

ABSTRACT

We examined the effects of a new antiangiogenic isocoumarin, NM-3, as a radiation modifier in vitro and in vivo. The present studies demonstrate that NM-3 is cytotoxic to human umbilical vein endothelial cells (HUVECs) but not to Lewis lung carcinoma (LLC) cells nor Seg-1, esophageal adenocarcinoma cells, in clonogenic survival assays. When HUVEC cultures are treated with NM-3 combined with ionizing radiation (IR), additive cytotoxicity is observed. In addition, the combination of NM-3 and IR inhibits HUVEC migration to a greater extent than either treatment alone. The effects of treatment with NM-3 and IR were also evaluated in tumor model systems. C57BL/6 female mice bearing LLC tumors were given injections for 4 consecutive days with NM-3 (25 mg/kg/day) and treated with IR (20 Gy) for 2 consecutive days. Combined treatment with NM-3 and IR significantly reduced mean tumor volume compared with either treatment alone. An increase in local tumor control was also observed in LLC tumors in mice receiving NM-3/IR therapy. When athymic nude mice bearing Seg-1 tumor xenografts were treated with NM-3 (100 mg/kg/day for 4 days) and 20 Gy (four 5 Gy fractions), significant tumor regression was observed after combined treatment (NM-3 and IR) compared with IR alone. Importantly, no increase in systemic or local tissue toxicity was observed after combined treatment (NM-3 and IR) when compared with IR alone. The bioavailability and nontoxic profile of NM-3 suggests that the efficacy of this agent should be tested in clinical radiotherapy.


Subject(s)
Coumarins/pharmacology , Neoplasms/drug therapy , Neoplasms/radiotherapy , Adenocarcinoma/drug therapy , Animals , Carcinoma, Lewis Lung/drug therapy , Cell Movement/drug effects , Cell Movement/radiation effects , Cells, Cultured , Collagen/metabolism , Coumarins/toxicity , Dose-Response Relationship, Drug , Dose-Response Relationship, Radiation , Drug Combinations , Endothelium, Vascular/cytology , Endothelium, Vascular/drug effects , Endothelium, Vascular/radiation effects , Esophageal Neoplasms/drug therapy , Female , Humans , Isocoumarins , Laminin/metabolism , Mice , Mice, Inbred C57BL , Mice, Nude , Neoplasm Transplantation , Proteoglycans/metabolism , Radiation, Ionizing , Time Factors , Tumor Cells, Cultured , Umbilical Veins/cytology , Umbilical Veins/drug effects , Umbilical Veins/radiation effects
11.
Cancer Res ; 59(14): 3374-8, 1999 Jul 15.
Article in English | MEDLINE | ID: mdl-10416597

ABSTRACT

The family of vascular endothelial growth factor (VEGF) proteins include potent and specific mitogens for vascular endothelial cells that function in the lation of angiogenesis Inhibition of VEGF-induced angiogenesis either by neutralizing antibodies or dominant-negative soluble receptor, blocks the growth of primary and metastatic experimental tumors Here we report that VEGF expression is induced in Lewis lung carcinomas (LLCs) both in vitro and vivo after exposure to ionizing radiation (IR) and in human tumor cell lines (Seg-1 esophageal adenocarcinoma, SQ20B squamous cell carcinoma, T98 and U87 glioblastomas, and U1 melanoma) in vitro. The biological significance of IR-induced VEGF production is supported by our finding that treatment of tumor-bearing mice (LLC, Seg-1, SQ20B, and U87) with a neutralizing antibody to VEGF-165 before irradiation is associated with a greater than additive antitumor effect. In vitro, the addition of VEGF decreases IR-induced killing of human umbilical vein endothelial cells, and the anti-VEGF treatment potentiates IR-induced lethality of human umbilical vein endothelial cells. Neither recombinant VEGF protein nor neutralizing antibody to VEGF affects the radiosensitivity of tumor cells These findings support a model in which induction of VEGF by IR contributes to the protection of tumor blood vessels from radiation-mediated cytotoxicity and thereby to tumor radioresistance.


Subject(s)
Antibodies, Monoclonal/pharmacology , Endothelial Growth Factors/antagonists & inhibitors , Gene Expression Regulation, Neoplastic/drug effects , Lymphokines/antagonists & inhibitors , Neoplasm Proteins/antagonists & inhibitors , Neoplasms, Experimental/radiotherapy , Neovascularization, Pathologic/physiopathology , Radiation-Sensitizing Agents/pharmacology , Stress, Physiological/physiopathology , Adenocarcinoma/genetics , Adenocarcinoma/pathology , Adenocarcinoma/radiotherapy , Animals , Carcinoma, Squamous Cell/genetics , Carcinoma, Squamous Cell/pathology , Carcinoma, Squamous Cell/radiotherapy , Cells, Cultured , Culture Media, Conditioned , Endothelial Growth Factors/immunology , Endothelial Growth Factors/physiology , Endothelium, Vascular/cytology , Esophageal Neoplasms/genetics , Esophageal Neoplasms/pathology , Esophageal Neoplasms/radiotherapy , Female , Glioblastoma/genetics , Glioblastoma/pathology , Glioblastoma/radiotherapy , Humans , Lung Neoplasms/genetics , Lung Neoplasms/pathology , Lymphokines/immunology , Lymphokines/physiology , Melanoma/genetics , Melanoma/pathology , Melanoma/radiotherapy , Mice , Mice, Inbred C57BL , Mice, Nude , Neoplasm Proteins/immunology , Neoplasm Proteins/physiology , Neoplasm Transplantation , Neoplasms, Experimental/blood supply , Neoplasms, Experimental/complications , Neoplasms, Experimental/physiopathology , RNA, Messenger/biosynthesis , RNA, Neoplasm/biosynthesis , Radiation Tolerance/drug effects , Stress, Physiological/genetics , Tumor Cells, Cultured/metabolism , Tumor Cells, Cultured/radiation effects , Tumor Stem Cell Assay , Vascular Endothelial Growth Factor A , Vascular Endothelial Growth Factors
12.
C R Acad Sci III ; 322(2-3): 225-8, 1999.
Article in English | MEDLINE | ID: mdl-10196676

ABSTRACT

The radiation-inducible chimeric genetic construct Egr-TNF alpha introduced into human xenografts produces cytotoxicity of infected tumor cells resulting in tumor growth inhibition. The interaction between Egr-TNF and radiation is selectively cytotoxic for the tumor microvasculature resulting in vascular thrombosis and tumor necrosis. Gene therapy combined with radiation therapy offers great potential for the treatment of localized human cancers.


Subject(s)
Gene Expression Regulation/radiation effects , Genetic Therapy/methods , Immediate-Early Proteins/genetics , Adenoviridae/genetics , DNA-Binding Proteins/genetics , Early Growth Response Protein 1 , Gene Targeting , Genetic Vectors , Humans , Transcription Factors/genetics , Transplantation, Heterologous , Tumor Necrosis Factor-alpha/therapeutic use
13.
Cancer Res ; 58(24): 5686-9, 1998 Dec 15.
Article in English | MEDLINE | ID: mdl-9865723

ABSTRACT

Angiostatin, a proteolytic fragment of plasminogen, inhibits the growth of primary and metastatic tumors by suppressing angiogenesis. When used in combination with ionizing radiation (IR), angiostatin demonstrates potent antitumor synergism, largely caused by inhibition of the tumor microvasculature. We report here the temporal interaction of angiostatin and IR in Lewis lung carcinoma (LLC) tumors growing in the hind limbs of syngeneic mice. Tumors with an initial mean volume of 510 +/- 151 mm3 were treated with IR alone (20 Gy x 2 doses on days 0 and 1), angiostatin alone (25 mg/kg/day divided twice daily) on days 0 through 13, or a combination of the two as follows: (a) IR plus angiostatin (days 0 through 13); (b) IR plus angiostatin (days 0 and 1); and (c) IR followed by angiostatin beginning on the day after IR completion and given daily thereafter (days 2 through 13). By day 14, tumors in untreated control mice had grown to 6110 +/- 582 mm3, whereas in mice treated with: (a) IR alone, tumors had grown to 2854 +/- 338 mm3 (P < 0.05 compared with untreated controls); and (b) angiostatin alone, tumors had grown to 3666 +/- 453 mm3 (P < 0.05 compared with untreated controls). In combined-treatment groups, in mice treated with: (a) IR plus longer-course angiostatin, tumors reached 2022 +/- 282 mm3 (P = 0.036 compared with IR alone); (b) IR followed by angiostatin, tumors reached 2677 +/- 469 mm3 (P > 0.05 compared with IR alone); and (c) IR plus short-course angiostatin, tumors reached 1032 +/- 78 mm3 (P < 0.001 compared with IR alone). These findings demonstrate that the efficacy of experimental radiation therapy is potentiated by brief concomitant exposure of the tumor vasculature to angiostatin.


Subject(s)
Antineoplastic Agents/therapeutic use , Carcinoma, Lewis Lung/drug therapy , Carcinoma, Lewis Lung/radiotherapy , Peptide Fragments/therapeutic use , Plasminogen/therapeutic use , Angiostatins , Animals , Combined Modality Therapy , Drug Administration Schedule , Female , Mice , Mice, Inbred C57BL , Peptide Fragments/administration & dosage , Plasminogen/administration & dosage , Time Factors , Tumor Cells, Cultured
14.
Nature ; 394(6690): 287-91, 1998 Jul 16.
Article in English | MEDLINE | ID: mdl-9685160

ABSTRACT

Angiogenesis, the formation of new capillaries from pre-existing vessels, is essential for tumour progression. Angiostatin, a proteolytic fragment of plasminogen that was first isolated from the serum and urine of tumour-bearing mice, inhibits angiogenesis and thereby growth of primary and metastatic tumours. Radiotherapy is important in the treatment of many human cancers, but is often unsuccessful because of tumour cell radiation resistance. Here we combine radiation with angiostatin to target tumour vasculature that is genetically stable and therefore less likely to develop resistance. The results show an antitumour interaction between ionizing radiation and angiostatin for four distinct tumour types, at doses of radiation that are used in radiotherapy. The combination produced no increase in toxicity towards normal tissue. In vitro studies show that radiation and angiostatin have combined cytotoxic effects on endothelial cells, but not tumour cells. In vivo studies show that these agents, in combination, target the tumour vasculature. Our results provide support for combining ionizing radiation with angiostatin to improve tumour eradication without increasing deleterious effects.


Subject(s)
Antineoplastic Agents/therapeutic use , Carcinoma, Lewis Lung/drug therapy , Carcinoma, Lewis Lung/radiotherapy , Peptide Fragments/therapeutic use , Plasminogen/therapeutic use , Angiostatins , Animals , Apoptosis , Carcinoma, Lewis Lung/blood supply , Combined Modality Therapy , Endothelium, Vascular/drug effects , Endothelium, Vascular/radiation effects , Female , Humans , Mice , Mice, Inbred C57BL , Neoplasm Transplantation , Neovascularization, Pathologic/drug therapy , Neovascularization, Pathologic/radiotherapy , Recombinant Proteins/therapeutic use , Tumor Cells, Cultured
15.
Gene Ther ; 5(3): 293-300, 1998 Mar.
Article in English | MEDLINE | ID: mdl-9614548

ABSTRACT

We evaluated the antitumor effects of ionizing radiation and tumor necrosis factor-alpha (TNF-alpha) gene therapy in human malignant glioma (D54) xenografts. An adenoviral vector (Ad5) containing DNA sequences of the Egr-1 promoter was linked to a cDNA encoding the TNF-alpha gene (Ad. Egr-TNF). Athymic nude mice bearing D54 xenografts received intratumoral injections of Ad.Egr-TNF or the null vector (Ad.null), with and without fractionated radiation, 5 gray (Gy) per day for 6 days, a total dose of 30 Gy. Administration of Ad.Egr-TNF and 30 Gy resulted in complete tumor regression in 71% of xenografts compared with xenografts treated with radiation alone (7.4%, P = 0.006), Ad.Egr-TNF alone (0%, P = 0.012) or Ad.null with 30 Gy (0%, P = 0.002). Combined treatment with Ad.Egr-TNF and 30 Gy significantly reduced mean fractional tumor volumes compared with radiation alone (P = 0.002), Ad.Egr-TNF alone (P = 0.002) and Ad.null plus 30 Gy (P = 0.018). Histopathologic analyses of glioma xenografts treated with Ad.Egr-TNF and radiation revealed tumor vessel thrombosis by day 4 and necrosis by day 7. Thrombosis was not observed in tumors treated with Ad.Egr-TNF alone and was significantly reduced in all other treatment groups. These studies suggest that in the D54 glioma xenograft model, the antitumor effects of combining radiation and Ad.Egr-TNF are mediated, in part, by the destruction of the tumor microvasculature.


Subject(s)
Adenoviridae/genetics , Genetic Therapy , Glioma/therapy , Immediate-Early Proteins , Tumor Necrosis Factor-alpha/genetics , Animals , Cell Survival/drug effects , Cell Survival/radiation effects , Combined Modality Therapy , DNA-Binding Proteins/genetics , Early Growth Response Protein 1 , Female , Genetic Vectors , Glioma/radiotherapy , Humans , In Situ Hybridization , Mice , Mice, Nude , Neoplasm Transplantation , Recombinant Proteins/pharmacology , Transcription Factors/genetics , Transplantation, Heterologous , Tumor Cells, Cultured , Tumor Necrosis Factor-alpha/pharmacology , Zinc Fingers/genetics
16.
Cancer Gene Ther ; 5(6): 344-9, 1998.
Article in English | MEDLINE | ID: mdl-9917088

ABSTRACT

The purpose of the present study was to determine the therapeutic potential of combining radiotherapy with tumor necrosis factor (TNF)-alpha-based gene therapy in the human prostate cancer PC-3 xenograft. PC-3 cells are highly resistant to TNF-alpha-induced cytotoxicity in vitro. A modest enhancement of radiation killing was observed with the addition of TNF-alpha in clonogenic survival assays. Combined treatment with Ad.Egr-TNF, a replication-deficient adenovirus modified to express TNF-alpha following the exposure of infected cells to ionizing radiation (40 Gy administered at 5 Gy per fraction) in vivo, resulted in increased tumor control, as defined by a reduction of tumor volume, when compared with treatment with Ad.Egr-TNF alone or with radiation alone (P < .03). The improvement in tumor control was achieved without increasing acute normal tissue damage when compared with tissue injury from radiation alone. The results of these studies support further development and clinical application of genetic radiotherapy for human prostate cancer.


Subject(s)
Genetic Therapy , Prostatic Neoplasms/therapy , Tumor Necrosis Factor-alpha/genetics , Adenoviridae/genetics , Animals , Cell Survival/drug effects , Combined Modality Therapy , Dose-Response Relationship, Drug , Female , Humans , Male , Mice , Neoplasm Transplantation , Prostatic Neoplasms/radiotherapy , Transfection , Transplantation, Heterologous , Tumor Cells, Cultured , Tumor Necrosis Factor-alpha/pharmacology , Tumor Necrosis Factor-alpha/therapeutic use
17.
Cancer Res ; 57(19): 4205-9, 1997 Oct 01.
Article in English | MEDLINE | ID: mdl-9331076

ABSTRACT

Gene therapy combined with radiation therapy to enhance selectively radiation cytotoxicity in malignant cells represents a new approach for cancer treatment. We investigated the efficacy of adenoviral (Ad5)-directed cytosine deaminase/5-fluorocytosine (CD/5-FC) enzyme/prodrug gene therapy to enhance selectively the tumoricidal action of ionizing radiation in human cancer xenografts derived from a human squamous carcinoma cell line (SQ-20B). Tumor xenografts grown in hindlimbs of nude mice were transfected with an adenoviral vector (Ad.CMV.CD) containing the cytosine deaminase (CD) gene under the control of a cytomegalovirus (CMV) promoter. Mice were injected i.p. with 800 mg/kg of 5-FC for 12 days, and tumors were treated with fractionated radiation at a dose of 5 Gy/day to a total dose of 50 Gy. In larger tumors with a mean volume of 1069 mm3, marked tumor regression to 11% of the original tumor volume was observed at day 21 (P = 0.01). The volumetric regression of smaller tumors with a mean volume of 199 mm3, which received the same combined treatment protocol, was significant at day 12 (P = 0.014). However, unlike large tumors, regression of the smaller tumors continued until day 36 (P = 0.01), with 43% cured at day 26. No cures or significant volumetric reduction in size was observed in tumors treated with radiation alone; Ad.CMV.CD with or without radiation; or with Ad.CMV.CD and 5-FC. These results suggest that the CD/5-FC gene therapy approach is an effective radiosensitizing strategy and may lead to substantial improvement in local tumor control that would translate into improved cure rates and better survival.


Subject(s)
Antimetabolites, Antineoplastic/therapeutic use , Flucytosine/therapeutic use , Genetic Therapy , Neoplasms, Experimental/therapy , Nucleoside Deaminases/therapeutic use , Radiation-Sensitizing Agents/therapeutic use , Adenoviruses, Human/genetics , Animals , Carcinoma, Squamous Cell/drug therapy , Carcinoma, Squamous Cell/radiotherapy , Carcinoma, Squamous Cell/therapy , Combined Modality Therapy , Cytosine Deaminase , Female , Genetic Vectors , Humans , Laryngeal Neoplasms/drug therapy , Laryngeal Neoplasms/radiotherapy , Laryngeal Neoplasms/therapy , Mice , Mice, Nude , Neoplasm Recurrence, Local , Neoplasm Transplantation , Neoplasms, Experimental/drug therapy , Neoplasms, Experimental/pathology , Neoplasms, Experimental/radiotherapy , Nucleoside Deaminases/genetics , Radiation Tolerance , Recombinant Fusion Proteins/therapeutic use , Transfection , Transplantation, Heterologous
18.
Cancer Res ; 57(19): 4340-7, 1997 Oct 01.
Article in English | MEDLINE | ID: mdl-9331096

ABSTRACT

Approximately 30% of cancer deaths result from the failure to control local and regional tumors. The goal of radiotherapy is to maximize local and regional tumor cell killing while minimizing normal tissue destruction. Attempts to enhance radiation-mediated tumor cell killing using halogenated pyrimidines, antimetabolites, and other DNA-damaging agents or sensitizers of hypoxic tumor cells have met with only modest clinical success. In an unique strategy to modify tumor radiosensitivity, we used an inhibitor of the protein kinase C group A and B isoforms, chelerythrine chloride (chelerythrine), to enhance the killing effects of ionizing radiation (IR). Protein kinase C activity plays a central role in cellular proliferation, differentiation, and apoptosis. Chelerythrine increases sphingomyelinase activity and enhances IR-mediated cell killing through induction of apoptotic tumor cell death in a radioresistant tumor model both in vitro and in vivo. Although previous reports have suggested that IR-mediated apoptosis correlates with tumor volume reduction, we demonstrate for the first time that lowering the apoptotic threshold increases tumor cell killing in vivo.


Subject(s)
Apoptosis/drug effects , Carcinoma, Squamous Cell/radiotherapy , Craniocerebral Trauma/radiotherapy , Enzyme Inhibitors/pharmacology , Isoenzymes/antagonists & inhibitors , Neoplasm Proteins/metabolism , Phenanthridines/pharmacology , Protein Kinase C/antagonists & inhibitors , Radiation-Sensitizing Agents/therapeutic use , Sphingomyelin Phosphodiesterase/metabolism , Alkaloids , Animals , Benzophenanthridines , Carcinoma, Squamous Cell/drug therapy , Carcinoma, Squamous Cell/enzymology , Ceramides/pharmacology , Chemotherapy, Adjuvant , Combined Modality Therapy , Craniocerebral Trauma/drug therapy , Craniocerebral Trauma/enzymology , Endopeptidases/metabolism , Enzyme Activation/drug effects , Isoenzymes/metabolism , Mice , Mice, Nude , Neoplasm Proteins/antagonists & inhibitors , Protein Kinase C/metabolism , Radiation-Sensitizing Agents/pharmacology , Transplantation, Heterologous
19.
Radiat Oncol Investig ; 5(5): 220-6, 1997.
Article in English | MEDLINE | ID: mdl-9372544

ABSTRACT

Intratumoral injection of an adenoviral vector containing radiation-inducible DNA sequences of the early growth response gene (Egr-1) promoter ligated to a cDNA encoding tumor necrosis factor-alpha (TNF-alpha; Ad.Egr-TNF) increases the radiation killing of a human radioresistant xenograft (SQ-20B). Viral dose-escalation experiments demonstrated that SQ-20B growth inhibition correlated with viral titer. Injection of 5 x 10(8) PFU Ad.Egr-TNF produced regression to a mean volume of 22 +/- 13% of the original tumor volume, 1 x 10(8) PFU to a mean of 62 +/- 24%, and 5 x 10(7) PFU to a mean of 67 +/- 27%. No regression was observed when tumors were injected with 1 x 10(7) PFU Ad.Egr-TNF or with the null viral vector (Ad.null). When two injections of vector (2 x 10(8) PFU Ad.Egr-TNF) were combined with 50 Gy, a significant increase in tumor regression was observed compared with injection of buffer, Ad.Egr-TNF, or 50 Gy. The interactive killing between TNF and radiation was enhanced significantly (P = 0.05) when the number of injections was increased from two to five while maintaining a constant viral titer (2 x 10(8) PFU Ad.Egr-TNF) and a constant radiation dose (50 Gy). Significant TNF-alpha levels were present in irradiated vs. unirradiated tumors following injection with Ad.Egr-TNF. Taken together, these data suggest that the volumetric reduction produced by the combined effects of Ad.Egr-TNF and radiation is enhanced with increasing vector concentration and the number of vector injections.


Subject(s)
Carcinoma, Squamous Cell/therapy , Genetic Therapy/methods , Genetic Vectors/administration & dosage , Immediate-Early Proteins , Adenoviridae , Animals , Carcinoma, Squamous Cell/radiotherapy , Combined Modality Therapy/methods , DNA-Binding Proteins/pharmacology , Early Growth Response Protein 1 , Female , Humans , Injections, Intralesional , Mice , Mice, Nude , Neoplasm Transplantation , Transcription Factors/pharmacology , Transplantation, Heterologous , Tumor Necrosis Factor-alpha/pharmacology , Zinc Fingers
20.
Cancer Res ; 56(19): 4311-4, 1996 Oct 01.
Article in English | MEDLINE | ID: mdl-8813113

ABSTRACT

Intratumoral injection of an adenoviral vector containing radiation-inducible DNA sequences of the Egr-1 promoter linked to a cDNA encoding tumor necrosis factor (TNF) alpha (Ad.Egr-TNF) enhances the tumoricidal action of ionizing radiation in a human epidermoid carcinoma xenograft (SQ-20B). The dominant histopathological feature in tumor-bearing animals treated with Ad.Egr-TNF and irradiation is extensive intratumoral vascular thrombosis and tumor necrosis. Thrombosis and necrosis are not observed in animals treated with either the viral construct encoding TNF-alpha or radiation and did not occur in irradiated normal tissues adjacent to tumor in animals injected with Ad.Egr-TNF. To determine if the occlusive effects of Ad.Egr-TNF and X-irradiation were specific for tumor vessels, non-tumor-bearing mice were irradiated after receiving i.m. injection of Ad.Egr-TNF at viral titers 20-100 times greater than titers injected intratumorally. No vascular thrombosis was observed in the treated normal tissues. Combined Ad.Egr-TNF and radiation produced occlusion of tumor microvessels without significant normal tissue damage. Taken together, these data suggest that the interaction between radiation inducible TNF-alpha and X-irradiation occurs selectively within the tumor vessels.


Subject(s)
Carcinoma, Squamous Cell/therapy , Gene Expression Regulation/radiation effects , Genetic Therapy , Neovascularization, Pathologic/genetics , Tumor Necrosis Factor-alpha/genetics , Animals , Carcinoma, Squamous Cell/blood supply , Carcinoma, Squamous Cell/pathology , Female , Genetic Vectors/genetics , Genetic Vectors/radiation effects , Humans , Mice , Mice, Nude , Necrosis , Neoplasm Transplantation , Recombinant Fusion Proteins/biosynthesis , Thrombosis/etiology , Tumor Necrosis Factor-alpha/biosynthesis
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