RESUMO
The combination of radiation with radiosensitizing gene delivery or oncolytic viruses promises to provide an advantage that could improve the therapeutic results for glioblastoma. X-rays can induce significant molecular changes in cancer cells. We isolated the GIRLRG peptide that binds to radiation-inducible 78 kDa glucose-regulated protein (GRP78), which is overexpressed on the plasma membranes of irradiated cancer cells and tumor-associated microvascular endothelial cells. The goal of our study was to improve tumor-specific adenovirus-mediated gene delivery by selectively targeting the adenovirus binding to this radiation-inducible protein. We employed an adenoviral fiber replacement approach to conduct a study of the targeting utility of GRP78-binding peptide. We have developed fiber-modified adenoviruses encoding the GRP78-binding peptide inserted into the fiber-fibritin. We have evaluated the reporter gene expression of fiber-modified adenoviruses in vitro using a panel of glioma cells and a human D54MG tumor xenograft model. The obtained results demonstrated that employment of the GRP78-binding peptide resulted in increased gene expression in irradiated tumors following infection with fiber-modified adenoviruses, compared with untreated tumor cells. These studies demonstrate the feasibility of adenoviral retargeting using the GRP78-binding peptide that selectively recognizes tumor cells responding to radiation treatment.
Assuntos
Adenoviridae/genética , Técnicas de Transferência de Genes , Vetores Genéticos/genética , Radiação Ionizante , Adenoviridae/metabolismo , Animais , Antígenos de Neoplasias/imunologia , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , Linhagem Celular Tumoral , Modelos Animais de Doenças , Chaperona BiP do Retículo Endoplasmático , Feminino , Expressão Gênica , Genes Reporter , Terapia Genética , Vetores Genéticos/administração & dosagem , Glioma/genética , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Integrina alfaVbeta3/metabolismo , Camundongos , Terapia Viral Oncolítica , Vírus Oncolíticos/genética , Receptores de Vitronectina/metabolismo , Transdução Genética , Transgenes , Raios X , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Pancreatic cancer is a significant clinical problem and novel therapeutic approaches are desperately needed. Recent advances in conditionally replicative adenovirus-based (CRAd) oncolytic virus design allow the application of CRAd vectors as a therapeutic strategy to efficiently target and eradicate chemoresistant pancreatic cancer cells, thereby improving the efficacy of pancreatic cancer treatment. The goal of this study was to construct and validate the efficacy of an infectivity-enhanced, liver-untargeted, tumor-specific CRAd vector. A panel of CRAds has been derived that embodies the C-X-C chemokine receptor type 4 promoter for conditional replication, two-fiber complex mosaicism for targeting expansion and hexon hypervariable region 7 (HVR7) modification for liver untargeting. We evaluated CRAds for cancer virotherapy using a human pancreatic tumor xenograft model. Employment of the fiber mosaic approach improved CRAd replication in pancreatic tumor xenografts. Substitution of the HVR7 of the Ad5 hexon for Ad serotype 3 hexon resulted in decreased liver tropism of systemically administrated CRAd. Obtained data demonstrated that employment of complex mosaicism increased efficacy of the combination of oncolytic virotherapy with chemotherapy in a human pancreatic tumor xenograft model.
Assuntos
Adenoviridae/genética , Terapia Viral Oncolítica/métodos , Neoplasias Pancreáticas/terapia , Neoplasias Pancreáticas/virologia , Receptores CXCR4/genética , Animais , Feminino , Vetores Genéticos/genética , Humanos , Camundongos , Camundongos Nus , Distribuição Aleatória , Receptores CXCR4/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Neoplasias PancreáticasRESUMO
Malignant forms of glioma, the most common primary brain tumors, remain poorly responsive to multimodality therapeutic interventions, including chemotherapy. Suppressed apoptosis and extraordinary invasiveness are important distinctive features that contribute to the malignant phenotype of glioma. We have developed the vascular endothelial growth factor receptor 1 (VEGFR-1/flt-1) conditional replicating adenoviral vector (CRAdRGDflt-IL24) encoding the interleukin-24 (IL-24) gene. We investigated whether a combination of CRAdRGDflt-IL24-mediated oncolytic virotherapy and chemotherapy using temozolomide (TMZ) produces increased cytotoxicity against human glioma cells in comparison with these agents alone. Combination of CRAdRGDflt-IL24 and TMZ significantly enhanced cytotoxicity in vitro, inhibited D54MG tumor growth and prolonged survival of mice harboring intracranial human glioma xenografts in comparison with CRAdRGDflt-IL24 or TMZ alone. These data indicate that combined treatment with CRAdRGDflt-IL24-mediated oncolytic virotherapy and TMZ chemotherapy provides a promising approach for glioma therapy.
Assuntos
Neoplasias Encefálicas/terapia , Dacarbazina/análogos & derivados , Glioma/terapia , Interleucinas/genética , Terapia Viral Oncolítica/métodos , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genética , Adenoviridae/genética , Animais , Antineoplásicos Alquilantes/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/virologia , Processos de Crescimento Celular/genética , Linhagem Celular Tumoral , Terapia Combinada , Dacarbazina/farmacologia , Feminino , Terapia Genética/métodos , Vetores Genéticos/genética , Glioma/tratamento farmacológico , Glioma/genética , Glioma/virologia , Humanos , Camundongos , Camundongos Nus , Regiões Promotoras Genéticas , Proteínas Recombinantes/farmacologia , Temozolomida , Fator A de Crescimento do Endotélio Vascular/farmacologia , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/biossíntese , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Combined treatment using adenoviral (Ad)-directed enzyme/prodrug therapy and radiation therapy has the potential to become a powerful method of cancer therapy. We have developed an Ad vector encoding a mutant bacterial cytosine deaminase (bCD) gene (AdbCD-D314A), which has a higher affinity for cytosine than wild-type bCD (bCDwt). The purpose of this study was to evaluate cytotoxicity in vitro and therapeutic efficacy in vivo of the combination of AdbCD-D314A with the prodrug 5-fluorocytosine (5-FC) and ionizing radiation against human glioma. The present study demonstrates that AdbCD-D314A infection resulted in increased 5-FC-mediated cell killing, compared with AdbCDwt. Furthermore, a significant increase in cytotoxicity following AdbCD-D314A and radiation treatment of glioma cells in vitro was demonstrated as compared to AdbCDwt. Animal studies showed significant inhibition of subcutaneous or intracranial tumor growth of D54MG glioma xenografts by the combination of AdbCD-D314A/5-FC with ionizing radiation as compared with either agent alone, and with AdbCDwt/5-FC plus radiation. The results suggest that the combination of AdbCD-D314A/5-FC with radiation produces markedly increased cytotoxic effects in cancer cells in vitro and in vivo. These data indicate that combined treatment with this novel mutant enzyme/prodrug therapy and radiotherapy provides a promising approach for cancer therapy.
Assuntos
Adenoviridae/genética , Neoplasias Encefálicas/terapia , Citosina Desaminase/genética , Terapia Genética/métodos , Vetores Genéticos/administração & dosagem , Glioma/terapia , Animais , Antimetabólitos/uso terapêutico , Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/metabolismo , Linhagem Celular Tumoral , Terapia Combinada , Citosina/metabolismo , Citosina Desaminase/metabolismo , Escherichia coli/enzimologia , Flucitosina/uso terapêutico , Genes Transgênicos Suicidas , Vetores Genéticos/genética , Glioma/diagnóstico por imagem , Glioma/metabolismo , Humanos , Camundongos , Camundongos Nus , Mutação , Transplante de Neoplasias , Pró-Fármacos/uso terapêutico , Radiografia , Transplante HeterólogoRESUMO
Combined treatment using adenoviral-directed enzyme/prodrug therapy and immunotherapy has the potential to become a powerful alternative method of cancer therapy. We have developed adenoviral vectors encoding the cytosine deaminase gene (Ad-CD) and cytosine deaminase:uracil phosphoribosyltransferase fusion gene (Ad-CD:UPRT). A monoclonal antibody, TRA-8, specifically binds to death receptor 5, one of two death receptors bound by tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). The purpose of this study was to evaluate cytotoxicity in vitro and therapeutic efficacy in vivo of the combination of Ad-CD:UPRT and TRA-8 against human pancreatic cancer and glioma cell lines. The present study demonstrates that Ad-CD:UPRT infection resulted in increased 5-FC-mediated cell killing, compared with Ad-CD. Furthermore, a significant increase of cytotoxicity following Ad-CD:UPRT/5-FC and TRA-8 treatment of cancer cells in vitro was demonstrated. Animal studies showed significant inhibition of tumor growth of MIA PaCa-2 pancreatic and D54MG glioma xenografts by the combination of Ad-CD:UPRT/5-FC plus TRA-8 as compared with either agent alone or no treatment. The results suggest that the combination of Ad-CD:UPRT/5-FC with TRA-8 produces an additive cytotoxic effect in cancer cells in vitro and in vivo. These data indicate that combined treatment with enzyme/prodrug therapy and TRAIL immunotherapy provides a promising approach for cancer therapy.
Assuntos
Adenoviridae/genética , Anticorpos Monoclonais/uso terapêutico , Citosina Desaminase/genética , Genes Transgênicos Suicidas/genética , Terapia Genética/métodos , Glioma/terapia , Imunoterapia/métodos , Neoplasias Pancreáticas/terapia , Análise de Variância , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/metabolismo , Western Blotting , Linhagem Celular Tumoral , Testes Imunológicos de Citotoxicidade , Feminino , Citometria de Fluxo , Glioma/imunologia , Humanos , Camundongos , Camundongos Nus , Microscopia de Fluorescência , Neoplasias Pancreáticas/imunologia , Pentosiltransferases/genética , Pró-Fármacos/uso terapêutico , Receptores do Ligante Indutor de Apoptose Relacionado a TNF , Receptores do Fator de Necrose Tumoral/metabolismoRESUMO
Overexpression of vascular endothelial growth factor (VEGF) and its cognate receptor KDR has been linked to a more aggressive phenotype of human prostate carcinomas. The importance of signal transduction through the VEGF receptor 2 is illustrated by use of soluble KDR, which binds to VEGF and sequesters this ligand before its binding to cellular receptor. Treatment with recombinant adenovirus AdVEGF-sKDR, encoding sKDR under control of the human VEGF promoter, significantly inhibited the proliferation of human vascular endothelial cells and prostate cancer cells. AdVEGF-sKDR infection decreased migration of endothelial 1P-1B cells (61% reduction) and DU145 prostate carcinoma cells (47%) in comparison with AdCMV-Luc-infected control cells. Ionizing radiation upregulated VEGF promoter activity in prostate carcinoma and endothelial cells. AdVEGF-sKDR infection significantly reduced human vascular endothelial and prostate cancer cell proliferation and sensitized cancer cells to ionizing radiation. In vivo tumor therapy studies demonstrated significant inhibition of DU145 tumor growth in mice that received combined AdVEGF-sKDR infection and ionizing radiation versus AdVEGF-sKDR alone or radiation therapy alone. These results suggest that selective transcriptional targeting of sKDR gene expression employing a radiation inducible promoter can effectively inhibit tumor growth and demonstrate the advantage of combination radiotherapy and gene therapy for the treatment of prostate cancer.
Assuntos
Adenoviridae/genética , Terapia Genética/métodos , Vetores Genéticos/administração & dosagem , Neoplasias da Próstata/terapia , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Animais , Proliferação de Células , Terapia Combinada , Endotélio Vascular/patologia , Regulação da Expressão Gênica/efeitos da radiação , Vetores Genéticos/genética , Humanos , Masculino , Camundongos , Camundongos Nus , Regiões Promotoras Genéticas , Neoplasias da Próstata/patologia , Neoplasias da Próstata/radioterapia , Proteínas Recombinantes/metabolismo , Transdução Genética/métodos , Células Tumorais Cultivadas , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
The parameters of nonspecific immunity (interferon, interleukin-1, tumor necrosis factor, and natural killers) changed in immune BALB/c mice after challenge with Machupo virus in doses of 1000 and 5000 PFU. After challenge with 1000 PFU the activity of the above parameters increased during the first three days and no cases of animal death occurred. After challenge with 5000 PFU the maximal values of interferon, interleukin-1, tumor necrosis factor, and natural killers were observed on days 5-7, the animals dying at the height of these values. Hence, the formation of specific humoral and cell-mediated immune response in BALB/c mice immunized with inactivated Machupo virus does not protect the animals from infection with the homologous virus in a dose of 5000 PFU. High mediator activity of nonspecific immunity factors on days 5-7 after infection augments the disease course and leads to earlier death of vaccinated animals in comparison with the same dose of the virus in nonimmune mice.
Assuntos
Arenavirus do Novo Mundo , Febre Hemorrágica Americana/imunologia , Vacinas de Produtos Inativados/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais/biossíntese , Divisão Celular , Linhagem Celular , Linfócitos/citologia , Linfócitos/imunologia , Camundongos , Camundongos Endogâmicos BALB CAssuntos
Infecções por Arenaviridae/terapia , Desferroxamina/uso terapêutico , Febres Hemorrágicas Virais/terapia , Choque/terapia , Fator de Necrose Tumoral alfa/imunologia , Animais , Infecções por Arenaviridae/complicações , Infecções por Arenaviridae/metabolismo , Moléculas de Adesão Celular/biossíntese , Febres Hemorrágicas Virais/complicações , Febres Hemorrágicas Virais/metabolismo , Soros Imunes , Interleucina-1/sangue , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos CBA , Choque/etiologiaRESUMO
Describes changes in nonspecific immunity parameters (interferon, interleukin-1, tumor necrosis factor, and natural killers) in the course of experimental Bolivian fever (Machupo virus). Changes of these parameters were followed up after urgent prophylactic injections of Ridostin and specific gamma-globulin to infected animals. The possibility of treatment of experimental Machupo fever by intranasal administration of interferon inductor Ridostin has been demonstrated.
Assuntos
Febre Hemorrágica Americana/prevenção & controle , Animais , Antivirais/farmacologia , Chlorocebus aethiops , Febre Hemorrágica Americana/imunologia , Indutores de Interferon/farmacologia , Interferons/biossíntese , Interferons/imunologia , Interleucina-1/imunologia , Células Matadoras Naturais/imunologia , Camundongos , Camundongos Endogâmicos BALB C , RNA de Cadeia Dupla/farmacologia , RNA Fúngico/farmacologia , Fator de Necrose Tumoral alfa/imunologia , Células Vero , gama-Globulinas/farmacologiaRESUMO
The possibility of using different strains of parotitis virus (Enders, L-3, Jeryl-Leen) as antigens for enzyme immunoassay (EIA) to titer antibodies in human and animal blood sera is analyzed. Methods for preparation and purification of antigen on the basis of the said parotitis virus strains have been developed. Conditions of EIA were optimized. The sensitivity and specificity of EIA and hemagglutination inhibition test were compared.
Assuntos
Respirovirus/imunologia , Animais , Anticorpos Antivirais/análise , Anticorpos Antivirais/imunologia , Antígenos Virais/imunologia , Western Blotting , Chlorocebus aethiops , Humanos , Técnicas Imunoenzimáticas , Microscopia Eletrônica , Caxumba/sangue , Caxumba/diagnóstico , Testes de Neutralização , Respirovirus/ultraestrutura , Células VeroRESUMO
Some immunity parameters (interferon, tumor necrosis factor, interleukin 1, etc.) were studied in CBA/Calac mice infected with Lassa virus. The results permit a hypothesis that a pathologic inflammatory reaction is responsible for the death of animals in experimental Lassa fever. One of the components of this reaction is endogenous shock involving a manifest production of immune response mediators, such as interferon, interleukin 1. and tumor necrosis factor.
Assuntos
Febre Lassa/imunologia , Animais , Linhagem Celular , Chlorocebus aethiops , Citotoxicidade Imunológica , Humanos , Interferons/biossíntese , Interleucina-1/biossíntese , Células Matadoras Naturais/imunologia , Febre Lassa/metabolismo , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos CBA , Células Tumorais Cultivadas , Fator de Necrose Tumoral alfa/biossíntese , Células VeroRESUMO
The paper presents the data characterizing parameters of specific and nonspecific immunity in BALB/c mice immunized with gamma-ray-inactivated Machupo virus antigen or its formalinized antigen. The gamma-ray inactivated preparation was shown to be more immunogenic for BALB/c mice. A certain relationship between the time course of activity of nonspecific immunity factors in the immunized animals and the protective activity of the preparation under study was also noted. The decisive role of the T-cell part of the immune system was demonstrated in the resistance of this model animal to Machupo virus infection.
Assuntos
Antígenos Virais/imunologia , Arenavirus do Novo Mundo/imunologia , Febre Hemorrágica Americana/imunologia , Imunização , Camundongos Endogâmicos BALB C/imunologia , Animais , Animais Lactentes , Formação de Anticorpos , Antígenos Virais/efeitos dos fármacos , Antígenos Virais/efeitos da radiação , Arenavirus do Novo Mundo/efeitos dos fármacos , Arenavirus do Novo Mundo/efeitos da radiação , Febre Hemorrágica Americana/prevenção & controle , Imunidade Celular , Imunidade Inata , Imunização/métodos , Camundongos , Vacinas de Produtos Inativados/imunologia , Vacinas de Produtos Inativados/efeitos da radiação , Vacinas Virais/imunologia , Vacinas Virais/efeitos da radiaçãoRESUMO
The pathomorphological patterns and the activity of serum interferon, interleukin-1, tumor necrosis factor, natural killers, and proliferative activity of lymphocytes were studied in BALB/c and C57B1/6 mice intracerebrally infected with Machupo virus. The BALB/c mice showed 100% lethality at 8-9 days after inoculation while C57B1/6 mice were found nonsusceptible to Machupo virus inoculation by this route. The pathomorphological findings at the peak of clinical manifestations in BALB/c mice revealed no organ whose functional deficiency could lead to the death of the animals. Investigations of nonspecific immunity parameters revealed a direct dependence between their high activity and susceptibility of the animals to Machupo virus infection. It is assumed that the endogenous shock due to the high activity of immune response mediators is the cause of death in Machupo virus infection.