RESUMO
Antiangiogenic therapy using Semliki Forest virus (SFV) carrying Endostatin gene for malignant brain tumor was investigated to improve the therapeutic efficacy. The efficiency of SFV-mediated gene delivery was first evaluated for B 16 cells and compared with the efficiency in cells of endothelial origin (HMVECs). HMVECs are more susceptible to SFV infection than B 16 cells. For the in vivo treatment model, phosphate-buffered saline, SFV-LacZ, retrovirus vector GCsap-Endostatin, and SFV-Endostatin were injected to mice bearing B 16 brain tumors. A very significant inhibition of tumor growth was observed in the group that had been treated with SFV-Endostatin. A marked reduction of intratumoral vascularization was seen in the tumor sections from the SFV-Endostatin group compared with tumor sections from the SFV-LacZ or GCsap-Endostatin groups. Moreover, at day 7 after intravenous administration of SFV-Endostatin, the serum level of endostatin was augmented more than 3-fold compared to that after intravenous administration of GCsap-Endostatin. The results indicated that treatment with SFV-Endostatin inhibited the angiogenesis with established tumors. Gene therapy with Endostatin delivered via SFV may be a candidate for the development of new therapy for brain tumors.
Assuntos
Neoplasias Encefálicas/terapia , Colágeno/genética , Endotélio Vascular/metabolismo , Terapia Genética/métodos , Melanoma Experimental/terapia , Neovascularização Patológica/terapia , Fragmentos de Peptídeos/genética , Vírus da Floresta de Semliki/fisiologia , Animais , Neoplasias Encefálicas/irrigação sanguínea , Neoplasias Encefálicas/virologia , Células Cultivadas , Colágeno/sangue , Endostatinas , Endotélio Vascular/virologia , Feminino , Expressão Gênica , Técnicas de Transferência de Genes , Vetores Genéticos/administração & dosagem , Humanos , Melanoma Experimental/irrigação sanguínea , Melanoma Experimental/virologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Fragmentos de Peptídeos/sangue , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Taxa de SobrevidaRESUMO
OBJECT: The aim of this study was to further investigate dendritic cell (DC)-based immunotherapy for malignant glioma to improve its therapeutic efficacy. METHODS: Dendritic cells were isolated from the bone marrow and pulsed with phosphate-buffered saline, tumor RNA, tumor lysate, Semliki Forest virus (SFV)-LacZ, SFV-mediated B16 complementary (c)DNA, or SFV-mediated 203 glioma cDNA, respectively, to treat mice bearing tumors of the 203 glioma cell line. The results indicated that pre-immunization with DCs pulsed with the same type of cDNA as in the tumor by a self-replicating RNA vector (that is, SFV) protected mice from tumor challenge, and that therapeutic immunization prolonged the survival of mice with established tumors. The SFV induced apoptosis in DCs and their death facilitated the uptake of apoptotic cells by other DCs, thus providing a potential mechanism for enhanced immunogenicity. CONCLUSIONS: Therapy with DCs that have been pulsed with SFV-mediated tumor cDNA may be an excellent procedure for the development of new cancer vaccines.
Assuntos
Neoplasias Encefálicas/terapia , Células Dendríticas/imunologia , Terapia Genética/métodos , Vetores Genéticos , Glioma/terapia , Imunoterapia/métodos , Vírus da Floresta de Semliki , Animais , Apoptose/genética , Apoptose/imunologia , Células da Medula Óssea/citologia , Neoplasias Encefálicas/mortalidade , Linfócitos T CD8-Positivos/imunologia , DNA Complementar , Células Dendríticas/citologia , Glioma/mortalidade , Imunização , Melanoma , Camundongos , Camundongos Endogâmicos C57BL , Taxa de Sobrevida , Transfecção , Células Tumorais CultivadasRESUMO
Alphavirus vectors based on Sindbis virus and Semliki Forest virus (SFV) were characterized as potential gene transfer vectors. Initial studies were performed using vectors engineered to transfer either lacZ or green fluorescent protein (GFP). High levels of gene transfer were achieved in human primary fibroblasts, BHK and 293T cells, with low levels of transduction observed in more than 20 other target cells. Alphavirus-based expression was generally very high, but transient in every cell type. Replication-competent alphavirus was never detected in SFV preparations but could be produced by Sindbis-based vectors at a frequency of up to 3 x 10(-3) infectious units per ml. We constructed a human clotting factor IX (hFIX) cDNA-containing Sindbis virus and compared it with hFIX cDNA-harboring adenoviral and retroviral vectors. In most cases, hFIX levels obtained with Sindbis vector were initially at least an order of magnitude higher than those obtained with other viral vectors. These data demonstrate that alphavirus vectors compare favorably with adenovirus vectors as systems to promote high-level transient gene expression and should be considered as an alternative vector for gene transfer and potential gene therapy studies.
Assuntos
Alphavirus/genética , Técnicas de Transferência de Genes , Terapia Genética/métodos , Vetores Genéticos/administração & dosagem , Animais , Linhagem Celular , Estudos de Avaliação como Assunto , Feminino , Expressão Gênica , Humanos , Óperon Lac , Camundongos , Ratos , Vírus da Floresta de Semliki/genética , Sindbis virus/genética , Células Tumorais CultivadasRESUMO
OBJECT: The authors investigated immunogene therapy for malignant glioma to determine whether its therapeutic efficacy could be improved. METHODS: Four groups of 203-glioma-bearing mice were treated with injections of phosphate-buffered saline, Semliki Forest virus (SFV)-LacZ, retrovirus vector DFG-interleukin (IL)-12, and SFV-IL12, respectively. The results indicated that therapeutic immunization with SFV-IL12 prolonged the survival of mice with established tumors. Semliki Forest virus induces apoptotic death to glioma cells, which facilitates the uptake of apoptotic cells by dendritic cells, providing a potential mechanism for enhanced immunogenicity. CONCLUSIONS: Immunogene therapy with IL-12 via SFV may be an excellent candidate for the development of new cancer vaccines.