RESUMO
IGF-I antisense gene therapy has been applied successfully to animal models of glioma, hepatoma and teratocarcinoma. The antisense strategy has shown that tumor cells transfected with vectors encoding IGF-I antisense RNA lose tumorigenicity, become immunogenic and are associated with tumor specific immune response involving CD8+ lymphocytes. An IGF-I triple helix approach to gene therapy for glioma was recently described. The approach we have taken is to establish parameters of change using the IGF-I triple helix strategy. PCC-3 embryonal carcinoma cells derived from murine teratocarcinoma which express IGF-I were used as a model. The cells were transfected with vector which encodes an oligoribonucleotide that forms RNA-IGF-I DNA triple-helix structure. The triple-helix stops the production of IGF-I. Cells transfected in this manner underwent changes in phenotype and an increase in MHC-I and B-7 cell surface molecules. They also showed enhancement in the production of apoptotic cells (60-70%). The "triple helix" transfected cells lost the ability to induce tumor when injected subcutaneously in syngeneic 129 Sv mice. When co-transfected in vitro with expression vectors encoding both MHC-I and B-7 cDNA in antisense orientation, the "triple-helix" transfected cells were down-regulated in expression of MHC-I and B-7 and the number of apoptotic cells was significantly decreased. Injection of the doubly co-transfected cells into 129 Sv mice was associated with induction of teratocarcinoma. Comparison between antisense and triple-helix transfected cells strategies showed similar immunogenic and apoptotic changes. The findings suggest that triple-helix technology may offer a new clinical approach to treatement of tumors expressing IGF-I.
Assuntos
Apoptose , Carcinoma Embrionário/imunologia , DNA , Fator de Crescimento Insulin-Like I/genética , Animais , Antígeno B7-1/genética , Antígeno B7-1/imunologia , Sequência de Bases , Carcinoma Embrionário/genética , Carcinoma Embrionário/patologia , Carcinoma Embrionário/terapia , Citometria de Fluxo , Técnica Indireta de Fluorescência para Anticorpo , Expressão Gênica , Terapia Genética , Vetores Genéticos , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/imunologia , Marcação In Situ das Extremidades Cortadas , Fator de Crescimento Insulin-Like I/metabolismo , Camundongos , Dados de Sequência Molecular , Transplante de Neoplasias , RNA Antissenso/genética , Transfecção , Células Tumorais CultivadasRESUMO
Recently we demonstrated that rat glioma cells when transfected with a vector encoding antisense IGF-I c-DNA lost tumorigenicity and induced a tumor specific immune response involving CD8+ lymphocytes. Here we showed, using immunostaining flow cytometry analysis, that the transfected cell lines, rat C-6 glioma and rat LF hepatoma, expressed an increase level of MHC-class I, and even the amount of MHC-I was found to be higher in the transfected hepatoma, than in the transfected glioma cells. This increased expression of MHC-I could contribute to the final immune recognition of tumour immunogenicity.
