Herpes simplex virus thymidine kinase/ganciclovir system in multicellular tumor spheroids

• Autores: Finocchiaro, L.M.E.; Bumaschny, V.F.; Karara, A.L.; Fiszman, G.L.; Casais, C.C.; Glikin, G.C.
• Año de publicación: 2004
• Idioma: inglés
• Tipo de recurso: artículo
• Fuente: Cancer Gene Ther. 2004;11(5):333-345
• Materia: Lipofection; Murine adenocarcinoma; Suicide gene therapy; ganciclovir; suicide substrate; thymidine kinase; animal cell; animal tissue; apoptosis; article; breast adenocarcinoma; bystander effect; cell growth; cell proliferation; cell specificity; cell type; controlled study; drug cytotoxicity; drug efficacy; gene expression regulation; gene transfer; Herpes simplex virus; in vitro study; in vivo study; intracellular transport; melanoma cell; mitochondrion; monolayer culture; mouse; multicellular spheroid; nonhuman; phenotype; population structure; priority journal; protein assembly; sensitivity analysis; statistical significance; transgene; virus resistance; Adenocarcinoma; Animals; Antiviral Agents; bcl-2-Associated X Protein; beta-Galactosidase; Bystander Effect; Cell Division; Cell Line, Tumor; Drug Resistance; Ganciclovir; Gene Therapy; Mammary Neoplasms, Animal; Melanoma; Mice; Mice, Inbred BALB C; Proto-Oncogene Proteins c-bcl-2; Simplexvirus; Spheroids, Cellular; Thymidine Kinase; Transfection; Viral Proteins; Animalia; Murinae; Simplexvirus
• Institución: Universidad Nacional de Buenos Aires. Facultad de Ciencias Exactas y Naturales

We have developed multicellular spheroids (MCS) established from LM05e and LM3 spontaneous Balb/c-murine mammary adenocarcinoma and B16 C57-murine melanoma derived cell lines as an in vitro model to study the efficacy of the herpes simplex virus thymidine kinase/ganciclovir (HSVtk/GCV) suicide system. We demonstrated for the first time that HSVtk-expressing cells assembled as MCS manifested a GCV resistance phenotype compared to the same cells grown as sparse monolayers. HSVtk-expressing LM05e, LM3 and B16 spheroids were 16-, three- and nine-fold less sensitive to GCV than their respective monolayers, even though they could express transgenes 10-, eight- and five-fold more efficiently. Mixed populations of HSVtk- and their respective βgal-expressing cells displayed a cell-type specific bystander effect that was higher in monolayers than in MCS. However, HSVtk-expressing cells in two- or three-dimensional cultures were always significantly more sensitive to GCV than the βgal-expressing counterparts, supporting the feasibility of this suicide approach in vivo. We present evidence showing that HSVtk-expressing tumor cells, when transferred from monolayers to MCS, displayed: (i) lower GCV cytotoxic activity and bystander effect; (ii) higher and efficient expression of genes transferred as lipoplexes; (iii) lower cell proliferation rates; and (iv) changes in intracellular Bax/Bcl-xL rheostat of mitochondria-mediated apoptosis.
Fil:Finocchiaro, L.M.E. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.
Fil:Casais, C.C. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.
Fil:Glikin, G.C. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.