ABSTRACT
Breast cancer is the most common cause of cancer-related death worldwide, thus remaining a crucial health problem among women despite advances in conventional therapy. Therefore, new alternative strategies are needed for effective diagnosis and treatment. One approach is the use of oncolytic viruses for gene-directed enzyme prodrug therapy. Here, the lacZ-carrying vaccinia virus (VACV) strain GLV-1h68 was used in combination with a ß-galactosidase-activatable prodrug derived from a seco-analog of the natural antibiotic duocarmycin SA. Tumor cell infection with the VACV strain GLV-1h68 led to production of ß-galactosidase, essential for the conversion of the prodrug to the toxic compound. Furthermore, drug-dependent cell kill and induction of the intrinsic apoptosis pathway in tumor cells was also observed on combination therapy using the prodrug and the GLV-1h68 strain, despite the fact that VACV strains encode antiapoptotic proteins. Moreover, GI-101A breast cancer xenografts were effectively treated by the combination therapy. In conclusion, the combination of a ß-galactosidase-activatable prodrug with a tumor-specific vaccinica virus strain encoding this enzyme, induced apoptosis in cultures of the human GI-101A breast cancer cells, in which a synergistic oncolytic effect was observed. Moreover, in vivo, additional prodrug treatment had beneficial effects on tumor regression in GLV-1h68-treated GI-101A-xenografted mice.
Subject(s)
Breast Neoplasms/drug therapy , Breast Neoplasms/therapy , Indoles/chemistry , Prodrugs/chemistry , Prodrugs/therapeutic use , Vaccinia virus/genetics , beta-Galactosidase/metabolism , Animals , Cell Line, Tumor , Cell Survival , Duocarmycins , Female , Humans , Mice , Mice, Nude , Microscopy, Fluorescence , Oncolytic Virotherapy , Pyrroles/chemistry , Vaccinia virus/physiology , Xenograft Model Antitumor Assays , beta-Galactosidase/geneticsABSTRACT
Mammary cancers together with cancers of the skin account for about 60% of the total cancers occurring in dogs. The veterinary options for therapeutic management of canine mammary cancer are limited and prognosis for such patients is poor. In this study, we analyzed the functionality of the oncolytic vaccinia virus strain GLV-1h68 as a possible therapeutic agent for canine mammary cancer. Cell culture data demonstrated that GLV-1h68 efficiently infected and destroyed cells of the canine mammary adenoma cell line ZMTH3. Furthermore, after systemic administration this attenuated vaccinia virus strain primarily replicated in canine tumor xenografts in nude mice. The efficient tumor colonization process resulted in inhibition of tumor growth and drastic reduction of tumor size. This is the first report demonstrating that vaccinia virus is an effective tool for the therapy of canine mammary cancers, which might next be applied to dogs with breast tumors.
Subject(s)
Adenoma/therapy , Adenoma/veterinary , Mammary Neoplasms, Animal/therapy , Oncolytic Virotherapy/methods , Vaccinia virus/physiology , Adenoma/blood , Animals , Antibodies, Viral/blood , Apoptosis/drug effects , Cell Line, Tumor , Dogs , Female , Haplorhini , Mammary Neoplasms, Animal/blood , Mice , Mice, Nude , Oncolytic Viruses/physiology , Virus Replication , Xenograft Model Antitumor AssaysABSTRACT
The addition of double-stranded circular or linear DNA encoding EGFP (the Enhanced Green Fluorescent Protein) to a Listeria -containing infection medium resulted in up to 8.6% COS-1 cells expressing the reporter protein. The transfer of naked DNA into host cells upon infection by Listeria was found to be dependent on the ability of the bacteria to synthesize internalins and listeriolysin. Since no binding of DNA to the bacterial cells was detected, DNA uptake seems to be the consequence of the simultaneous entry of infection medium, and thus of naked DNA, via the phagosomes induced by the bacterium to facilitate its own entry into the host cells.
Subject(s)
DNA/metabolism , Gene Transfer Techniques , Genetic Vectors , Green Fluorescent Proteins/metabolism , Listeria monocytogenes/metabolism , Animals , Bacterial Proteins/metabolism , Bacterial Toxins/metabolism , COS Cells , Chlorocebus aethiops , DNA Primers , Flow Cytometry , Green Fluorescent Proteins/genetics , Heat-Shock Proteins/metabolism , Hemolysin Proteins , Listeria monocytogenes/genetics , Microscopy, Electron, Transmission , Microscopy, Fluorescence , Mutagenesis , PhagocytosisABSTRACT
Bacteria-mediated transfer of plasmid DNA into mammalian cells (bactofection) is a potent approach to express plasmid-encoded heterologous proteins (protein antigens, toxins or enzymes) in a large set of different cell types including phagocytic and nonphagocytic mammalian cells. Previously, we have described a Listeria monocytogenes-mediated DNA delivery system, which releases plasmid DNA directly into the cytosol of mammalian cells by partial self-destruction of the carrier bacteria. Here we report on a second generation of this phage lysin supported bactofection system, which is greatly improved with respect to plasmid stability, transfer efficacy and biosafety. In this case, DNA release is initiated by spontaneous bacterial lysis in the infected cells cytosol which is subsequently enhanced by the simultaneously released phage lysin produced by the intracellular carrier bacteria. Bacteria that are capable of cell-to-cell spread are found to be much more efficient in bactofection than their non spreading counterparts.
