AvidinOX™ for tissue targeted delivery of biotinylated cells (original) (raw)
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Purpose: the “avidin-biotin system” has been extensively studied for multiple applications to treat solid tumors. In this work, we investigated the effect of a new avidin chase, PWT- Biot1, on the uptake of two different 111In-labelled biotin derivatives in mice limbs intramuscularly pre-injected with avidin. Methods: two new biotin derivatives (r-BHD and Bis18) were radiolabelled with Indium- 111 and injected intravenously in a Balb/c mice (n=48) pretreated with an intramuscular injection of avidin in the left limb and an inert injection of Matrigel® in the right limb, as negative control. Twelve mice received 111In-r-BHD and twelve received 111In-Bis18. After 30 min, 1 h, 4 h and 24 h, 3 mice per group were anesthetized to acquire in vivo planar images, followed by ex-vivo organ counting. After the synthesis of PWT-Biot1, the same in vivo experiments were performed in two more groups of 12 mice each, but administering the PWT-Biot1 chase, for removing circulating avidin, 10 min before radiolabelled biotin injection.
Stem Cells, 2009
Muscle injuries in sport activities can pose challenging problems in traumatology and sports medicine. The best treatment for muscle injury has not been clearly established except for the conservative treatment that is routinely performed. We investigated the potential of human adult CD1331 cells to contribute to skeletal muscle regeneration in an athymic rat model. We tested whether CD1331 cells locally transplanted to the skeletal muscle lacerated models could (a) induce vasculogenesis/angiogenesis, (b) differentiate into endothelial and myogenic lineages, and (c) finally promote histological and functional skeletal myogenesis. Granulocyte colony stimulating factor-mobilized peripheral blood (PB) CD1331 cells, PB mononuclear cells, or phosphate-buffered saline was locally injected after creating a muscle laceration in the tibialis anterior muscle in athymic rats. After treatment, histological and functional skeletal myogenesis was observed significantly in the CD1331 group. The injected CD1331 cells differentiated into endothelial and myogenic lineages. Using real-time polymerase chain reaction analysis, we found that the gene expressions related to microenvironment conduction for host angiogenesis, fibrosis, and myogenesis were ideally up/downregulated. Our results show that CD1331 cells have the potential to enhance the histological and functional recovery from skeletal muscle injury rather via indirect contribution to environment conduction for muscular regeneration. It would be relatively easy to purify this cell fraction from PB, which could be a feasible and attractive autologous candidate for skeletal muscle injuries in a clinical setting. These advantages could accelerate the progression of cell-based therapies for skeletal muscle injuries from laboratory to clinical implementation. STEM CELLS 2009;27:949-960
AvidinOX™ for Highly Efficient Tissue-Pretargeted Radionuclide Therapy
Cancer Biotherapy and Radiopharmaceuticals, 2010
Avidin is widely used in vitro for its capacity to bind biotin. However, avidin's in vivo use is limited by its short residence in blood and tissues. An avidin variant, named AvidinOX,Ô has been recently described. This product is obtained by 4-hydroxyazobenzene-2 0-carboxylic acid-assisted sodium periodate oxidation of avidin. This method generates aldehyde groups from avidin carbohydrates, sparing biotin-binding sites from inactivation. AvidinOX binds cellular and interstitial protein amino groups through Schiff's bases, resulting in a tissue halflife of 2 weeks, compared with 2 hours of native avidin. Binding of AvidinOX occurs in normal and neoplastic tissues. Data show that AvidinOX, administered intranipple in the breast of transgenic BALB=neuT mice, is highly efficient for capturing 90 Y-biotinDOTA, intravenously injected after 48 hours, leading to eradication of multifocal cancer lesions. Efficacy data, together with good tolerability results, indicate that AvidinOX is a highly innovative reagent for tissue-pretargeted radionuclide therapy.
Poor nuclear entry, especially into nondividing cells, is a limiting factor in nonviral gene delivery. We have engineered a novel chimeric vector relying on the controlled assembly of a TAT-tagged multisubunit DNA binding protein (EcoR124I) with expression plasmids containing the EcoR124I recognition site. Molecular interactions of this molecular assembly were studied by electrophoretic mobility shift assay and atomic force microscopy. Maintenance of nanocomplexes in an appropriate stoichiometric ratio was both necessary and sufficient to produce a significant (>8-fold) increase in the activity of the therapeutic ␣-galactosidase A enzyme after intramuscular administration in the mouse model of Fabry disease. To our knowledge, this is the first molecular targeting system significantly enhancing plasmid-based expression in skeletal muscle. Coinjection with pluronic SP1017 produced further enhancement of gene expression, demonstrating cumulative effects of the increased nuclear delivery by TAT chimeras and transcription activation by the pluronic. Cell penetration peptides (CPP), such as TAT, have been shown to improve delivery of macromolecules, when linked directly. However, in our system TAT-enhanced targeting took place even though it was linked to the plasmid DNA molecule indirectly via two noncovalent bonds. Therefore, this proof-of principle result indicates that TAT (and potentially other CPP) can be used for targeting modular chimeric vectors and therapeutic nanodevices.-Lavigne, M. D., Yates, L., Coxhead, P., Gó recki, D. C. Nuclear-targeted chimeric vector enhancing nonviral gene transfer into skeletal muscle of Fabry mice in vivo. FASEB J. 22,
Faseb Journal, 2008
Poor nuclear entry, especially into nondividing cells, is a limiting factor in nonviral gene delivery. We have engineered a novel chimeric vector relying on the controlled assembly of a TAT-tagged multisubunit DNA binding protein (EcoR124I) with expression plasmids containing the EcoR124I recognition site. Molecular interactions of this molecular assembly were studied by electrophoretic mobility shift assay and atomic force microscopy. Maintenance of nanocomplexes in an appropriate stoichiometric ratio was both necessary and sufficient to produce a significant (>8-fold) increase in the activity of the therapeutic ␣-galactosidase A enzyme after intramuscular administration in the mouse model of Fabry disease. To our knowledge, this is the first molecular targeting system significantly enhancing plasmid-based expression in skeletal muscle. Coinjection with pluronic SP1017 produced further enhancement of gene expression, demonstrating cumulative effects of the increased nuclear delivery by TAT chimeras and transcription activation by the pluronic. Cell penetration peptides (CPP), such as TAT, have been shown to improve delivery of macromolecules, when linked directly. However, in our system TAT-enhanced targeting took place even though it was linked to the plasmid DNA molecule indirectly via two noncovalent bonds. Therefore, this proof-of principle result indicates that TAT (and potentially other CPP) can be used for targeting modular chimeric vectors and therapeutic nanodevices.-Lavigne, M. D., Yates, L., Coxhead, P., Gó recki, D. C. Nuclear-targeted chimeric vector enhancing nonviral gene transfer into skeletal muscle of Fabry mice in vivo. FASEB J. 22,
Journal of Pharmaceutical Sciences, 2000
Here we examine whether attachment of biotinylated antibodies to proteins on the cell surface increases the transfection efficiency of polyethylenimine-avidin-DNA bioconjugate gene transfer. Preliminary experiments were performed to compare avidin endocytosis into cells incubated with biotinylated antibodies. Antibody biotinylation resulted in the endocytosis of avidin-FITC into nearly 100% of cells compared with no detectable binding or entry into unbiotinylated cells. Gene transfer was accomplished with avidin conjugated to polyethylenimine (PEI) at a molar ratio of 4:1 (PA4). Plasmid DNA encoding the green fluorescent protein (GFP) gene was condensed on the PA4, and transfection efficiencies were measured by flow cytometry as the percentage of cells that fluoresced at levels greater than two standard deviations above the negative control. Gene transfer efficiencies were compared among K562, HEL, and Jurkat leukemia cell lines. Control transfections with DNA alone or untargeted PEI-DNA resulted in Յ2% GFP positive cells. Targeting PEI-avidin-DNA to antibody biotinylated cells increased transfection efficiency several fold over untargeted PEI. For each cell type, the increase in transfection efficiency was not significantly different among four biotinylated antibodies tested (antiCD55, antiCD59, antiCD71, and antiCD98). These data suggest biotinylated antibodies may be useful for targeting polyethylenimine-avidin mediated gene transfer.
Journal of Histochemistry & Cytochemistry, 1986
The capacity of the avidin-biotin system for post-embedding cytochemical detection of a biotin-labeled proteinaceous tracer was investigated. By testing modifications of the fixatives, the epoxy embedding medium, and various etching solutions, a procedure was developed to localize specifically the biotinylated tracer on 1-micron thick and thin sections. By use of this technique, a systemically administered IgG tracer was demonstrated after 24 hr throughout the endomysium of mouse skeletal muscle. In the adjoining sciatic nerve, the tracer IgG occurred at the perineurium and within endoneurial blood vessels, although the endoneurium itself was spared because of the presence of the blood-nerve barrier. Because of the small size of the biotin ligand and the non-denaturing method of labeling proteins, our mode of application of the avidin-biotin system appears suitable for tracer studies.
Targeted and Stable Gene Delivery into Muscle Cells by a Two-Step Transfer System
Biochemical and Biophysical Research Communications, 2000
We developed a muscle-specific gene delivery system based on two-step gene transfer. The first step involved adenovirus-mediated transfer of the ecotropic retrovirus receptor (EcoRec) gene driven by the muscle-specific desmin promoter. Both human primary myoblasts and fibroblasts were efficiently transduced with this adenovirus vector. However, expression of EcoRec was detected only in myoblasts. In the second step, EcoRec-expressing myoblasts could be stably transduced with the ecotropic retroviral vector with the -galactosidase gene. Approximately 15% of myoblasts were transduced by this two-step strategy. When the transduced myoblasts were differentiated into myotubes, extensive cell-cell fusion occurred, and the apparent number of -galactosidase-positive cells increased to 28%. These results indicate that our two-step gene delivery system could be used for targeted and stable gene transfer into muscle cells.