Gene delivery luciferase

Polyanhydride Nanoparticles Polyanhydrides have been more commonly used to prepare microparticles than nanoparticles. However, the technology is adaptable for nanoparticles. The transfection efficiency of firefly luciferase DNA was enhanced when delivered in nanoparticles prepared from polyanhydride-lactic acid blends, demonstrating the potential application in gene delivery [120], The degradation and elimination of polyanhydrides have been reviewed [97], In vivo, the anhydride bond degrades to form diacid monomers that are eliminated from the body. 

Fig. 3. Property of gene delivery with BLs and US exposure (a) Schema of transfection mechanism by BLs and US. The mechanical effect based on the disruption of BLs by US exposure, which results in generation of some pores on plasma membrane, is associated with direct delivery of extracellular plasmid DNA into cytosol, (b) Luciferase expression in COS-7 cells transfected by BLs and US. COS-7 cells (1x10 cells/500 pLAube) were mixed wifh pCMV-Luc (5 pg) and BLs (60 pg). The cell mixture was exposed with US (Frequency 2 MHz, Duty 50%, Burst rate 2 Hz, Intensity 2.5 W/ cm. Time 10 s). The cells were washed and cultured for 2 days. Affer fhaf, luciferase acfivify was measured, (c) Effecf of US condition on transfection efficiency with BLs. COS-7 cells were exposed with US (Frequency 2 MHz, Duty 50%, Burst rate 2 Hz, Intensity 2.5 W/cm Time 0,1, 5,10 s) in the presence of pCMV-Luc (0.25 pg) and BLs (60 pg). Luciferase activity was measured as above, (d) Effect of serum on transfection efficiency of BLs. COS-7 cells in the medium containing EBS (0,10, 30, 50% (v/v)) were treated with US (Erequency 2 MHz, Duty 50%, Burst rate 2 Hz, Intensity 2.5 W/cm, Time 10 s), pCMV-Luc (0.25 pg) and BLs (60 pg) or transfected with lipoplex of pCMV-Luc (0.25 pg) and lipofectin (1.25 pg). (e) In vitro gene delivery to various types of cell using BLs and US. The method of gene delivery was same as above. S-180 mouse sarcoma cells, Colon26 mouse colon adenocarcinoma cells, B16BL6 mouse melanoma cells, Jurkat human T cell line, HUVEC human umbilical endothelial cells. Luciferase activity was measured as above. <10 RLU/mg protein, <10 RLU/mg protein Each data represents the mean S.D. n=3). L PEG-liposomes, LF Lipotectin...

Fig. 4. In vivo gene delivery into mouse ascites tumor cells with Bubble liposomes. S-180 cells (1x10 cells) were i.p. injected into ddY mice. After 8 days, the mice were anaesthetized, then injected with 510 p.L of pCMV-Luc (10 p.g) and Bubble liposomes (500 pg) in PBS. Ultrasound (frequency 1 MHz, duty 50% intensity 1.0 W/cm, time 1 min) was transdermally applied to the abdominal area. In another experiment, pCMV-Luc (10 pg) - Lipofectin (50 pg) or Lipofectamine 2000 (50 pg) complex was suspended in PBS (510 pL) and injected into the peritoneal cavity of mice. After 2 days, S-180 cells were recovered from the abdomens of the mice. Luciferase activity was determined, as described in Materials and Methods. Each bar represents the mean S.D. (/j=3-6). P<0.01 compared to the group treated with plasmid DNA, Bubble liposomes, ultrasound exposure or lipofection with Lipofectin or Lipofectamine 2000. LF, Lipofectin. LF2000, Lipofectamine 2000. <10 BLU/mg protein...

Figure 19 Construct 28 was used for luciferase gene delivery to hepatocytes by noncovalent association of the gene to GalNAc cluster 24.

While the applications of SELP-mediated controlled gene delivery are numerous, we have focused our efforts on controlled delivery to solid tumors. Our initial studies in this arena involved the intratumoral delivery of a reporter plasmid Renilla luciferase) to solid tumors in a murine (athymic nu/nu) model of human breast cancer (MDA-MB-435 cell line) (50). Delivery of the Renilla luciferase plasmid from SELP-47K matrices resulted in significantly enhanced tumor transfection for up to 21 days compared to naked DNA (Fig. 5). In particular, delivery of the plasmid from matrices containing 4 or 8 wt.% polymer resulted in enhanced transfection... 

Figure 11 (a) Structures of the click cluster delivery vehicles five monodisperse structures were developed that vary in the length of the oligoethyleneamine arms where x- 0 (9a), 1 (9b), 2 (9c), 3 (9d), and 4 (9e). (b) Luciferase reporter gene delivery experiments with the analogs described in (a) at N/P ratios 5,10,20, and 50. Transfection increases as the arm length increases, without having a dramatic effect on cytotoxicity. Adapted from Srinivasachari, S. Fichter, K. M. Reineke, T. M. J. Am. Chem. Soc. 2008, 130, 4618. ... 

The PIC micelles showed prolonged in vivo circulation, as shown by luciferase gene expression in the liver that lasted for 3 days after systemic administration. Various PEG-containing PLL-based architectures were synthesized and the effects of PEGylation and molecular shape on their physicochemical and biological properties in gene delivery were evaluated. ... 

Figure 4.2 (See color insert) Adeno-associated viral delivery of dual function reporter gene to the lungs of Balb/c mice. Mice were treated with two different AAV vectors encoding the same dual function reporter gene comprised of Flue and YFP (Brindle and Contag, Unpubhshed data). Animals were imaged for luciferase expression (a and b) and after the study fluorescence micrographs (c and d) were used to validate gene delivery to the lungs. Effective gene delivery is shown in b and d, whereas the absence of effective delivery is shown in a and b.

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