Effect of innate glutathione levels on activity of redox-responsive gene delivery vectors

Abstract

Redox-responsive polyplexes represent a promising class of non-viral gene delivery vectors. The reducible disulfide bonds in the polyplexes undergo intracellular reduction owing to the presence of high concentrations of reduced glutathione (GSH). Available evidence suggests improved transfection activity of redox-sensitive polyplexes upon artificial modulation of intracellular GSH. This study investigates the effect of innate differences in GSH concentration in a panel of human pancreatic cancer cell lines on activity of reducible polyplexes of the four major classes of nucleic acid therapeutics: plasmid DNA (pDNA), messenger RNA (mRNA), antisense oligodeoxynucleotides (AON) and siRNA. In general, reducible polyplexes of linear poly(amido amines) (PAA) show improved activity compared to non-reducible polyplexes of PAA. Results demonstrate that increased GSH levels are associated with improved transfection of mRNA polyplexes but no clear trend is observed for pDNA, AON and siRNA polyplexes.

Tuning multiple arms for camptothecin and folate conjugations on star-shaped copolymers to enhance glutathione-mediated intracellular drug delivery

“…8 Recently, the release of payloads from micelles in response to intrinsic intracellular signals, in particular cytoplasmic glutathione (GSH), has been explored for the construction of stimulus-responsive carriers. 9 GSH is produced intracellularly and is maintained at mM levels in the cytosol and subcellular compartments, whereas rapid enzymatic degradation limits GSH concentrations to μM levels in plasma. 10 Additionally, GSH metabolism is associated with cancer development and drug resistance, and the GSH concentration in some cancer cells is several times higher than that in normal cells.…”

The yin of exofacial protein sulfhydryls and the yang of intracellular glutathione in in vitro transfection with SS14 bioreducible lipoplexes

“…Moreover, we speculate that this behavior is specific for bioreducible lipoplexes. Nonetheless, it is worth noting that the outcome of the reduction process mediated by EPTs is likely to be determined by numerous factors including the site of disulfide introduction within the gene transfer vector [9,47], the type of non-viral vehicle used [13,49], the dose of complexes delivered to cells [23] as well as the concentration and the site where the reducing species reside [6][7][8][9].…”

“…Within a variety of antioxidant molecules, early reports pinpointed intracellular GSH as being central for transfection with bioreducible liposomes and polymers [7,8]. GSH is the most abundant cytoplasmic and nuclear reductant and powerful antioxidant.…”

“…We therefore focused on whether the intracellular redox status in general or GSH levels in particular could impact the transfection effectiveness of SS14 lipoplexes. As GSH is homeostatically controlled, being continually self-adjusting with respect to the balance between GSH synthesis, its recycling from GSSG, and its utilization, one of the most compelling lines of evidence for a role for GSH in gene delivery took advantage of the use of exogenous GSH depletors to temporarily decrease its intracellular concentration [7,8]. We therefore challenged MG63 cells with L-buthionine-sulfoximine (BSO) for 20 h and, as expected, found markedly high oxidative stress due to progressive GSH depletion (not shown).…”

“…Thus transfectants specifically tailored with disulfide functionalities turned out to be stable extracellularly but have proved amenable to reduction by intracellular sulfhydryl pool [6]. Despite a strong interdependence between intracellular glutathione (GSH) levels and the extent of transfection for bioreducible gene delivery vectors [7,8], alternative routes independent of GSH have also been elucidated [9]. In light of these somewhat puzzling results, it is not possible to dissect precisely the relative contribution of each of these mechanisms to the overall outcome of lipofection.…”