Enhanced cellular uptake of maleimide-modified liposomes via thiol-mediated transport

Abstract

With a small amount of maleimide modification on the liposome surface, enhanced cellular uptake of liposomes and drug-delivery efficiency can be obtained both in vitro and in vivo. Herein, we describe the mechanisms underlying this enhanced cellular uptake. Suppression of the cellular uptake of maleimide-modified liposomes (M-GGLG, composed of 1,5-dihexadecyl N,N -diglutamyl-lysyl-L-glutamate [GGLG]/cholesterol/poly(ethylene glycol) – 1,2-distearoyl- sn -glycero-3-phosphoethanolamine [PEG 5000 -DSPE]/maleimide [M]-PEG 5000 -Glu2C 18 at a molar ratio of 5:5:0.03:0.03) caused by temperature block and addition of serum was alleviated compared with that of liposomes without maleimide modification (GGLG liposomes, composed of GGLG/cholesterol/PEG 5000 -DSPE/PEG 5000 -Glu2C 18 at a molar ratio of 5:5:0.03:0.03). When 0.01 nM N -ethylmaleimide was used to pre-block cellular thiols, the cellular uptake of M-GGLG liposomes was decreased to approximately 70% in HeLa, HCC1954, MDA-MB-468, and COS-7 cell lines. Moreover, inhibition of a thiol-related reductase such as protein disulfide isomerase resulted in a 15%–45% inhibition of the cellular uptake of M-GGLG liposomes, whereas GGLG liposomes were not influenced. Further, single and mixed inhibitors of clathrin-mediated endocytosis, caveolae-mediated endocytosis, and macropinocytosis did not efficiently inhibit the cellular uptake of M-GGLG liposomes. Using confocal microscopy, we verified that M-GGLG liposomes were localized partially in lysosomes after inhibition of the mentioned conventional endocytic pathways. Therefore, it was hypothesized that the mechanisms underlying the enhanced cellular uptake of liposomes by maleimide modification was thiol-mediated membrane trafficking, including endocytosis and energy-independent transport.

Epidithiodiketopiperazines: Strain-Promoted Thiol-Mediated Cellular Uptake at the Highest Tension

“…Similarly, the presence of 10% serum 22 caused only a minor ∼25% reduction of ETP uptake (Figures 4C and S5). DTNB 25 was special because this reagent converts thiols on cell surfaces into activated disulfides.…”

“…Contrary to AspA controls, 12,13 the removal of thiols on cell surfaces with maleimide 26 , 22 iodoacetamide 27 , 12 and the most powerful hypervalent iodine reagent 28 (29) failed to inactivate ETP 6 (Figure 4C). Similarly, the presence of 10% serum 22 caused only a minor ∼25% reduction of ETP uptake (Figures 4C and S5).…”

“…Namely, insensitivity toward chlorpromazine (CPZ) excluded clathrin-mediated endocytosis, methyl-β-cyclodextrin (mβCD) caveolae-mediated endocytosis, and wortmannin and cytochalasin B (cytoB) ruled out macropinocytosis (Figure 4B). 11,22,26−28 …”

Effects of Liposomes Charge on Extending Sciatic Nerve Blockade of N-ethyl Bromide of Lidocaine in Rats

“…It is well known that liposomes have good affinity to cell membranes and lipid tissues71819. Previously, hydrocortisone or insulin administered with liposomes resulted in targeted delivery through skins20.…”

Improving cellular uptake of therapeutic entities through interaction with components of cell membrane

“…reported that thiol-reactive small molecules, Alexa Fluor 488-C5-maleimide dye, can be conjugated to plasma membrane thiol groups in the absence of endocytosis at 4 °C (Fretz et al., 2007). Moreover, when liposomes were modified with a small amount of maleimide, the cellular uptake and drug-delivery efficiency were significantly enhanced (Li & Takeoka, 2014). The studies of the mechanism revealed that the interaction of maleimide with cellular thiols triggered alternative cellular internalization via thiol-mediated transport.…”