In Vitro and In Vivo Antimetastatic Effect of Glutathione Disulfide Liposomes

Abstract

Cancer metastasis is the major cause of cancer mortality. Despite extensive research efforts, effective treatment for cancer metastasis is still lacking. Cancer metastasis involves 4 essential steps: cell detachment, migration, invasion, and adhesion. Detachment is the first and required step for metastasis. Glutathione disulfide (GSSG) is derived from the oxidation of glutathione (GSH), which is present in biological systems in millimolar concentration. Although GSSG is commercially available, the impact of GSSG on cell functions/dysfunctions has not been fully explored due to the fact that GSSG is not cell membrane permeable and a lack of method to specifically increase GSSG in cells. We have developed GSSG liposomes that effectively deliver GSSG to cells. Unexpectedly, cells treated with GSSG liposomes were resistant to detachment by trypsinization. This observation led to the investigation of the antimetastatic effect of GSSG liposomes. Our data demonstrate that GSSG liposomes at 1 mg/mL completely blocked cell detachment and migration, and significantly inhibited cancer cell invasion. Aqueous GSSG showed no such effect, confirming that the effects on cell detachment, migration, and invasion were caused by the intracellular delivery of GSSG. An in vivo experiment with a murine melanoma experimental metastasis model showed that GSSG liposomes prevented melanoma lung metastasis. The unique antimetastatic mechanism through the effects on detachment and migration, and effective in vitro and in vivo metastasis inhibition, warrants further investigation of the GSSG liposomes as a potential treatment for cancer metastasis.

In Vitro and In Vivo Tumor Growth Inhibition by Glutathione Disulfide Liposomes

“…β-Actin in the figure was initially used as a loading control but found to be downregulated in cells treated with GSSG liposomes. 3 Downregulation of β-actin was consistent with the observation that GSSG liposomes affected cell migration and invasion through affecting cell motility as discussed in a separate study. 3 …”

“…The reason that cells had to be collected by a scraper instead of trypsinization is because cells treated with GSSG liposomes would not detach by trypsinization. 3 As shown in Figure 3, the number of cells treated with GSSG liposomes remained almost constant over the 72-hour period for both B16-F10 and NCI-H226 cells when compared with the control in which cells were treated with the growth medium containing PBS (control 1). In the meantime, cells treated with BLS (control 2) and GAQ (control 3) exhibited the same growth rates as those in control 1, confirming that it was GSSG being delivered into cells that produced a halt in cell proliferation (Figure 3).…”

“…In addition, in a separate study, GSSG liposomes were found to effectively prevent cancer cell detachment and migration, significantly inhibit invasion of 3 human cancer cell lines (NCI-H226, PC-3: prostate cancer, and HCT 116: colon cancer) and 1 murine melanoma cancer cell line (B16-F10), and exhibit significant antimetastatic effect in an experimental murine melanoma metastasis model. 3 The effective cancer proliferation retardation and strong metastasis blocking effect warrant GSSG liposomes for further investigation as a therapeutic agent for cancer treatment.…”

“…4,5 Consistently, an investigation with an in vivo murine melanoma metastasis model demonstrated that GSSG liposomes at 0.48 g/kg (a comparable concentration that produced the in vitro prevention of cell detachment and migration) produced almost a complete inhibition of melanoma metastasis. 3 …”

Controlled drug delivery vehicles for cancer treatment and their performance

“…Glutathione protects biological systems from oxidizing factors, such as reactive oxygen species, by terminating them; GSH is oxidized to GSSG, and then it is reduced back to GSH by glutathione reductase (GR). The unique antimetastatic mechanism of glutathione disulfide (GSSG) based liposomes completely prevents cell detachment and migration and significantly inhibits cancer cell invasion and has been confirmed as a potential treatment for cancer metastasis 154 . Temperature-sensitive liposomes with temperature-sensitive amphiphilic polymer poly(EOEOVE)-OD4 are used for tumor-specific chemotherapy.…”