The Polar Lipid Fraction E from Sulfolobus acidocaldarius Can Be Used as Liposomal Drug Stabilizing Agents to Reduce the Leakage of the Antivascular Drug Combretastatin A4 Disodium Phosphate from Tetraether/Diester Hybrid Archaeosomes

Abstract

Liposomes have many advantages as therapeutic capsules over free drugs such as small molecule drugs and nucleic acids. Cholesterol is commonly used as a membrane stabilizing agent in liposomal drugs (e.g., mRNA-lipid nanoparticle COVID-19 vaccines). However, due to the vulnerability of cholesterol to oxidation and the etiological role of cholesterol in many disorders, it is desirable to find an alternative means to stabilize liposomal membranes for drug delivery. In this study, we demonstrated that the polar lipid fraction E (PLFE), which contains exclusively bipolar tetraether macrocyclic lipids, isolated from the thermoacidophilic archaeon S. acidocaldarius can greatly stabilize the liposomal formulation of the anti-vascular drug, combretastatin A4 disodium phosphate (CA4P). Stability was assessed by determining the leakage rate constant k of entrapped CA4P fluorometrically. We found that, at 37 °C, PLFE decreases the k value monotonically from 1.54 × 10−2 s−1 for 100% 1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC) liposomes to 3.4 × 10−5 s−1 for 100% PLFE archaeosomes, a change of k by two orders of magnitude. The changes in k of CA4P leakage are correlated well with the changes in liposomal CA4P’s cytotoxicity against MCF-7 breast cancer cells. We further showed that the reduction in spontaneous leakage of entrapped CA4P by PLFE can be attributed to the increased membrane surface charge and the increased membrane order and packing tightness in liposomes, as reflected by the zeta potential (−6.83 to −41.1 mV from 0 to 100 mol% PLFE) and diphenylhexatriene (DPH) fluorescence polarization (0.13 to 0.4 from 0 to 100 mol% PLFE) measurements. Moreover, we showed that PLFE slows down CA4P leakage more than cholesterol in POPC liposomes. These results together suggest that PLFE lipids can serve as an effective stabilizing agent for liposomal drugs and could potentially be useful for the optimization of liposomal CA4P for cancer treatment.

Vesicular and Planar Membranes of Archaea Lipids: Unusual Physical Properties and Biomedical Applications

“…Physical properties of PLFE tetraether archaeosomes have been studied extensively (reviewed in [ 3 , 35 ]) and have been utilized to make archaeosomal therapeutics [ 16 , 26 , 36 , 37 , 38 , 39 ]. In terms of therapeutics delivery, archaeosomes are favored over conventional liposomes mainly because archaeosomes, particularly those containing tetraether lipids, are of much higher stability in many different aspects.…”

“…Membrane volume fluctuations are also tightly associated with solute permeability [ 47 ]. Low membrane volume fluctuation explains why the spontaneous leakage rate constant of dyes or drugs entrapped inside tetraether archaeosomes is significantly lower than inside conventional liposomes [ 27 , 37 , 44 ]. The leakage rate constant of the anti-vasculature drug combretastatin A4 disodium phosphate (CA4P) entrapped in PLFE archaeosomes is two orders of magnitude lower than that in 1-palmitoyl-2-oleoyl- sn -glycero-3-phosphocholine (POPC) diester liposomes [ 37 ].…”

“…Like cholesterol, PLFE and GCTE can stabilize liposomal membranes [ 19 , 37 ]. To evaluate whether PLFE or cholesterol is a better liposome stabilizing agent, a parallel comparison was made by using the same drug (CA4P) leakage and cytotoxicity (CyQuan) assays and it was found that the CA4P leakage rate constant in 20 mol% PLFE/80 mol% POPC [ 37 ] is significantly smaller than that in 20 mol% cholesterol/80 mol% POPC [ 57 ]. Apparently, PLFE is more efficient than cholesterol in keeping liposomal drugs.…”

“…As a stabilizing agent for liposomal drugs, PLFE tetraether lipids have advantages over cholesterol [ 37 ]. First, the amount of cholesterol in a liposome is restricted by the cholesterol solubility limit, which is 66.7 mol% for phosphatidylcholine liposomes [ 58 ].…”

“…Third, although cholesterol can tighten membrane packing, cholesterol does not change vesicle zeta potential and, thus, does not increase liposome stability against colloidal aggregation/coalescence. In sharp contrast, the zeta potential of PLFE/POPC liposomes can be changed from slightly negative to much more negative [ 37 ]. When the zeta potential becomes <−30 mV, the colloidal particles are usually considered as stable against particle aggregation and coalescence during long-term storage.…”