Fliposomes: pH-triggered conformational flip of newtrans-2-aminocyclohexanol-based amphiphiles causes instant cargo release in liposomes

Abstract

A new type of pH-sensitive liposomes (fliposomes) was designed based on the amphiphiles that are able to perform a pH-triggered conformational flip (flipids). This flip disrupts the liposome membrane and causes rapid release of the liposome cargo, specifically in response to lowered pH. The flipids (1) and (2) are equipped with a trans-2-aminocyclohexanol conformational switch. pH-sensitive fliposomes containing one or both of these flipids, as well as POPC and PEG ceramide, were constructed and characterized. These compositions were stable at 4°C and pH 7.4 for several months. Fliposomes loaded with ANTS/DPX performed an unusually quick content release within a few seconds at pH below 8.5 (in case of 2) and 6.0 (in case of 1). This difference in pH sensitivity demonstrates a potential for the custom design of flipids by variation of the amino group to target areas with specific pH values. The pH titration curves for the fliposome leakage parallel the curves for the acid-induced conformational flip of 1 and 2 studied by ¹H NMR. A plausible mechanism of pH sensitivity starts with an acid-triggered conformational flip of 1 or 2, which changes the molecular size and shape, shortens the lipid tails, and perturbs the liposome membrane, resulting in the content leakage.

trans-2-Aminocyclohexanol-based amphiphiles as highly efficient helper lipids for gene delivery by lipoplexes

“…1) [21][22][23][24]. All the lipids (1-8) share the common structural features of a trans-2-aminocyclohexanol (TACH) moiety as the polar headgroup and two trans-alkoxycarbonyl groups at the 4-and 5-positions of the cyclohexane ring as the lipid tails.…”

“…The spin-spin coupling constants between several pairs of vicinal protons attached to the cyclohexane moiety are strongly conformationdependent, which allowed the assignment of a predominant conformation and the estimation of a position of the conformational equilibrium as described previously [21][22][23][24]31,32]. To characterize the pH-induced change of conformational equilibrium, a TACH-lipid was dissolved in CD 3 OD (0.02-0.03 M), and the changes of its 1 H NMR spectrum (600 MHz) were monitored during titration of the solution with d-trifluoroacetic acid (d-TFA) and with a base: 1,8-diazabicyclo [5.4.0]undec-7-ene (DBU).…”

“…The amphiphiles 1-3, 6, and 9, and the corresponding intermediates were prepared as described earlier [21][22][23][24]. Other TACH-lipids ( Fig.…”

“…This impulse is mechanically transmitted via the conformational flip of the cyclohexane ring to force both ester groups into axial positions (BH + on Scheme 1), thereby drastically increasing the spatial separation of the lipid tails and perturbing the bilayer structure. After incorporation into liposome membranes, TACH-lipids significantly enhanced the release from both the traditional liposomes based on phospholipids and the sterically hindered liposomes containing mPEG2000-ceramide (PEG-ceramide) upon exposure to lowered pH [21][22][23][24]. To highlight the key role of the conformational flip in the pHtriggered liposome leakage, we introduced the terms "flipids" for the amphiphiles containing a pH-sensitive conformational switch and "fliposomes" for the liposomes comprising these amphiphiles [22][23][24].…”

“…We recently designed a novel type of amphiphiles based on trans-2-aminocyclohexanol (TACH) and introduced these TACH-lipids as a pHsensitive conformational switch of liposome bilayer [21][22][23][24] (Scheme 1). In the original conformational equilibrium A ⇆ B (Scheme 1), the amine and the hydroxy groups of TACH-lipid are predominantly in axial positions while both lipidic trans-ester groups adopt equatorial positions (A).…”