Hyaluronic acid-bearing lipoplexes: Physico-chemical characterization and in vitro targeting of the CD44 receptor

Abstract

The mechanism by which hyaluronic acid (HA)-bearing lipoplexes target the A549 lung cancer cell line was evaluated. For this purpose, cationic liposomes targeting the CD44 receptor were designed thanks to the incorporation in their composition of a conjugate between high molecular weight HA and the lipid DOPE (HA-DOPE). Liposomes containing HA-DOPE were complexed at different lipids:DNA ratios with a reporter plasmid encoding the green fluorescent protein (GFP). Diameter, zeta potential, lipoplex stability and DNA protection from nucleases have been determined.Lipids:DNA ratios of 2, 4 and 6 provided a diameter around 250 nm with a zeta potential of − 30 mV.The strength of lipids:DNA interaction and the fraction of DNA protected from enzymatic degradation increased with the lipids:DNA ratio. 2D-immunoelectrophoresis demonstrated the low capacity to activate the C3 fraction of the complement system of any of these three ratios, with and without HA-DOPE. Transfection efficiency in the presence of 0, 10 and 15% of HA-DOPE or unconjugated HA, was determined on the CD44-expressing A549 cells by flow cytometry.Lipoplexes at a lipids:DNA ratio of 2 containing 10% (w/w) of HA-DOPE were the most efficient for transfection. The maximal level of GFP expression was obtained after 6 h of incubation demonstrating a slow transfection kinetics of lipoplexes. Finally, lipoplex cellular uptake, measured indirectly by the level of transfection using flow cytometry and validated by fluorescence microscopy, was shown to be mediated by the CD44 receptor and caveolae. These results demonstrate the strong specificity of DNA targeting through the CD44 receptor using HA of high molecular weight as a ligand.

Supramolecular Organization and siRNA Binding of Hyaluronic Acid-Coated Lipoplexes for Targeted Delivery to the CD44 Receptor

“…This amount was previously shown to be an optimal amount for the transfection of MDA-MB231 and A549 cancer cells expressing CD44. (18,25) The complexation of liposomes with siRNA at low concentrations (± ratios of 200, 100, and 50) resulted in lipoplexes with diameters comparable to those of liposomes, around 90 nm for the noncoated and 120 nm for the HA-liposomes (Figure 2). When larger amounts of siRNA were complexed (at ratios 4, 3, 2, and 1), the diameters of noncoated and HA-liposome increased to around 140 and 230 nm, respectively (Figure 2).…”

“…(15) It does not induce an expression of genes involved in proliferation or inflammation (16) and counteracts proangiogenic effects of the HA oligomers. (17,18) Surface modification of cationic liposomes with high-molecularweight HA can improve their efficacy by mediating active CD44 targeting in tumors and can also increase their circulation time because of a possible dysopsonization effect due to the hydrophilic coating effect of HA. (19)(20)(21)(22) In this study, we describe the design and physicochemical characterization of novel HA-lipoplexes entrapping siRNA.…”

Progress in the development of Lipoplex and Polyplex modified with Anionic Polymer for efficient Gene Delivery

“…HA can also be conjugated to lipids ( Figure 3B) or polymers (Figure 5B) using chemical linkers [36][37][38][39][40][41]. HA-DOPE conjugate can be obtained through the creation of an amide bond between the carboxylic groups of HA and the amino group of DOPE [37].…”

“…HA-DOPE conjugate can be obtained through the creation of an amide bond between the carboxylic groups of HA and the amino group of DOPE [37]. Inclusion of HA-DOPE conjugates into cationic liposomes composed of [2-(2,3-didodecyloxypropyl) hydroxyethyl] ammonium bromide (DC) and DOPE ( Figure 3B) could improve transfection into tumor cells expressing the CD44 receptor [36][37][38][39]. Furthermore, DOTAP/DOPE liposomes modi ied with HA-DOPE conjugate ( Figure 3B) also exhibited improved stability in cell culture medium and a reduced cytotoxicity [40].…”

Hyaluronan drug delivery systems are promising for cancer therapy because of their selective attachment, enhanced uptake, and superior efficacy

“…The addition of a delivery system will result in localization that reduces the concentration of the drug needed to achieve therapeutic outcomes, and have the added benefit of reducing the patients' side effects. Although many delivery systems and targeting molecules exist, hyaluronan is an interesting molecule because this polymer can be derivatized to conjugate chemotherapeutics drugs [16] and/or fabricated into many types of delivery systems including liposomes [17], nanoparticles [18], and pendant-chain systems [16]. Controlling the chemical reaction is a key for preserving the ability to bind hyaluronan receptors [16], which are overexpressed by many carcinomas [3].…”