Cationic DOPC–Detergent Conjugates for Safe and Efficient in Vitro and in Vivo Nucleic Acid Delivery

Abstract

The ability of a nonviral nucleic acid carrier to deliver its cargo to cells with low associated toxicity is a critical issue for clinical applications of gene therapy. We describe biodegradable cationic DOPC-C12 E4 conjugates in which transfection efficiency is based on a Trojan horse strategy. In situ production of the detergent compound C12 E4 through conjugate hydrolysis within the acidic endosome compartment was expected to promote endosome membrane destabilization and subsequent release of the lipoplexes into cytosol. The transfection efficiency of the conjugates has been assessed in vitro, and associated cytotoxicity was determined. Cellular uptake and intracellular distribution of the lipoplexes have been investigated. The results show that direct conjugation of DOPC with C12 E4 produces a versatile carrier that can deliver both DNA and siRNA to cells in vitro with high efficiency and low cytotoxicity. SAR studies suggest that this compound might represent a reasonable compromise between the membrane activity of the released detergent and susceptibility of the conjugate to degradation enzymes in vitro. Although biodegradability of the conjugates had low impact on carrier efficiency in vitro, it proved critical in vivo. Significant improvement of transgene expression was obtained in the mouse lung tuning biodegradability of the carrier. Importantly, this also allowed reduction of the inflammatory response that invariably characterizes cationic-lipid-mediated gene transfer in animals.

Enhanced gene delivery to the lung using biodegradable polyunsaturated cationic phosphatidylcholine-detergent conjugates

“…Preparation of the cationic conjugates 1 and 6 has been described elsewhere (Pierrat et al, 2016a;Pierrat et al, 2016b). GL67 and formulation GL67A were prepared as reported in the literature (Chadwick et al, 1997;Griesenbach et al, 2011b).…”

“…Optimum conditions for in vitro DNA delivery with cationic lipid 1 to various cell lines were obtained at charge neutrality (N/P = 1) and without the need for addition of helper lipid (e.g. 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, DOPE) to improve transfection efficiency or decrease cytotoxicity (Pierrat et al, 2016a). Furthermore, the introduction of PEGylated lipids in the formulations led to significant loss in transfection efficiency in vitro (Fig.…”

“…The first one entails intrinsic fusogenicity of the conjugates (Pierrat et al, 2016b). The second one implies a Trojan horse mechanism, where potency of the carriers relies on their biodegradability (Pierrat et al, 2013a;Pierrat et al, 2013b) and on the membrane disrupting properties of the detergent molecules produced upon hydrolysis of the conjugates within the endosome compartment (Pierrat et al, 2016a). These two mechanisms rely on separate properties of the conjugates in the extracellular and intracellular compartments and are not mutually exclusive.…”

Dual Gene Delivery Reagents From Antiproliferative Alkylphospholipids for Combined Antitumor Therapy

“…In the course of our quest for nucleic acid carriers with improved transfection properties and safeness, we previously developed cationic derivatives of 1,2-dioleoyl- sn -glycero-3-phosphatidylcholine (DOPC), a major component of the cell membranes. These phosphotriester compounds proved highly efficient nucleic acid delivery reagents both in vitro (Pierrat et al, 2012 , 2013a , b ) and in vivo (Pierrat et al, 2016a , b , c ). This inspired us to develop erufosine-based biolabile phosphotriesters (pro-APLs) as dual gene delivery reagents for combined cancer therapy (Gaillard et al, 2019 ).…”

Tuning the cationic interface of simple polydiacetylene micelles to improve siRNA delivery at the cellular level

“… 4 The design of nanocarriers is thus of prime importance to maximize the amount of siRNA to be delivered into the cells. At the cellular level, examples of siRNA delivery systems include cationic polymers, 5 lipids, 6 peptides, 7 carbon nanotubes, 8 nanofibers 9 and micelles. 10 Although the latter systems are efficient and could behave in a synergistic fashion, 11 there is a lack of understanding on how the chemical structures of the cationic region (responsible for electrostatic interactions with the nucleic acids) impacts the overall transfection process, and a rationale is yet to be formulated.…”