Drug Release by Direct Jump from Poly(ethylene-glycol-b-ε-caprolactone) Nano-Vector to Cell Membrane.

Drug delivery by nanovectors involves numerous processes, one of the most important being its release from the carrier. This point still remains unclear. The current work focuses on this point using poly(ethyleneglycol-b-ε-caprolactone) micelles containing either pheophorbide-a (Pheo-a) as a fluorescent probe and a phototoxic agent or fluorescent copolymers. This study showed that the cellular uptake and the phototoxicity of loaded Pheo-a are ten times higher than those of the free drug and revealed a very low cellular penetration of the fluorescence-labeled micelles. Neither loaded nor free Pheo-a displayed the same cellular localization as the labeled micelles. These results imply that the drug entered the cells without its carrier and probably without a disruption, as suggested by their stability in cell culture medium. These data allowed us to propose that Pheo-a directly migrates from the micelle to the cell without disruption of the vector. This mechanism will be discussed.

Randomised comparison of a bioresorbable everolimus-eluting scaffold with a metallic everolimus-eluting stent for ischaemic heart disease caused by de novo native coronary artery lesions: the 2-year clinical outcomes of the ABSORB II trial.

AIMS: The one-year randomised data of the ABSORB II trial showed that the everolimus-eluting bioresorbable scaffold and the everolimus-eluting metallic stent were comparable for the composite secondary clinical outcomes of patient-oriented composite endpoint (PoCE) and device-oriented composite endpoint (DoCE)/target lesion failure (TLF), MACE and TVF. This report describes the two-year clinical outcomes of the ABSORB II trial. METHODS AND RESULTS: Patients were randomly assigned in a 2:1 ratio to receive treatment with an everolimus-eluting bioresorbable scaffold (Absorb; Abbott Vascular, Santa Clara, CA, USA) or treatment with an everolimus-eluting metallic stent (XIENCE; Abbott Vascular). The trial enrolled 501 patients. Clinical follow-up at two years was available in 320 patients in the Absorb BVS arm and 160 patients in the XIENCE arm. At two years, the PoCE for the Absorb and XIENCE arms was 11.6% and 12.8% (p=0.70) and the DoCE/TLF was 7.0% and 3.0% (p=0.07), respectively. The hierarchical ID-MACE rate was 7.6% vs. 4.3% (p=0.16) and the rate of TVF was 8.5% vs. 6.7% (p=0.48). The definite/probable thrombosis rate was 1.5% in the Absorb arm vs. 0% in the XIENCE arm (p=0.17). Thirty-six percent and 34% of patients remained on DAPT at two years, respectively. Ninety-two percent of patients in both arms remained on aspirin. CONCLUSIONS: Two-year clinical results demonstrate sustained low rates of PoCE, MACE, DoCE and TVF with the Absorb BVS as compared to the XIENCE stent.

A Polylactide Bioresorbable Scaffold Eluting Everolimus for Treatment of Coronary Stenosis: 5-Year Follow-Up.

BACKGROUND: Long-term benefits of coronary stenosis treatment with an everolimus-eluting bioresorbable scaffold are unknown. OBJECTIVES: This study sought to evaluate clinical and imaging outcomes 5 years after bioresorbable scaffold implantation. METHODS: In the ABSORB multicenter, single-arm trial, 45 (B1) and 56 patients (B2) underwent coronary angiography, intravascular ultrasound (IVUS), and optical coherence tomography (OCT) at different times. At 5 years, 53 patients without target lesion revascularization underwent final imaging. RESULTS: Between 6 months/1 year and 5 years, angiographic luminal late loss remained unchanged (B1: 0.14 ± 19 mm vs. 0.13 ± 0.33 mm; p = 0.7953; B2: 0.23 ± 0.28 mm vs. 0.18 ± 0.32 mm; p = 0.5685). When patients with a target lesion revascularization were included, luminal late loss was 0.15 ± 0.20 mm versus 0.15 ± 0.24 mm (p = 0.8275) for B1 and 0.30 ± 0.37 mm versus 0.32 ± 0.48 mm (p = 0.8204) for B2. At 5 years, in-scaffold and -segment binary restenosis was 7.8% (5 of 64) and 12.5% (8 of 64). On IVUS, the minimum lumen area of B1 decreased from 5.23 ± 0.97 mm(2) at 6 months to 4.89 ± 1.81 mm(2) at 5 years (p = 0.04), but remained unchanged in B2 (4.95 ± 0.91 mm(2) at 1 year to 4.84 ± 1.28 mm(2) at 5 years; p = 0.5). At 5 years, struts were no longer discernable by OCT and IVUS. On OCT, the minimum lumen area in B1 decreased from 4.51 ± 1.28 mm(2) at 6 months to 3.65 ± 1.39 mm(2) at 5 years (p = 0.01), but remained unchanged in B2, 4.35 ± 1.09 mm(2) at 1 year and 4.12 ± 1.38 mm(2) at 5 years (p = 0.24). Overall, the 5-year major adverse cardiac event rate was 11.0%, without any scaffold thrombosis. CONCLUSIONS: At 5 years, bioresorbable scaffold implantation in a simple stenotic lesion resulted in stable lumen dimensions and low restenosis and major adverse cardiac event rates. (ABSORB Clinical Investigation, Cohort B [ABSORB B]; NCT00856856).

Incidence and Potential Mechanism(s) of Post-Procedural Rise of Cardiac Biomarker in Patients With Coronary Artery Narrowing After Implantation of an Everolimus-Eluting Bioresorbable Vascular Scaffold or Everolimus-Eluting Metallic Stent.

OBJECTIVES: This study sought to evaluate the mechanism of post-procedural cardiac biomarker (CB) rise following device implantation. BACKGROUND: A fully bioresorbable Absorb scaffold, compared with everolimus-eluting metallic stents (EES), might be associated with a higher incidence of periprocedural myocardial injury. METHODS: In 501 patients with stable or unstable angina randomized to either Absorb (335 patients) or EES (n = 166) in the ABSORB II trial, 3 types of CB (creatine kinase, creatine kinase-myocardial band, and troponin) were obtained before and after procedure. Per protocol, periprocedural myocardial infarction (PMI) was defined as creatine kinase rise >2× the upper limit of normal with creatine kinase-myocardial band rise. RESULTS: Incidence of side branch occlusion and any anatomic complications assessed by angiography was similar between the 2 treatment arms (side branch occlusion: Absorb: 5.3% vs. Xience: 7.6%, p = 0.07; any anatomic complication: Absorb: 16.4% vs. EES: 19.9%, p = 0.39). Fourteen patients who presented with recent myocardial infarction at entry with normalized creatine kinase-myocardial band according to the protocol were excluded for post-CB analysis. The overall compliance for CB was 97.8%. The CB rise subcategorized in 7 different ranges was comparable between the 2 treatment arms. PMI rate was numerically higher in the Absorb arm according to the per-protocol definitions, and treatment with overlapping devices was the only independent determinant of per-protocol PMI (odds ratio: 5.07, 95% confidence interval: 1.78 to 14.41, p = 0.002). CONCLUSIONS: There were no differences in the incidence of CB rise and PMI between Absorb and EES. Device overlap might be a precipitating factor of myocardial injury. (ABSORB II Randomized Clinical Trial: A Clinical Evaluation to Compare the Safety, Efficacy, and Performance of Absorb Everolimus Eluting Bioresorbable Vascular Scaffold System Against Xience Everolimus Eluting Coronary Stent System in the Treatment of Subjects With Ischemic Heart Disease Caused by De Novo Native Coronary Artery Lesions [ABSORB II]; NCT01425281).

A bioresorbable everolimus-eluting scaffold versus a metallic everolimus-eluting stent for ischaemic heart disease caused by de-novo native coronary artery lesions (ABSORB II): an interim 1-year analysis of clinical and procedural secondary outcomes from a randomised controlled trial.

BACKGROUND: Despite rapid dissemination of an everolimus-eluting bioresorbable scaffold for treatment for coronary artery disease, no data from comparisons with its metallic stent counterpart are available. In a randomised controlled trial we aimed to compare an everolimus-eluting bioresorbable scaffold with an everolimus-eluting metallic stent. Here we report secondary clinical and procedural outcomes after 1 year of follow-up. METHODS: In a single-blind, multicentre, randomised trial, we enrolled eligible patients aged 18-85 years with evidence of myocardial ischaemia and one or two de-novo native lesions in different epicardial vessels. We randomly assigned patients in a 2:1 ratio to receive treatment with an everolimus-eluting bioresorbable scaffold (Absorb, Abbott Vascular, Santa Clara, CA, USA) or treatment with an everolimus-eluting metallic stent (Xience, Abbott Vascular, Santa Clara, CA, USA). Randomisation was stratified by diabetes status and number of planned target lesions. The co-primary endpoints of this study are vasomotion (change in mean lumen diameter before and after nitrate administration at 3 years) and difference between minimum lumen diameter (after nitrate administration) after the index procedure and at 3 years. Secondary endpoints were procedural performance assessed by quantitative angiography and intravascular ultrasound; composite clinical endpoints based on death, myocardial infarction, and coronary revascularisation; device and procedural success; and angina status assessed by the Seattle Angina Questionnaire and exercise testing at 6 and 12 months. Cumulative angina rate based on adverse event reporting was analysed post hoc. This trial is registered at ClinicalTrials.gov, number NCT01425281. FINDINGS: Between Nov 28, 2011, and June 4, 2013, we enrolled 501 patients and randomly assigned them to the bioresorbable scaffold group (335 patients, 364 lesions) or the metallic stent group (166 patients, 182 lesions). Dilatation pressure and balloon diameter at the highest pressure during implantation or postdilatation were higher and larger in the metallic stent group, whereas the acute recoil post implantation was similar (0.19 mm for both, p=0.85). Acute lumen gain was lower for the bioresorbable scaffold by quantitative coronary angiography (1.15 mm vs 1.46 mm, p<0.0001) and quantitative intravascular ultrasound (2.85 mm(2)vs 3.60 mm(2), p<0.0001), resulting in a smaller lumen diameter or area post procedure. At 1 year, however, cumulative rates of first new or worsening angina from adverse event reporting were lower (72 patients [22%] in the bioresorbable scaffold group vs 50 [30%] in the metallic stent group, p=0.04), whereas performance during maximum exercise and angina status by SAQ were similar. The 1-year composite device orientated endpoint was similar between the bioresorbable scaffold and metallic stent groups (16 patients [5%] vs five patients [3%], p=0.35). Three patients in the bioresorbable scaffold group had definite or probable scaffold thromboses (one definite acute, one definite sub-acute, and one probable late), compared with no patients in the metallic stent group. There were 17 (5%) major cardiac adverse events in the bioresorbable scaffold group compared with five (3%) events in the metallic stent group, with the most common adverse events being myocardial infarction (15 cases [4%] vs two cases [1%], respectively) and clinically indicated target-lesion revascularisation (four cases [1%] vs three cases [2%], respectively). INTERPRETATION: The everolimus-eluting bioresorbable scaffold showed similar 1-year composite secondary clinical outcomes to the everolimus-eluting metallic stent. FUNDING: Abbott Vascular.

Shock waves associated with electric pulses affect cell electro-permeabilization.

New features of cell electro-permeabilization are obtained by using high field (several tens of kV/cm) with short (sub-microsecond, nanosecond) pulse duration. Arcing appears as a main safety problem when air gaps are present between electrodes. A new applicator design was chosen to obtain a closed chamber where high field pulses could be delivered in a safe way with very short pulse duration. The safety issue of the system was validated under millisecond, microsecond and nanosecond pulses. The closed chamber applicator was then checked for its use under classical electro-mediated permeabilization and electro-gene transfer (EGT). A 20 times decrease in gene expression was observed compared with classical open chambers. It was experimentally observed that shock waves were present under the closed chamber configuration of the applicator. This was not the case with an open chamber design. Electropulsation chamber design plays a role on pulsing conditions and in the efficiency of gene electro transfer.

Antitumor drug delivery in multicellular spheroids by electropermeabilization.

Electrochemotherapy (ECT) is a physical technique that allows cytotoxic molecules to be efficiently released in tumor cells by inducing transient cell plasma membrane permeabilization. The main antitumoral drugs used in ECT are nonpermeant bleomycin and low permeant cisplatin. The method is nowadays applied in clinics as a palliative treatment. In order to improve it, we took advantage of a human 3D multicellular tumor spheroid as a model of tumor to visually and molecularly assess the effect of ECT. We used bleomycin and cisplatin to confirm its relevance and doxorubicin to show its potential to screen new antitumor drug candidates for ECT. Confocal microscopy was used to visualize the topological distribution of permeabilized cells in 3D spheroids subjected to electric pulses. Our results revealed that all cells were efficiently permeabilized, whatever their localization in the spheroid, even those in the core. The combination of antitumor drugs and electric pulses (ECT) led to changes in spheroid macroscopic morphology and cell cohesion, to tumor spheroid growth arrest and finally to its complete apoptosis-mediated dislocation, mimicking previously observed in vivo situations. Taken together, these results indicate that the spheroid model is relevant for the study and optimization of electromediated drug delivery protocols.

Investigating relationship between transfection and permeabilization by the electric field and/or the Pluronic® L64 in vitro and in vivo.

BACKGROUND: Electrotransfer can be obtained by the successive delivery of a high voltage short duration pulse (HV) inducing membrane destabilization and then a low voltage long duration pulse (LV), allowing DNA electrophoresis (HVLV mode). Pluronic® L64 (L64) (Fluka, Sigma-Aldrich, L'Isle-d'Abeau Chesnes, Saint-Quentin Fallavier, France) has permeabilizing properties and amplifies the expression of DNA. We aimed to determine whether L64 could have an adjuvant effect on transfection by electrotransfer and whether the sequence L64 injection and then application of a LV pulse could induce transfection comparable to that observed with the HVLV mode. METHODS: In vitro, we used fluorescence-activated cell sorting to evaluate Chinese hamster ovary (CHO) cell transfection by a plasmid coding green fluorescent protein, and permeabilization to propidium iodide. In vivo, the transfection efficiency of mice tibial cranial muscle was evaluated by optical imaging using a plasmid DNA encoding luciferase. For the same animals, permeabilization indices were evaluated by magnetic resonance imaging from the uptake of a T(1) contrast agent. RESULTS: Using the HVLV mode, transfection efficiency was low in vitro on CHO cells but high for muscles in vivo. Pre-treatment by L64 increased the transfection efficiency of electrotransfer for CHO cells but not for muscle. In mice muscles, the L64 amplified the expression of DNA. Nevertheless, neither transgene expression, nor permeability indices were further amplified by subsequent delivery of one LV pulse. CONCLUSIONS: A major finding of the present study is that the nature of the membrane modification induced by electric pulses is not comparable to that mediated by L64. The electrophoretic LV pulse does not induce additive effects to that of L64 for transfection improvement.

Cationic and anionic lipoplexes inhibit gene transfection by electroporation in vivo.

BACKGROUND: Nonviral gene therapy still suffers from low efficiency. Methods that would lead to higher gene expression level of longer duration would be a major advance in this field. Lipidic vectors and physical methods have been investigated separately, and both induced gene expression improvement. METHODS: We sought to combine both chemical and physical methods. Cationic or anionic lipids can potentially destabilize the cell membrane and could consequently enhance gene delivery by a physical method such as electrotransfer. A plasmid model encoding luciferase was used, either free or associated with differently-charged lipoplexes before electrotransfer. RESULTS: Electrotransfer alone strongly enhanced gene expression after intramuscular and intradermal injection of naked DNA. On the other hand, cationic and anionic lipoplex formulations decreased gene expression after electrotransfer, whereas poorly-charged thiourea-based complexes, brought no benefit. Pre-injection of the lipids, followed by administration of naked DNA, did not modified gene expression induced by electroporation in the skin. CONCLUSIONS: The results obtained in the present study suggest that packing of DNA plasmid in lipoplexes strongly decreases the efficiency of gene electrotransfer, independently of the lipoplex charge. Non-aggregating complexes, such as poorly-charged thiourea-based complexes, should be preferred to increase DNA release.

What is (still not) known of the mechanism by which electroporation mediates gene transfer and expression in cells and tissues.

Cell membranes can be transiently permeabilized under application of electric pulses. This treatment allows hydrophilic therapeutic molecules, such as anticancer drugs and DNA, to enter into cells and tissues. This process, called electropermeabilization or electroporation, has been rapidly developed over the last decade to deliver genes to tissues and organs, but there is a general agreement that very little is known about what is really occurring during membrane electropermeabilization. It is well accepted that the entry of small molecules, such as anticancer drugs, occurs mostly through simple diffusion after the pulse while the entry of macromolecules, such as DNA, occurs through a multistep mechanism involving the electrophoretically driven interaction of the DNA molecule with the destabilized membrane during the pulse and then its passage across the membrane. Therefore, successful DNA electrotransfer into cells depends not only on cell permeabilization but also on the way plasmid DNA interacts with the plasma membrane and, once into the cytoplasm, migrates towards the nucleus. The focus of this review is to describe the different aspects of what is known of the mechanism of membrane permeabilization and associated gene transfer and, by doing so, what are the actual limits of the DNA delivery into cells.

Gene electrotransfer: from biophysical mechanisms to in vivo applications : Part 2 - In vivo developments and present clinical applications.

Gene electrotransfer can be obtained not just on single cells in diluted suspension. For more than 10 years, this is a quasi routine strategy in tissue on the living animal and a few clinical trials have now been approved. New problems have been brought by the close contacts of cells in tissue both on the local field distribution and on the access of DNA to target cells. They need to be solved to provide a further improvement in the efficacy and safety of protein expression. There is a competition between gene transfer and cell destruction. Nevertheless, present results are indicative that electrotransfer is a promising approach for gene therapy. High level and long-lived expression of proteins can be obtained in muscles. This is used for a successful method of electrovaccination.

Gene electrotransfer: from biophysical mechanisms to in vivo applications : Part 1- Biophysical mechanisms.

Electropulsation is one of the nonviral methods successfully used to deliver genes into living cells in vitro and in vivo. This approach shows promise in the field of gene and cellular therapies. The present review focuses on the processes supporting gene electrotransfer in vitro. In the first part, we will report the events occurring before, during, and after pulse application in the specific field of plasmid DNA electrotransfer at the cell level. A critical discussion of the present theoretical considerations about membrane electropermeabilization and the transient structures involved in the plasmid uptake follows in a second part.

Cationic lipids, lipoplexes and intracellular delivery of genes.

As a consequence of several setbacks encountered by viral technology in achieving efficient and safe gene therapy in clinical trials, non-viral gene delivery vectors are considered to date as a valuable alternative and to hold promise for future therapeutic applications. Nevertheless, the transfection efficiency mediated by these non-viral gene delivery vectors has to be improved, especially in vivo, to benefit fully from their advantages. Cationic lipid/nucleic acid complexes or lipoplexes have been the subject of intensive investigations in recent years to understand the parameters governing the efficiency of transfection. Specifically, the comprehension of such mechanisms, from the formation of the complexes to their intracellular delivery, will lead to the design of better adapted non-viral vectors for gene therapy applications. Here, we will discuss some recent developments in the field on the structure/function relationship of cationic lipids in the mechanism of transfection, and where appropriate, we will make a comparison with mechanisms of viral and polyplex-mediated gene delivery. Cationic lipids are often used in combination with helper lipids such as DOPE or cholesterol. The effect of DOPE on lipoplex assembly and the relevance of the structural properties of the lipoplexes in destabilizing endosomal membranes and mediating endosomal escape of DNA will be discussed.

Lipoplexes formed from sugar-based gemini surfactants undergo a lamellar-to-micellar phase transition at acidic pH. Evidence for a non-inverted membrane-destabilizing hexagonal phase of lipoplexes.

The present study aims at a better understanding of the mechanism of transfection mediated by two sugar-based gemini surfactants GS1 and GS2. Previously, these gemini surfactants have been shown to be efficient gene vectors for transfection both in vitro and in vivo. Here, using Nile Red, a solvatochromic fluorescent probe, we investigated the phase behavior of these gemini surfactants in complexes with plasmid DNA, so-called lipoplexes. We found that these lipoplexes undergo a lamellar-to-non-inverted micellar phase transition upon decreasing the pH from neutral to mildly acidic. This normal (non-inverted) phase at acidic pH is confirmed by the colloidal stability of the lipoplexes as shown by turbidity measurements. We therefore propose a normal hexagonal phase, H(I), for the gemini surfactant lipoplexes at acidic endosomal pH. Thus, we suggest that besides an inverted hexagonal (H(II)) phase as reported for several transfection-potent cationic lipid systems, another type of non-inverted non-bilayer structure, different from H(II), may destabilize the endosomal membrane, necessary for cytosolic DNA delivery and ultimately, cellular transfection.

Transfection mediated by pH-sensitive sugar-based gemini surfactants; potential for in vivo gene therapy applications.

In this study, the in vitro and in vivo transfection capacity of novel pH-sensitive sugar-based gemini surfactants was investigated. In an aqueous environment at physiological pH, these compounds form bilayer vesicles, but they undergo a lamellar-to-micellar phase transition in the endosomal pH range as a consequence of an increased protonation state. In the same way, lipoplexes made with these amphiphiles exhibit a lamellar morphology at physiological pH and a non-lamellar phase at acidic pH. In this study, we confirm that the gemini surfactants are able to form complexes with plasmid DNA at physiological pH and are able to transfect efficiently CHO cells in vitro. Out of the five compounds tested here, two of these amphiphiles, GS1 and GS2, led to 70% of transfected cells with a good cell survival. These two compounds were tested further for in vivo applications. Because of their lamellar organisation, these lipoplexes exhibited a good colloidal stability in salt and in serum at physiological pH compatible with a prolonged stability in vivo. Indeed, when injected intravenously to mice, these stable lipoplexes apparently did not substantially accumulate, as inferred from the observation that transfection of the lungs was not detectable, as examined by in vivo bioluminescence. This potential of avoiding 'preliminary capture' in the lungs may, thus, be further exploited in developing devices for specific targeting of gemini lipoplexes.

Effect of manganese on in vitro replication of damaged DNA catalyzed by the herpes simplex virus type-1 DNA polymerase.

In vitro bypass of damaged DNA by replicative DNA polymerases is usually blocked by helix-distorting or bulky DNA lesions. In this study, we report that substitution of the divalent metal ion Mg2+ with Mn2+ promotes quantitative replication of model DNA substrates containing the major cisplatin or N-2-acetylaminofluorene adducts by the catalytic subunit (UL30) of the replicative DNA polymerase of herpes simplex virus. The ability of Mn2+ ions to confer bypass of bulky lesions was not observed with other replicative DNA polymerases of the B family, such as bacteriophage T4 or delta polymerases. However, for these enzymes, manganese induced the incorporation of one nucleotide opposite the first (3') guanine of the d(GpG) intrastrand cisplatin lesion. Translesion replication of the cisplatin adduct by UL30 led to the incorporation of mismatched bases, with the preferential incorporation of dAMP opposite the 3' guanine of the lesion. Furthermore, substitution of MgCl2 with MnCl2 greatly inhibited the 3' to 5' exonuclease of UL30 but had a far lesser effect on that of T4 DNA polymerase. Finally, manganese induced a conformational change in the structure of UL30 bound to the platinated substrate. Taken together, the latter findings suggest a mechanism by which manganese might allow UL30 to efficiently promote translesion DNA synthesis in vitro.

Interference of poly(ethylene glycol)-lipid analogues with cationic-lipid-mediated delivery of oligonucleotides; role of lipid exchangeability and non-lamellar transitions.

Cationic liposomes are applied to transfer oligonucleotides (ODNs) into cells to regulate gene expression for gene therapeutic or cell biological purposes. In vivo, poly(ethylene glycol) (PEG)-lipid derivatives are employed to stabilize and prolong the circulation lifetime of nucleic acid-containing particles, and to improve targeting strategies. In this study, we have studied the effects of PEG-lipid analogues, i.e. PEG coupled to either phosphatidylethanolamine (PE) or ceramide, on cationic-lipid-DNA complex ('lipoplex') assembly and the mechanism of cationic-lipid-mediated delivery of ODNs in vitro. Inclusion of 10 mol% PEG-PE in ODN lipoplexes inhibited their internalization in Chinese hamster ovary cells by more than 70%. The intracellular fraction remained entrapped in the endosomal/lysosomal pathway, and no release of ODNs was apparent. Similar observations were made for complexes prepared from liposomes that contained PEG-ceramides. Interestingly, delivery resumed when lipoplexes had been externally coated with PEG-ceramides. In this case, the kinetics of delivery were dependent on the length of the ceramide acyl chain, consistent with a requirement for the PEG-lipid to dissociate from the complex. Moreover, although the chemical nature of the PEG-ceramides distinctly affected the net internalization of the complexes, impediment of delivery was largely related to an inhibitory effect of the PEG-lipid on the release of ODNs from the endosomal compartment. Cryo-electron microscopy and small-angle X-ray scattering revealed that the PEG-lipids stabilize the lamellar phase of the lipoplexes, while their acyl-chain-length-dependent transfer from the complex enables adaptation of the hexagonal phase. Within the endosomal compartment, this transition appears to be instrumental in causing the dissociation and cytosolic release of the ODNs for their nuclear homing.