ABSTRACT
Ionizable lipids are a class of pharmaceutical excipients with a main application in lipid nanoparticles for nucleic acid delivery. New ionizable lipids are needed to tune characteristics of lipid-based nucleic acid delivery systems, e.g. stability, nucleic acid loading capacity and binding strength, as well as bio-distribution. Herein, we present the synthesis of three novel ionizable lipids as putative excipients for lipid-based nucleic acid delivery systems. Langmuir monolayer experiments with classical surface pressure/area isotherm evaluation were used to understand the self-assembly behavior of the lipids. Additional experiments with surface sensitive techniques, namely grazing incidence x-ray scattering and infrared reflection-absorption spectroscopy (IRRAS), were performed to understand structural characteristics of lipid associates. The latter technique was also used to investigate the nucleic acid binding process between DNA and the ionizable lipids. Finally, first transfection experiments with the novel lipids formulated as cationic liposomes were performed providing first efficacy data. Although the alkyl chain pattern was comparable for all three ionizable lipids, the results demonstrated that with increasing head-group size the DNA binding capacity changed and the alkyl chain fluidity was increased. The lipid with the lowest phase transition temperature and the smallest packing parameter showed the highest DNA transfer efficiency.
Subject(s)
DNA , Fatty Acids , Lipids , Lipids/chemistry , Fatty Acids/chemistry , DNA/chemistry , Liposomes/chemistry , Excipients/chemistry , Nanoparticles/chemistry , Surface PropertiesABSTRACT
The physicochemical properties and transfection efficacies of two samples of a cationic lipid have been investigated and compared in 2D (monolayers at the air/liquid interface) and 3D (aqueous bulk dispersions) model systems using different techniques. The samples differ only in their chain composition due to the purity of the oleylamine (chain precursor). Lipid 8 (using the oleylamine of technical grade for cost-efficient synthesis) shows lateral phase separation in the Langmuir layers. However, the amount of attached DNA, determined by IRRAS, is for both samples the same. In 3D systems, lipid 8 p forms cubic phases, which disappear after addition of DNA. At physiological temperatures, both lipids (alone and in mixture with cholesterol) assemble to lamellar aggregates and exhibit comparable DNA delivery efficiency. This study demonstrates that non-lamellar structures are not compulsory for high transfection rates. The results legitimate the utilization of oleyl chains of technical grade in the synthesis of cationic transfection lipids.
Subject(s)
Amines/chemistry , DNA/chemistry , Lipids/chemistry , Liposomes/chemistry , Amines/chemical synthesis , Amines/standards , Amines/toxicity , Animals , Cattle , Cell Line, Tumor , Cholesterol/chemistry , Gene Transfer Techniques/standards , Humans , Lipids/chemical synthesis , Lipids/standards , Lipids/toxicity , Liposomes/standards , Liposomes/toxicity , Molecular Structure , Phase Transition , Swine , Transfection/standards , Transition TemperatureABSTRACT
Based on previous work, the influence of the chain composition on the physical-chemical properties of five new transfection lipids (TH10, TT10, OH10, OT10 and OO10) containing the same lysine-based head group has been investigated in aqueous dispersions. For this purpose, the chain composition has been gradually varied from saturated tetradecyl (T, C14:0) and hexadecyl (H, C16:0) chains to longer but unsaturated oleyl (O, C18:1) chains with double bonds in the cis configuration. In this work, the lipid dispersions have been investigated in the absence and presence of the helper lipid DOPE and calf thymus DNA by small-angle and wide-angle X-ray scattering (SAXS/WAXS) supplemented by differential scanning calorimetry (DSC), attenuated total reflection Fourier-transform infrared spectroscopy (ATR-FTIR) and Fourier-transform Raman spectroscopy (FTRS). Lamellar and inverted hexagonal mesophases have been observed in single-component systems. In the binary mixtures, the aggregation behaviour changes with an increasing amount of DOPE from lamellar to cubic. The lipid mixtures with DNA show a panoply of mesophases. Interestingly, TT10 and OT10 form cubic lipoplexes, whereas OO10 complexes the DNA sandwich-like between lipid bilayers in a lamellar lipoplex. Surprisingly, the latter is the most effective lipoplex.
Subject(s)
DNA/chemistry , Lipid Bilayers/chemistry , Liposomes/chemistry , Lysine/chemistry , Transfection/methods , A549 Cells , Animals , DNA/metabolism , Gene Expression , Green Fluorescent Proteins/genetics , HeLa Cells , Humans , LLC-PK1 Cells , Swine , ThermodynamicsABSTRACT
The influence of the chain composition on the physical-chemical properties will be discussed for five transfection lipids containing the same lysine-based head group. For this purpose, the chain composition will be gradually varied from saturated tetradecyl (C14:0) and hexadecyl (C16:0) chains to longer but unsaturated oleyl (C18:1) chains with double bonds in the cis configuration. In this work, we investigated the lipids as Langmuir monolayers at the air-water-interface in the absence and presence of calf thymus DNA applying different techniques such as infrared reflection absorption spectroscopy (IRRAS) and grazing incidence X-ray diffraction (GIXD). The replacement of saturated tetradecyl (C14:0) and hexadecyl (C16:0) chains by unsaturated oleyl (C18:1) chains increases the fluidity of the lipid monolayer: TH10 < TT10 < OH10 < OT10 < OO10 resulting in a smaller packing density. TH10 forms the stiffest and OO10 the most fluid monolayer in this structure-property study. OO10 has a higher protonation degree compared to the saturated lipids TT10 and TH10 as well as to the hybrids OT10 and OH10 because of a better accessibility of the amine groups. Depending on the bulk pH, different scenarios of DNA coupling to the lipid monolayers have been proposed.
Subject(s)
Amines/chemistry , Lipids/chemistry , Lysine/chemistry , Animals , Cattle , DNA/chemistry , Hydrophobic and Hydrophilic Interactions , Molecular Structure , Surface Properties , Transfection , Water/chemistryABSTRACT
BACKGROUND: home enteral nutrition (HEN) is the best option for chronic. patients without the ability to swallow, but with intact digestive tract. Despite the increasing use of home enteral tube feeding (HETF), there is little published information about the types of patients receiving home enteral nutrition. The purpose of this paper to present the evolution of HETF. MATERIAL AND METHODS: the retrospective multicenter observational study was performed using questionnaires, which were distributed among the biggest Polish HEN centres. The study covered all patients treated between January, 2007 and January, 2014. RESULTS: in total 196 adult patients in 2008 (M:104. F: 92, mean age 58.1 [41-75]) and 2842 in 2013 (M: 1541, F: 1301, mean age 61.4 range: 1-91) were assessed. The number of patients grew significantly between 2008 and 2013 (p < 0.05), rising from 196 up to 2 842 (and 1 716 at the moment of study). The predominant primary disease was neurology in both time periods, but the profile switched from neurovascular to neurodegenerative (p > 0.05). Percutaneous endoscopic gastrostomy was the most common GI access ( > 60%), its use and the use of gastrostomies increased significantly since 2008 (p < 0.05). Although the reimbursement for HETF started in 2007, HEN centres expressed doubts about unclear rules for the qualification to HEN and its use. CONCLUSIONS: HETF is a safe, well-tolerated and cost-effective procedure. The profile of patients and techniques may vary at the beginning, but becomes similar to other HETF countries relatively soon. The number of patients grows quickly, and that fact suggests that the prevalence of HETF is similar in all countries.
Antecedentes: el inicio con nutrición enteral (HEN) es la mejor opción para los pacientes crónicos sin capacidad de tragar, pero con el tracto digestivo intacto. A pesar del aumento en el uso de la alimentación por sonda enteral domiciliaria (HETF), hay poca información publicada sobre los tipos de pacientes que reciben nutrición enteral domiciliaria. El propósito de este trabajo es presentar la evolución de HETF. Material y métodos: el estudio observacional multicéntrico retrospectivo se realizó mediante cuestionarios que se distribuyeron entre los mayores centros HEN polacos. El estudio abarcó a todos los pacientes tratados entre enero de 2007 y enero de 2014. Resultados: en total fueron evaluados 196 pacientes adultos en 2008 (M: 104 F: 92, edad media 58,1 [41-75]) y 2.842 en 2013 (M: 1541, F: 1.301, con una edad media de 61,4 rango: 1-91). El número de pacientes aumentó significativamente entre 2008 y 2013 (p < 0,05), al pasar de 196 hasta 2.842 (y 1.716 en el momento del estudio). La enfermedad primaria predominante fue la neurología en ambos períodos de tiempo, pero el perfil cambia de neurovascular a neurodegenerativa (p > 0,05). La gastrostomía endoscópica percutánea fue el acceso GI más frecuentes (> 60%), su uso y el uso de gastrostomías aumentó significativamente desde 2008 (p < 0,05). Aunque el reembolso de HETF comenzó en 2007, los centros de HEN expresaron dudas sobre las reglas poco claras para la calificación para HEN y su uso. Conclusiones: HETF es un procedimiento seguro, bien tolerado y rentable. El perfil de los pacientes y las técnicas puede variar al principio, pero se vuelve similar a otros países HETF relativamente pronto. El número de pacientes crece rápidamente, y ese hecho sugiere que la prevalencia de HETF es similar en todos los países.
Subject(s)
Enteral Nutrition , Home Care Services/statistics & numerical data , Adolescent , Adult , Aged , Aged, 80 and over , Child , Child, Preschool , Enteral Nutrition/methods , Female , Humans , Infant , Male , Middle Aged , Outcome Assessment, Health Care , Poland , Retrospective Studies , Young AdultABSTRACT
The front cover artwork is provided by the groups of Prof. Bodo Dobner, Prof. Andreas Langner, and research partners Dr. Gerd Hause, Dr. Simon Drescher, and Dr. Annette Meister (MLU Halle-Wittenberg) as well as the group of Prof. Gerald Brezesinski (MPI of Colloids and Interfaces). The image shows the space-filling model of a three-chain amino-functionalized lipid designed for gene transfer and the preferred pH-dependent aggregates (multilamellar stacks, vesicles, rod-like micelles). The background shows a Cryo-TEM image of rod-like micelles. Read the full text of the article at 10.1002/cphc.201500188.
ABSTRACT
Background: home enteral nutrition (HEN) is the best option for chronic. patients without the ability to swallow, but with intact digestive tract. Despite the increasing use of home enteral tube feeding (HETF), there is little published information about the types of patients receiving home enteral nutrition. The purpose of this paper to present the evolution of HETF. Material and methods: the retrospective multicenter observational study was performed using questionnaires, which were distributed among the biggest Polish HEN centres. The study covered all patients treated between January, 2007 and January, 2014. Results: in total 196 adult patients in 2008 (M:104. F: 92, mean age 58.1 [41-75]) and 2842 in 2013 (M: 1541, F: 1301, mean age 61.4 range: 1-91) were assessed. The number of patients grew significantly between 2008 and 2013 (p0.05). Percutaneous endoscopic gastrostomy was the most common GI access (>60%), its use and the use of gastrostomies increased significantly since 2008 (p <0.05). Although the reimbursement for HETF started in 2007, HEN centres expressed doubts about unclear rules for the qualification to HEN and its use. Conclusions: HETF is a safe, well-tolerated and cost-effective procedure. The profile of patients and techniques may vary at the beginning, but becomes similar to other HETF countries relatively soon. The number of patients grows quickly, and that fact suggests that the prevalence of HETF is similar in all countries (AU)
Antecedentes: el inicio con nutrición enteral (HEN) es la mejor opción para los pacientes crónicos sin capacidad de tragar, pero con el tracto digestivo intacto. A pesar del aumento en el uso de la alimentación por sonda enteral domiciliaria (HETF), hay poca información publicada sobre los tipos de pacientes que reciben nutrición enteral domiciliaria. El propósito de este trabajo es presentar la evolución de HETF. Material y métodos: el estudio observacional multicéntrico retrospectivo se realizó mediante cuestionarios que se distribuyeron entre los mayores centros HEN polacos. El estudio abarcó a todos los pacientes tratados entre enero de 2007 y enero de 2014. Resultados: en total fueron evaluados 196 pacientes adultos en 2008 (M: 104 F: 92, edad media 58,1 [41-75]) y 2.842 en 2013 (M: 1541, F: 1.301, con una edad media de 61,4 rango: 1-91). El número de pacientes aumentó significativamente entre 2008 y 2013 (p0,05). La gastrostomía endoscópica percutánea fue el acceso GI más frecuentes (>60%), su uso y el uso de gastrostomías aumentó significativamente desde 2008 (p< 0,05). Aunque el reembolso de HETF comenzó en 2007, los centros de HEN expresaron dudas sobre las reglas poco claras para la calificación para HEN y su uso. Conclusiones: HETF es un procedimiento seguro, bien tolerado y rentable. El perfil de los pacientes y las técnicas puede variar al principio, pero se vuelve similar a otros países HETF relativamente pronto. El número de pacientes crece rápidamente, y ese hecho sugiere que la prevalencia de HETF es similar en todos los países (AU)
Subject(s)
Humans , Enteral Nutrition/trends , Nutritional Support/methods , Nutrition Disorders/diet therapy , Poland , Home Care Services, Hospital-Based/trendsABSTRACT
The aggregation behavior of a cationic lipid, N-[6-amino-1-oxo-1-(N-tetradecylamino)hexan-(2S)-2-yl]-N'-{2-[N,N-bis(2-aminoethyl)amino]ethyl}-2,2-ditetradecylpropandiamide (DiTT4), is investigated in aqueous dispersions at different pH values (5, 7.3, and 10). An unusual aggregation behavior is observed whereby DiTT4 forms bilayer structures at pH 10 and 7.3. At pH 5, rod-like micelles are the dominant aggregate form. The thermotropic and lyotropic behavior is studied using differential scanning calorimetry, small-angle X-ray scattering, and FTIR spectroscopy. In addition, investigations at the air-water interface are performed by recording area-pressure-isotherms and infrared reflection-absorption (IRRA) spectra. Complementary dynamic light scattering experiments and transmission electron microscopy (TEM and cryoTEM) are also used. The ability of DiTT4 to complex plasmid DNA is investigated using fluorescence techniques and zeta potential measurements. Cell culture experiments demonstrate the ability of DiTT4 to enhance plasmid transfer in A549 cells.
Subject(s)
Lipids/chemistry , Cations/chemical synthesis , Cations/chemistry , Cell Line, Tumor , DNA/chemistry , Humans , Hydrogen-Ion Concentration , Lipids/chemical synthesis , Micelles , Molecular Structure , PlasmidsABSTRACT
Two novel micelle-forming amino-functionalized lipids (OT6 and TT6) bearing two alkyl chains connected to a large positively charged hexavalent headgroup, which might be interesting polynucleotide transferring agents with the advantage of an easy and reproducible production of micelle dispersions, have been characterized. The critical micelle concentration (cmc) of both lipids has been determined by two different methods, namely, isothermal titration calorimetry (ITC) and 1,6-diphenyl-1,3,5-hexatriene (DPH) fluorescence experiments. In addition, the lipid dispersions were studied as a function of temperature using differential scanning calorimetry (DSC), dynamic light scattering (DLS), Fourier-transform infrared (FT-IR) spectroscopy, and cryo-transmission electron microscopy (cryo-TEM). The OT6 and TT6 micelles effectively complex DNA as determined by ITC and DSC measurements. In addition, DLS and ζ-potential measurements were performed to determine lipoplex formulations that exhibit colloidal stability. Finally, the structures of OT6/DNA complexes were investigated by means of X-ray scattering and TEM.
