Quantcast


CURATOR
A pinboard by
Mays Al-Dulaymi

Ph.D Student , University of Saskatchewan

PINBOARD SUMMARY

A better way of delivering genes into the skin: Bridging the gap between rational and empirical desi

Gene therapy holds a promising future for the treatment of many acquired and inherited diseases. Current focus is on the design of “intelligent” nanoparticles that respond to stimuli and carry their cargo to a targeted site. Gemini surfactants (GS) have emerged as a promising class of gene delivery vectors. They composed of two ionic head-groups attached to their hydrocarbon tails and connected by a spacer. Their unique structure offers possibilities for structural modification, allowing for the production of compounds specifically designed to overcome delivery barriers. Despite the success of traditional GS, their efficiency was not satisfactory and concerns regarding toxicity profile impeded their progress toward a clinical success. Several approaches can be undertaken to address this problem, including the insertion of biocompatible moieties such as peptides that respond to environmental stimuli resulting in enhanced gene expression. In this research, a series of 22 GS was synthesized having various hydrocarbon tails and spacer group substitutions ranging from dipeptide to branched peptides. The new family of GS were evaluated for their transgene efficiency and cytotoxicity in vitro. A wide variation between the tested compounds was observed and was highly connected to the compounds’ architecture. My second objective is to assess the application of the new generation of GS-based delivery systems for topical gene delivery. Topical application of genes into the skin is an attractive method for gene delivery. It is a convenient and easy to administer, thus could enhance patient compliance. Upon topical application, GS distribute within various tissues and cellular components. Although extensive research was focused on the design of new GS, little is known about their post transfection fate. Correlating the biodistribution and the biological fate of the nanoparticles to their chemical structure will provide insights into the rational design process to produce gene carriers with higher efficiency and reduced toxicity. In my research, I utilized mass spectrometry (MS) to detect and quantify topically applied within skin tissue of CD1 mice in order to determine their biological fate and distribution. Preliminary results reveled that GS was confined to the skin with minimal escape into the circulation, suggesting the potential to of the peptide-modified GS to be used for cutaneous applications to treat conditions like scleroderma or melanoma.

2 ITEMS PINNED

Tandem Mass Spectrometric Analysis of Novel Peptide-Modified Gemini Surfactants Used as Gene Delivery Vectors.

Abstract: Diquaternary ammonium gemini Surfactants have emerged as effective gene delivery vectors. A novel series of eleven peptide-modified compounds was synthesized, showing promising results in delivering genetic materials. The purpose of this work is to elucidate the tandem mass spectrometric (MS/MS) dissociation behaviour of these novel molecules establishing a generalized MS/MS fingerprint. Exact mass measurements were achieved using a hybrid quadrupole orthogonal time-of-flight mass spectrometer (QqToF-MS) and a multi-stage tandem mass spectrometric analysis was conducted using a triple quadrupole-linear ion trap mass spectrometer (QqQLIT-MS). Both instruments were operated in the positive ionization mode and are equipped with electrospray ionization (ESI). Abundant triply charged [M + H](3+) species were observed in the single stage analysis of all the evaluated compounds with mass accuracies of less than 8 ppm in mass error. MS/MS analysis showed that the evaluated gemini surfactants exhibited peptide-related dissociation characteristics due to the presence of amino acids within the compounds' spacer region. In particular, diagnostic product ions were originated from the neutral loss of ammonia from the amino acids' side chain resulting in the formation of pipecolic acid at the N-terminus part of the gemini surfactants. In addition, a charge directed amide bond cleavage was initiated by the amino acids' side chain producing a protonated α-amino-ε-caprolactam ion and its complimentary c-terminus ion that contains quaternary amines. MS/MS and MS(3) analysis revealed common fragmentation behaviour among all tested compounds, resulting in the production of a universal MS/MS fragmentation pathway.

Pub.: 15 Apr '17, Pinned: 30 Jun '17

Di-Peptide-Modified Gemini Surfactants as Gene Delivery Vectors: Exploring the Role of the Alkyl Tail in Their Physicochemical Behavior and Biological Activity

Abstract: The aim of this work was to elucidate the structure-activity relationship of new peptide-modified gemini surfactant-based carriers. Glycyl-lysine modified gemini surfactants that differ in the length and degree of unsaturation of their alkyl tail were used to engineer DNA nano-assemblies. To probe the optimal nitrogen to phosphate (N/P) ratio in the presence of helper lipid, in vitro gene expression and cell toxicity measurements were carried out. Characterization of the nano-assemblies was accomplished by measuring the particle size and surface charge. Morphological characteristics and lipid organization were studied by small angle X-ray scattering technique. Lipid monolayers were studied using a Langmuir-Blodgett trough. The highest activity of glycyl-lysine modified gemini surfactants was observed with the 16-carbon tail compound at 2.5 N/P ratio, showing a 5- to 10-fold increase in the level of reporter protein compared to the 12 and 18:1 carbon tail compounds. This ratio is significantly lower compared to the previously studied gemini surfactants with alkyl or amino- spacers. In addition, the 16-carbon tail compound exhibited the highest cell viability (85%). This high efficiency is attributed to the lowest critical micelle concentration of the 16-tail gemini surfactant and a balanced packing of the nanoparticles by mixing a saturated and unsaturated lipid together. At the optimal N/P ratio, all nanoparticles exhibited an inverted hexagonal lipid assembly. The results show that the length and nature of the tail of the gemini surfactants play an important role in determining the transgene efficiency of the delivery system. We demonstrated here that the interplay between the headgroup and the nature of tail is specific to each series, thus in the process of rational design, the contribution of the latter should be assessed in the appropriate context.

Pub.: 16 May '16, Pinned: 30 Jun '17