Cationic Vitamin E-TPGS Mixed Micelles of Berberine to Neutralize Doxorubicin-Induced Cardiotoxicity via Amelioration of Mitochondrial Dysfunction and Impeding Apoptosis.

Anthracycline antibiotics, namely, doxorubicin (DOX) and daunorubicin, are among the most widely used anticancer therapies, yet are notoriously associated with severe myocardial damage due to oxidative stress and mitochondrial damage. Studies have indicated the strong pharmacological properties of Berberine (Brb) alkaloid, predominantly mediated via mitochondrial functions and nuclear networks. Despite the recent emphasis on Brb in clinical cardioprotective studies, pharmaceutical limitations hamper its clinical use. A nanoformulation for Brb was developed (mMic), incorporating a cationic lipid, oleylamine (OA), into the TPGS-mixed corona of PEGylated-phosphatidylethanolamine (PEG-PE) micelles. Cationic TPGS/PEG-PE mMic with superior Brb loading and stability markedly enhanced both intracellular and mitochondria-tropic Brb activities in cardiovascular muscle cells. Sub-lethal doses of Brb via cationic OA/TPGS mMic, as a DOX co-treatment, resulted in significant mitochondrial apoptosis suppression. In combination with an intense DOX challenge (up to ~50 µM), mitochondria-protective Brb-OA/TPGS mMic showed a significant 24 h recovery of cell viability (p ≤ 0.05-0.01). Mechanistically, the significant relative reduction in apoptotic caspase-9 and elevation of antiapoptotic Bcl-2 seem to mediate the cardioprotective role of Brb-OA/TPGS mMic against DOX. Our report aims to demonstrate the great potential of cationic OA/TPGS-mMic to selectively enhance the protective mitohormetic effect of Brb to mitigate DOX cardiotoxicity.

Computer-Assisted Drug Formulation Design: Novel Approach in Drug Delivery.

We hypothesize that, by using several chemo/bio informatics tools and statistical computational methods, we can study and then predict the behavior of several drugs in model nanoparticulate lipid and polymeric systems. Accordingly, two different matrices comprising tripalmitin, a core component of solid lipid nanoparticles (SLN), and PLGA were first modeled using molecular dynamics simulation, and then the interaction of drugs with these systems was studied by means of computing the free energy of binding using the molecular docking technique. These binding energies were hence correlated with the loadings of these drugs in the nanoparticles obtained experimentally from the available literature. The obtained relations were verified experimentally in our laboratory using curcumin as a model drug. Artificial neural networks were then used to establish the effect of the drugs' molecular descriptors on the binding energies and hence on the drug loading. The results showed that the used soft computing methods can provide an accurate method for in silico prediction of drug loading in tripalmitin-based and PLGA nanoparticulate systems. These results have the prospective of being applied to other nano drug-carrier systems, and this integrated statistical and chemo/bio informatics approach offers a new toolbox to the formulation science by proposing what we present as computer-assisted drug formulation design (CADFD).

Quantitative silencing of EGFP reporter gene by self-assembled siRNA lipoplexes of LinOS and cholesterol.

Nonviral siRNA vectors prepared by the direct mixing of siRNA and mixtures of an asymmetric N(4),N(9)-diacyl spermine conjugate, N(4)-linoleoyl-N(9)-oleoyl-1,12-diamino-4,9-diazadodecane (LinOS), with either cholesterol or DOPE, at various molar ratios of the neutral lipids, are reported. The effects of varying the lipid formulation and changing the N/P charge ratio on the intracellular delivery of siRNA to HeLa cells and on the siRNA-mediated gene silencing of a stably expressed reporter gene (EGFP) were evaluated. The presence of either cholesterol or DOPE in the mixture resulted in a marked increase in the delivery of the siRNA as well as enhanced EGFP silencing as evaluated by FACS. A LinOS/Chol 1:2 mixture resulted in the highest siRNA delivery and the most efficient EGFP silencing (reduced to 20%) at N/P = 3.0. Lowering the amount of siRNA from 15 pmol to 3.75 pmol, thus increasing the N/P charge ratio to 11.9, resulted in decreasing the amount of delivered siRNA, while the efficiency of gene silencing was comparable to that obtained with 15 pmol (N/P = 3.0) of siRNA. Mixtures of symmetrical N(4),N(9)-dioleoyl spermine (DOS) with cholesterol at 1:2 molar ratio showed less siRNA delivery than with LinOS/Chol at N/P = 3.0 (15 pmol of siRNA), and comparable delivery at N/P = 11.9 (3.75 pmol of siRNA). The EGFP silencing was comparable with LinOS and with DOS when mixed with cholesterol 1:2 (lipoplexes prepared with 15 pmol of siRNA), but LinOS mixtures showed better EGFP silencing when the siRNA was reduced to 3.75 pmol. Lipoplex particle size determination by DLS of cholesterol mixtures was 106-118 nm, compared to 194-356 nm for lipoplexes prepared with the spermine conjugates only, and to 685 nm for the LinOS/DOPE 1:1 mixture. Confocal microscopy showed successful siRNA delivery of red tagged siRNA and quantitative EGFP knockdown in HeLa EGFP cells; Z-stack photomicrographs showed that the delivered siRNA is distributed intracellularly. Cryo-TEM of siRNA LinOS/Chol 1:2 lipoplexes shows the formation of multilamellar spheres with a size of ∼100 nm, in good agreement with the particle size measured by DLS. The constant distance between lamellar repeats is ∼6 nm, with the electron-dense layers fitting a monolayer of siRNA. AlamarBlue cell viability assay showed that the lipoplexes resulted in cell viability ≥81%, with LinOS/Chol 1:2 mixtures resulting in cell viabilities of 89% and 94% at siRNA 15 nM and 3.75 nM respectively. These results show that lipoplexes of siRNA and LinOS/Chol mixtures prepared by the direct mixing of the lipid mixture and siRNA, without any preceding preformulation steps, result in enhanced siRNA delivery and EGFP knockdown, with excellent cell viability. Thus, LinOS/Chol 1:2 mixture is a promising candidate as a nontoxic nonviral siRNA vector.

Novel thymoquinone lipidic core nanocapsules with anisamide-polymethacrylate shell for colon cancer cells overexpressing sigma receptors.

The biggest challenge in colorectal cancer therapy is to avoid intestinal drug absorption before reaching the colon, while focusing on tumor specific delivery with high local concentration and minimal toxicity. In our work, thymoquinone (TQ)-loaded polymeric nanocapsules were prepared using the nanoprecipitation technique using Eudragit S100 as polymeric shell. Conjugation of anisamide as a targeting ligand for sigma receptors overexpressed by colon cancer cells to Eudragit S100 was carried out via carbodiimide coupling reaction, and was confirmed by thin layer chromatography and 1H-NMR. TQ nanocapsules were characterized for particle size, surface morphology, zeta potential, entrapment efficiency % (EE%), in vitro drug release and physical stability. A cytotoxicity study on three colon cancer cell lines (HT-29, HCT-116, Caco-2) was performed. Results revealed that the polymeric nanocapsules were successfully prepared, and the in vitro characterization showed a suitable size, zeta potential, EE% and physical stability. TQ exhibited a delayed release pattern from the nanocapsules in vitro. Anisamide-targeted TQ nanocapsules showed higher cytotoxicity against HT-29 cells overexpressing sigma receptors compared to their non-targeted counterparts and free TQ after incubation for 48 h, hence delineating anisamide as a promising ligand for active colon cancer targeting.

Gelatinized core liposomes: A new Trojan horse for the development of a novel timolol maleate glaucoma medication.

Glaucoma treatment with ocular medications requires overcoming the corneal barrier to drug penetration. Liposomes have a great corneal penetration ability and affinity while suffering from poor stability and low entrapment of hydrophilic drugs accompanied by rapid drug release. This work aims to develop a new, effective and stable glaucoma medication with sustained drug release properties; Timolol maleate gelatinized core liposomes. A full factorial design was utilized to study the effects of three formulation variables on drug loading and vesicle particle size. Vesicles were prepared by the thin-film hydration method, and characterized for in-vitro drug release and stability. Intra-ocular pressure (IOP) reduction was evaluated in-vivo on glaucomatous rabbit's eyes. The safety profile was assessed using histopathological examinations. Gelatin significantly increased the drug entrapment percentage reaching 50% with a particle size of 38.81 µm. Sustained drug release was recorded compared to a marketed product and to a conventional liposomal formulation. The prepared vesicles caused the highest reduction in IOP accompanied by safe histological findings. This work provided a new, safe and effective ocular glaucoma medication; Timolol maleate gelatinized core liposomes, solving the main problems of ocular liposomal formulations of hydrophilic drugs, suitable for the pharmaceutical industry and comprising abundant and relatively cheap components.

Gelatinized-core liposomes: Toward a more robust carrier for hydrophilic molecules.

The use of liposomes as a delivery system for hydrophobic and hydrophilic drugs is well recognized. However, they possess several limitations that remained unresolved, including stability problems, low entrapment of the hydrophilic drugs, and the subsequent rapid release. This study introduces a novel approach to incorporate gelatin in the liposomal core to overcome these limitations. A rheological study was conducted to select suitable masses of the gelatin used in the liposomal formulations. Moreover, a full-factorial experimental design was utilized to compare the newly produced gel-core liposomes to the conventional liposomes with respect to the amount of a model hydrophilic molecule loading. An advanced machine learning method, namely, artificial neural networks was utilized to capture the effects of gelatin and cholesterol incorporation in the liposomes on the entrapment efficiency. The results revealed the successful preparation of the novel vesicles and their superiority over the conventional liposomes in drug loading, sustaining the drug release and stability which pose the newly introduced liposomal system as a successful delivery carrier for hydrophilic molecules and drugs. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 3086-3092, 2017.

Self-assembled amphiphilic core-shell nanocarriers in line with the modern strategies for brain delivery.

Disorders of the central nervous system (CNS) represent increasing social and economic problems all over the world which makes the effective transport of drugs to the brain a crucial need. In the last decade, many strategies were introduced to deliver drugs to the brain trying to overcome the challenge of the blood brain barrier (BBB) using both invasive and non-invasive methods. Non-invasive strategy represented in the application of nanocarriers became very common. One of the most hopeful nanoscopic carriers for brain delivery is core-shell nanocarriers or polymeric micelles (PMs). They are more advantageous than other nanocarriers. They offer small size, ease of preparation, ease of sterilization and the possibility of surface modification with various ligands. Hence, the aim of this review is to discuss modern strategies for brain delivery, micelles as a successful delivery system for the brain and how micelles could be modified to act as "magic bullets" for brain delivery.

Towards better modelling of drug-loading in solid lipid nanoparticles: Molecular dynamics, docking experiments and Gaussian Processes machine learning.

This study represents one of the series applying computer-oriented processes and tools in digging for information, analysing data and finally extracting correlations and meaningful outcomes. In this context, binding energies could be used to model and predict the mass of loaded drugs in solid lipid nanoparticles after molecular docking of literature-gathered drugs using MOE® software package on molecularly simulated tripalmitin matrices using GROMACS®. Consequently, Gaussian processes as a supervised machine learning artificial intelligence technique were used to correlate the drugs' descriptors (e.g. M.W., xLogP, TPSA and fragment complexity) with their molecular docking binding energies. Lower percentage bias was obtained compared to previous studies which allows the accurate estimation of the loaded mass of any drug in the investigated solid lipid nanoparticles by just projecting its chemical structure to its main features (descriptors).

Measurement of polyamine pKa values.

The extent of ionization of the polyamines is an important factor in their interactions with cellular components. The pK(a) is the pH at which a functional group is 50% ionized. For compounds such as polyamines with more than one ionizable center (atom or functional group), there is a pK(a) value for each center of ionization. This chapter describes the pK(a) values for each amine group in many important polyamines, the factors influencing these values and methods for their determination using potentiometric titration and nuclear magnetic resonance spectroscopy.