Development of tamoxifen-loaded surface-modified nanostructured lipid carrier using experimental design: in vitro and ex vivo characterisation.

The present study aimed to develop a surface-modified biocompatible nanostructured lipid carrier (NLCs) system using polyoxyethylene (40) stearate (POE-40-S) to improve the oral bioavailability of poorly water-soluble Biopharmaceutics Classification System class-II drug like tamoxifen (TMX). Also aimed to screen the most influential factors affecting the particle size (PS) using Taguchi (L12 (211)) orthogonal array design (TgL12OA). Then, to optimize the TMX loaded POE-40-S (P) surface-modified NLCs (TMX-loaded-PEG-40-S coated NLC (PNLCs) or PNLCs) by central composite design (CCD) using a four-factor, five-level model. The most influential factors affecting the PS was screened and optimized. The in-vitro study showed that increased drug-loading (DL) and encapsulation efficiency (EE), decreased PS and charge, sustained drug release for the prolonged period of the time with good stability and suppressed protein adsorption. The Ex-vivo study showed that decreased mucous binding with five-fold enhanced permeability of PNLC formulation after surface modification with POE-40-S. The in-vitro cytotoxicity study showed that the blank carrier is biocompatible and cytotoxicity of the formulation was dependent on the concentration of the drug. Finally, it can be concluded that the surface-modified PNLCs formulation was an effective, biocompatible, stable formulation in the enhancement of dissolution rate, solubility, stability with reduced mucus adhesion and increased permeability thereby which indicates its enhanced oral bioavailability.

Improved Oral Pharmacokinetics of Pentoxifylline with Palm Oil and Capmul® MCM Containing Self-Nano-Emulsifying Drug Delivery System.

Pentoxifylline (PTX), an anti-hemorrhage drug used in the treatment of intermittent claudication, is extensively metabolized by the liver resulting in a reduction of the therapeutic levels within a short duration of time. Self-nano-emulsifying drug delivery system (SNEDDS) is well reported to enhance the bio-absorption of drugs by forming nano-sized globules upon contact with the biological fluids after oral administration. The present study aimed to formulate, characterize, and improve the oral bioavailability of PTX using SNEDDS. The formulated SNEDDS consisted of palm oil, Capmul® MCM, and Tween® 80 as oil, surfactant, and co-surfactant, respectively. The mixture design module under the umbrella of the design of experiments was used for the optimization of SNEDDS. The dynamic light-scattering technique was used to confirm the formation of nanoemulsion based on the globule size, in addition to the turbidity measurements. In vivo bioavailability studies were carried out on male Wistar rats. The pharmacokinetic parameters upon oral administration were calculated using the GastroPlus software. The optimized SNEDDS had a mean globule size of 165 nm with minimal turbidity in an aqueous medium. Bioavailability of PTX increased 1.5-folds (AUC = 1013.30 ng h/mL) as SNEDDS than the pure drug with an AUC of 673.10 ng h/mL. In conclusion, SNEDDS was seen to enhance the bioavailability of PTX and can be explored to effectively control the incidents of intermittent claudication.

Novel Phospholipid-Based Labrasol Nanomicelles Loaded Flavonoids for Oral Delivery with Enhanced Penetration and Anti-Brain Tumor Efficiency.

BACKGROUND: Owing to the rich anticancer properties of flavonoids, there is a need for their incorporation into drug delivery vehicles like nanomicelles for safe delivery of the drug into the brain tumor microenvironment. OBJECTIVE: This study, therefore, aimed to prepare the phospholipid-based Labrasol/Pluronic F68 modified nano micelles loaded with flavonoids (Nano-flavonoids) for the delivery of the drug to the target brain tumor. METHODS: Myricetin, quercetin and fisetin were selected as the initial drugs to evaluate the biodistribution and acute toxicity of the drug delivery vehicles in rats with implanted C6 glioma tumors after oral administration, while the uptake, retention, release in human intestinal Caco-2 cells and the effect on the brain endothelial barrier were investigated in Human Brain Microvascular Endothelial Cells (HBMECs). RESULTS: The results demonstrated that nano-flavonoids loaded with myricetin showed more evenly distributed targeting tissues and enhanced anti-tumor efficiency in vivo without significant cytotoxicity to Caco-2 cells and alteration in the Trans Epithelial Electric Resistance (TEER). There was no pathological evidence of renal, hepatic or other organs dysfunction after the administration of nanoflavonoids, which showed no significant influence on cytotoxicity to Caco-2 cells. CONCLUSION: In conclusion, Labrasol/F68-NMs loaded with MYR and quercetin could enhance antiglioma effect in vitro and in vivo, which may be better tools for medical therapy, while the pharmacokinetics and pharmacodynamics of nano-flavonoids may ensure optimal therapeutic benefits.

Absorption mechanism of DHP107, an oral paclitaxel formulation that forms a hydrated lipidic sponge phase.

Paclitaxel is a most widely used anticancer drug with low oral bioavailability, thus it is currently administered via intravenous infusion. DHP107 is a lipid-based paclitaxel formulation that can be administered as an oral solution. In this study, we investigated the mechanism of paclitaxel absorption after oral administration of DHP107 in mice and rats by changing the dosing interval, and evaluated the influence of bile excretion. DHP107 was orally administered to mice at various dosing intervals (2, 4, 8, 12, 24 h) to examine how residual DHP107 affected paclitaxel absorption during subsequent administration. Studies with small-angle X-ray diffraction (SAXS) and cryo-transmission electron microscopy (cryo-TEM) showed that DHP107 formed a lipidic sponge phase after hydration. The AUC values after the second dose were smaller than those after the first dose, which was correlated to the induction of expression of P-gp and CYP in the livers and small intestines from 2 h to 7 d after the first dose. The smaller AUC value observed after the second dose was also attributed to the intestinal adhesion of residual formulation. The adhered DHP107 may have been removed by ingested food, thus resulting in a higher AUC. In ex vivo and in vivo mucoadhesion studies, the formulation adhered to the villi for up to 24 h, and the amount of DHP107 that adhered was approximately half that of monoolein. The paclitaxel absorption after administration of DHP107 was not affected by bile in the cholecystectomy mice. The dosing interval and food intake affect the oral absorption of paclitaxel from DHP107, which forms a mucoadhesive sponge phase after hydration. Bile excretion does not affect the absorption of paclitaxel from DHP107 in vivo.

4'-O-methylhonokiol increases levels of 2-arachidonoyl glycerol in mouse brain via selective inhibition of its COX-2-mediated oxygenation.

BACKGROUND AND PURPOSE: 4'-O-methylhonokiol (MH) is a natural product showing anti-inflammatory, anti-osteoclastogenic, and neuroprotective effects. MH was reported to modulate cannabinoid CB2 receptors as an inverse agonist for cAMP production and an agonist for intracellular [Ca2+]. It was recently shown that MH inhibits cAMP formation via CB2 receptors. In this study, the exact modulation of MH on CB2 receptor activity was elucidated and its endocannabinoid substrate-specific inhibition (SSI) of cyclooxygenase-2 (COX-2) and CNS bioavailability are described for the first time. METHODS: CB2 receptor modulation ([35S]GTPγS, cAMP, and ß-arrestin) by MH was measured in hCB2-transfected CHO-K1 cells and native conditions (HL60 cells and mouse spleen). The COX-2 SSI was investigated in RAW264.7 cells and in Swiss albino mice by targeted metabolomics using LC-MS/MS. RESULTS: MH is a CB2 receptor agonist and a potent COX-2 SSI. It induced partial agonism in both the [35S]GTPγS binding and ß-arrestin recruitment assays while being a full agonist in the cAMP pathway. MH selectively inhibited PGE2 glycerol ester formation (over PGE2) in RAW264.7 cells and significantly increased the levels of 2-AG in mouse brain in a dose-dependent manner (3 to 20 mg kg(-1)) without affecting other metabolites. After 7 h from intraperitoneal (i.p.) injection, MH was quantified in significant amounts in the brain (corresponding to 200 to 300 nM). CONCLUSIONS: LC-MS/MS quantification shows that MH is bioavailable to the brain and under condition of inflammation exerts significant indirect effects on 2-AG levels. The biphenyl scaffold might serve as valuable source of dual CB2 receptor modulators and COX-2 SSIs as demonstrated by additional MH analogs that show similar effects. The combination of CB2 agonism and COX-2 SSI offers a yet unexplored polypharmacology with expected synergistic effects in neuroinflammatory diseases, thus providing a rationale for the diverse neuroprotective effects reported for MH in animal models.

Physico-chemical characterisation, cytotoxic activity, and biocompatibility studies of tamoxifen-loaded solid lipid nanoparticles prepared via a temperature-modulated solidification method.

CONTEXT: Solid lipid nanoparticles (SLNs) can efficiently and efficaciously incorporate anti-cancer agents. OBJECTIVE: To prepare and characterise tamoxifen (TAM)-loaded SLNs. MATERIALS AND METHODS: Glyceryl monostearate, Tween-80, and trehalose were used in SLNs. SLNs were tested via dynamic light scattering (DLS), transmission electron microscopy (TEM), differential scanning calorimetry (DSC), X-ray diffraction (XRD), and Fourier transform infrared spectroscopy (FTIR). RESULTS: Characterisation studies revealed SLNs of about 540 nm with a negative surface charge and confirmed the entrapment of TAM in the SLNs. The entrapment efficiency was estimated to be 60%. DISCUSSION: The in vitro drug release profile demonstrated a gradual increase followed by a release plateau for several days. A drug concentration-dependent increase in cytotoxic activity was observed when the SLNs were evaluated in cell cultures. CONCLUSION: Biocompatible and stable lyophilised SLNs were successfully prepared and found to possess properties that may be utilised in an anti-cancer drug delivery system.

Toxicological evaluation of neem (Azadirachta indica) oil: acute and subacute toxicity.

Neem (Azadirachta indica), popularly known as traditional medicine is a native plant in India. Neem oil is a vegetable oil derived from seeds or fruits of the neem tree through pressing or solvent extraction, and largely used in popular medicine to have antifungal, antibacterial, antimalarial, antiparasitic, anti-inflammatory, as well as immunemodulatory properties in different animal species. In the present study, acute and 28-day subacute toxicity tests were carried out. In the acute toxicity test, the LD50 values of neem oil were found to be 31.95g/kg. The subacute treatment with neem oil failed to change body weight gain, food and water consumption. Serum biochemistry analysis showed no significant differences in any of the parameters examined under the dose of 1600mg/kg/day. Histopathological exams showed that the target organs of neem oil were testicle, liver and kidneys up to the dose of 1600mg/kg/day.

Antitumor efficacy, tumor distribution and blood pharmacokinetics of chitosan/glyceryl-monooleate nanostructures containing paclitaxel.

AIMS: This investigation compared the tumor distribution, efficacy, blood pharmacokinetic parameters and hematological alterations following treatment with chitosan/glyceryl-monooleate (GMO) nanostructures containing paclitaxel (PTX) to a conventional formulation of PTX (Taxol(®)) in BALB/c female mice. MATERIALS & METHODS: The tumor and blood concentrations of PTX were evaluated by HPLC and the pharmacokinetic parameters were determined through noncompartmental methods. Tumor development was evaluated by histopathological methods and hematological composition was monitored through differential white blood cells counts. RESULTS: Lower localized or intravenous doses of PTX-chitosan/GMO nanostructures significantly increased the antitumor activity of paclitaxel. The tumor distribution studies showed effective concentrations in the tumors with the chitosan/GMO formulation while systemic blood levels remained lower than after administration of the conventional formulation. CONCLUSION: Delivery systems consisting of chitosan/GMO and PTX are safe and effective administered locally (intratumorally) or intravenously.

Preparation of multiparticulate vaginal tablet using glyceryl monooleate for sustained progesterone delivery.

Most of the sustained release vaginal formulations are in the form of bioadhesive gels and tablets. Though proved efficient, their presence in the vagina for a longer time as a bulk produces discomfort and interference with body functioning including sexual activities. Hence, they lack complete patient compliance. In this study, multiparticulate vaginal tablets were prepared by utilizing progesterone (PRO) loaded dry powder precursor of cubic phase (DPPCP) of glyceryl monooleate (GMO). DPPCP were obtained by spray drying GMO with magnesium trisilicate (MTS) and have presented PRO sustained release in simulated vaginal fluid (SVF) for 14 hours. The effect of hydrophilic and hydrophobic tableting excipients on compression, phase, bioadhesion and drug release properties of prepared tablets was evaluated. The effervescent hydrophilic tablet (EHT) prepared with hydrophilic excipients showed rapid disintegration but, diminished sustaining ability owing to transformation into lamellar phase whereas the multiparticulate hydrophobic tablet (MHT) obtained from hydrophobic excipients presented both rapid disintegration and sustained release in SVF by virtue of cubic phase retention. During bioadhesivity testing, fast disintegration of MHT with formation of uniform and viscous bioadhesive layer on cow mucosa was observed even with a small volume of SVF. As MHT may not produce discomfort and interference, it will be preferred over bioadhesive gel or tablet.

Enhanced in vitro percutaneous absorption and in vivo anti-inflammatory effect of a selective cyclooxygenase inhibitor using microemulsion.

Celecoxib, a specific COX-2 inhibitor, was recently approved for the treatment of rheumatoid and osteoarthritis, acute pain, familial adenomatous polyposis and primary dysmenorrhea. Oral administration of celecoxib is effective against ultraviolet B radiation (UVB)-induced skin carcinogenesis; however, its clinical use is restricted because of its failure to block the characteristic cutaneous inflammatory response and lower availability at the site of inflammation. Topical application of celecoxib has been effective compared with oral in certain clinical conditions. The present study was undertaken to develop and investigate the development of microemulsion system (isopropyl myristate/medium-chain glyceride/polysorbate 80/water) for topical delivery of celecoxib. The pseudoternary phase diagram was constructed with constant surfactant concentration, and several compositions were identified and characterized by using dynamic light scattering. The in vitro permeation rate of celecoxib through rat skin was determined for microemulsions, microemulsion gel, and cream by using the modified Franz-type diffusion cell. In all formulations tested, celecoxib permeated more quickly, and the microemulsions increased the permeation rate of celecoxib up to 5 and 11 times compared with those of microemulsion gel and cream, respectively. Increasing the concentration of medium-chain mono-/di-glyceride in microemulsion imparted increased droplet size and viscosity and decreased diffusion coefficient. In vivo anti-inflammatory study suggested that the developed microemulsion formulations might serve as potential drug vehicle for the prevention of UVB-induced skin cancer.

The uptake by cells of 2-arachidonoylglycerol, an endogenous agonist of cannabinoid receptors.

It is not yet clear if the endocannabinoid 2-arachidonoylglycerol (2-AG) is transported into cells through the same membrane transporter mediating the uptake of the other endogenous cannabinoid, anandamide (N-arachidonoylethanolamine, AEA), and whether this process (a) is regulated by cells and (b) limits 2-AG pharmacological actions. We have studied simultaneously the facilitated transport of [14C]AEA and [3H]2-AG into rat C6 glioma cells and found uptake mechanisms with different efficacies but similar affinities for the two compounds (Km 11.0 +/- 2.0 and 15.3 +/- 3.1 microM, Bmax 1.70 +/- 0.30 and 0.24 +/- 0.04 nmol.min-1.mg protein-1, respectively). Despite these similar Km values, 2-AG inhibits [14C]AEA uptake by cells at concentrations (Ki = 30.1 +/- 3.9 microM) significantly higher than those required to either 2-AG or AEA to inhibit [3H]2-AG uptake (Ki = 18.9 +/- 1.8 and 20.5 +/- 3.2 microM, respectively). Furthermore: (a) if C6 cells are incubated simultaneously with identical concentrations of [14C]AEA and [3H]2-AG, only the uptake of the latter compound is significantly decreased as compared to that observed with [3H]2-AG alone; (b) the uptake of [14C]AEA and [3H]2-AG by cells is inhibited with the same potency by AM404 (Ki = 7.5 +/- 0.7 and 10.2 +/- 1.7 microM, respectively) and linvanil (Ki = 9.5 +/- 0.7 and 6.4 +/- 1.2 microM, respectively), two inhibitors of the AEA membrane transporter; (c) nitric oxide (NO) donors enhance the uptake of both [14C]AEA and [3H]2-AG, thus suggesting that 2-AG action can be regulated through NO release; (d) AEA and 2-AG induce a weak release of NO that can be blocked by a CB1 cannabinoid receptor antagonist, and significantly enhanced in the presence of AM404 and linvanil, thus suggesting that transport into C6 cells limits the action of both endocannabinoids.

Rationale and applications of lipids as prodrug carriers.

Lipidic prodrugs, also called drug-lipid conjugates, have the drug covalently bound to a lipid moiety, such as a fatty acid, a diglyceride or a phosphoglyceride. Drug-lipid conjugates have been prepared in order to take advantage of the metabolic pathways of lipid biochemistry, allowing organs to be targeted or delivery problems to be overcome. Endogenous proteins taking up fatty acids from the blood stream can be targeted to deliver the drug to the heart or liver. For glycerides, the major advantage is the modification of the pharmacokinetic behavior of the drug. In this case, one or two fatty acids of a triglyceride are replaced by a carboxylic drug. Lipid conjugates exhibit some physico-chemical and absorption characteristics similar to those of natural lipids. Non-steroidal, anti-inflammatory drugs such as acetylsalicylic acid, indomethacin, naproxen and ibuprofen were linked covalently to glycerides to reduce their ulcerogenicity. Mimicking the absorption process of dietary fats, lipid conjugates have also been used to target the lymphatic route (e.g., L-Dopa, melphalan, chlorambucil and GABA). Based on their lipophilicity and resemblance to lipids in biological membranes, lipid conjugates of phenytoin were prepared to increase intestinal absorption, whereas glycerides or modified glycerides of L-Dopa, glycine, GABA, thiorphan and N-benzyloxycarbonylglycine were designed to promote brain penetration. In phospholipid conjugates, antiviral and antineoplasic nucleosides were attached to the phosphate moiety. After presenting the biochemical pathways of lipids, the review discusses the advantages and drawbacks of lipidic prodrugs, keeping in mind the potential pharmacological activity of the fatty acid itself.

Differential regulation of phosphatidylcholine biosynthesis by 12-O-tetradecanoylphorbol-13-acetate and diacylglycerol in NG108-15 neuroblastoma x glioma hybrid cells.

12-O-Tetradecanoylphorbol-13-acetate (TPA), a tumor promoter and potent activator of protein kinase C, stimulates [3H]choline incorporation into phosphatidylcholine (PtdCho) in NG108-15 cells (Liscovitch, M., Freese, A., Blusztajn, J. K. and Wurtman, R. J. (1986) J. Neurochem. 47, 1936-1941). In the present study we demonstrate that two cell-permeant diacylglycerols, sn-1-oleoyl-2-acetylglycerol and sn-1,2-dioctanoylglycerol, also stimulate [3H]choline incorporation into PtdCho. However, the effect of diacylglycerol is additional to that produced by a maximally effective concentration of TPA (0.5 microM), suggesting that the two agents may not act via the same mechanism. In addition, the protein kinase inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (at 200 microM) inhibits the action of TPA by 59% while not affecting that of diacylglycerol. Finally, preincubation of the cells with TPA (0.1 microM) for 24 h reduces protein kinase C activity in the cells and completely abolishes the effect of additional TPA on choline incorporation. In contrast, diacylglycerol-induced stimulation of PtdCho biosynthesis was not inhibited in the cells that were desensitized to TPA. These results suggest that the effect of the two cell-permeant diacylglycerols on PtdCho biosynthesis either is not mediated by protein kinase C activation, or, is mediated by a TPA-insensitive isoenzyme of protein kinase C.