Facile fabrication of multi-pocket nanoparticles with stepwise size transition for promoting deep penetration and tumor targeting.

BACKGROUND: Nanocarriers-derived antitumor therapeutics are often associated with issues of limited tumor penetration and dissatisfactory antitumor efficacies. Some multistage delivery systems have been constructed to address these issues, but they are often accompanied with complicated manufacture processes and undesirable biocompatibility, which hinder their further application in clinical practices. Herein, a novel dual-responsive multi-pocket nanoparticle was conveniently constructed through self-assembly and cross-linking of amphiphilic methoxypolyethylene glycol-lipoic acid (mPEG-LA) conjugates to enhance tumor penetration and antitumor efficacy. RESULTS: The multi-pocket nanoparticles (MPNs) had a relatively large size of ~ 170 nm at physiological pH which results in prolonged blood circulation and enhanced accumulation at the tumor site. But once extravasated into acidic tumor interstices, the increased solubility of PEG led to breakage of the supramolecular nanostructure and dissolution of MPNs to small-sized (< 20 nm) nanoparticles, promoting deep penetration and distribution in tumor tissues. Furthermore, MPNs exhibited not only an excellent stable nanostructure for antitumor doxorubicin (DOX) loading, but rapid dissociation of the nanostructure under an intracellular reductive environment. With the capacity of long blood circulation, deep tumor penetration and fast intracellular drug release, the DOX-loaded multi-pocket nanoparticles demonstrated superior antitumor activities against large 4T1 tumor (~ 250 mm3) bearing mice with reduced side effect. CONCLUSIONS: Our facile fabrication of multi-pocket nanoparticles provided a promising way in improving solid tumor penetration and achieving a great therapeutic efficacy.

Characterization and pharmacokinetic evaluation of microcomposite particles of alpha lipoic acid/hydrogenated colza oil obtained in supercritical carbon dioxide.

The work reported here is an extension of our previous findings in which supercritical composite particles (SCP) of alpha lipoic acid (ALA) masked with hydrogenated colza oil (HCO) named as ALA/HCO/SCP were obtained by the modified particles from gas-saturated solutions (PGSS) process in supercritical carbon dioxide in order to obscure the unpleasant taste and odor of ALA. The masking effect on ALA/HCO/SCP was compared with the widely used mechano-chemically masked formulation of ALA and HCO named as MC-50F. In the present study, ALA/HCO/SCP particles were found to have a significant improvement in regard to bitterness, numbness, and smell compared to ALA bulk powders suggesting they were well coated. The pharmacokinetic parameters for ALA/HCO/SCP and ALA bulk powder gave similar values but were significantly different from those of MC-50F. The amount of ALA absorbed into the body, in the administered ALA/HCO/SCP, was comparable to that absorbed by ALA bulk powder, whereas about half portion of ALA of the MC-50F was not absorbed, because the ALA/HCO/SCP particles were small enough and the particles of MC-50F were relatively large and had smaller specific surface area. Therefore, this study suggested a newly masked candidate may offer functional particles with maintained efficacy.

Lipoic Acid Stimulates cAMP Production in Healthy Control and Secondary Progressive MS Subjects.

Lipoic acid (LA) exhibits antioxidant and anti-inflammatory properties; supplementation reduces disease severity and T lymphocyte migration into the central nervous system in a murine model of multiple sclerosis (MS), and administration in secondary progressive MS (SPMS) subjects reduces brain atrophy compared to placebo. The mechanism of action (MOA) of LA's efficacy in suppression of MS pathology is incompletely understood. LA stimulates production of the immunomodulator cyclic AMP (cAMP) in vitro. To determine whether cAMP could be involved in the MOA of LA in vivo, we performed a clinical trial to examine whether LA stimulates cAMP production in healthy control and MS subjects, and whether there are differences in the bioavailability of LA between groups. We administered 1200 mg of oral LA to healthy control, relapsing remitting MS (RRMS) and SPMS subjects, and measured plasma LA and cAMP levels in peripheral blood mononuclear cells (PBMCs). There were no significant differences between the groups in pharmacokinetic (PK) parameters. Healthy and SPMS subjects had increased cAMP at 2 and 4 h post-LA treatment compared to baseline, while RRMS subjects showed decreases in cAMP. Additionally, plasma concentrations of prostaglandin E2 (PGE2, a known cAMP stimulator) were significantly lower in female RRMS subjects compared to female HC and SPMS subjects 4 h after LA ingestion. These data indicate that cAMP could be part of the MOA of LA in SPMS, and that there is a divergent response to LA in RRMS subjects that may have implications in the efficacy of immunomodulatory drugs. This clinical trial, "Defining the Anti-inflammatory Role of Lipoic Acid in Multiple Sclerosis," NCT00997438, is registered at https://clinicaltrials.gov/ct2/show/record/NCT00997438 .

Mutations in SLC5A6 associated with brain, immune, bone, and intestinal dysfunction in a young child.

The human sodium-dependent multivitamin transporter (hSMVT) is a product of the SLC5A6 gene and mediates biotin, pantothenic acid, and lipoate uptake in a variety of cellular systems. We report here the identification of mutations R94X, a premature termination, and R123L, a dysfunctional amino acid change, both in exon 3 of the SLC5A6 gene in a child using whole genome-scanning. At 15 months of age, the child showed failure to thrive, microcephaly and brain changes on MRI, cerebral palsy and developmental delay, variable immunodeficiency, and severe gastro-esophageal reflux requiring a gastrostomy tube/fundoplication, osteoporosis, and pathologic bone fractures. After identification of the SLC5A6 mutations, he responded clinically to supplemental administration of excess biotin, pantothenic acid, and lipoate with improvement in clinical findings. Functionality of the two mutants was examined by 3H-biotin uptake assay following expression of the mutants in human-derived intestinal HuTu-80 and brain U87 cells. The results showed severe impairment in biotin uptake in cells expressing the mutants compared to those expressing wild-type hSMVT. Live cell confocal imaging of cells expressing the mutants showed the R94X mutant to be poorly tolerated and localized in the cytoplasm, while the R123L mutant was predominantly retained in the endoplasmic reticulum. This is the first reporting of mutations in the SLC5A6 gene in man, and suggests that this gene is important for brain development and a wide variety of clinical functions.

Targeted systemic delivery of siRNA to cervical cancer model using cyclic RGD-installed unimer polyion complex-assembled gold nanoparticles.

For systemic delivery of small interfering RNA (siRNA) to solid tumors, we developed an actively-targeted unimer polyion complex-assembled gold nanoparticle (uPIC-AuNP) by a two-step assembling process. First is the monodispersed uPIC formation from the single molecules of therapeutic siRNA and the block catiomer, cyclic RGD (cRGD) peptide-installed poly(ethylene glycol)-block-poly(l-lysine) modified with lipoic acid (LA) at the ω-end (cRGD-PEG-PLL-LA). Second is the surface decoration of a 20nm-sized AuNP with uPICs. The cRGD-installed uPIC-AuNPs (cRGD-uPIC-AuNP) provided the targetability for selective binding to the cancer and cancer-related endothelial cellular surface, while regulating their size <50nm with a quite narrow distribution. The targeting efficacy of the cRGD-uPIC-AuNP was confirmed by in vitro cellular uptake in cultured cervical cancer (HeLa) cells and in vivo tumor accumulation in a subcutaneous HeLa model after systemic administration, compared with a non-targeted control uPIC-AuNP. Due to the targetability of the ligand, the cRGD-uPIC-AuNP achieved the significantly enhanced gene silencing ability in the subcutaneous HeLa tumor. Ultimately, the systemic delivery of siRNA targeted for papilloma virus-derived E6 oncogene by cRGD-uPIC-AuNP significantly inhibited the growth of subcutaneous HeLa tumor. This research demonstrates that the bottom-up construction of nanocarriers using monodispersed building blocks can be employed as delivery platforms for RNA interference-based cancer therapy.

Epidermal Growth Factor Receptor-Specific Nanoprobe Biodistribution in Mouse Models.

Nanotechnology offers a targeted approach to both imaging and treatment of cancer, the leading cause of death worldwide. Previous studies have found that nanoparticles with a wide variety of coatings initiate an immune response leading to sequestration in the liver and spleen. In an effort to find a nanoparticle platform which does not elicit an immune response, we created 43 nm and 44 nm of gold and silver nanoparticles coated with biomolecules normally produced by the body, α-lipoic acid and the epidermal growth factor (EGF), and have used mass spectroscopy to determine their biodistribution in mouse models, 24 h after tail vein injection. Relative to controls, mouse EGF (mEGF)-coated silver and gold nanoprobes are found at background levels in all organs including the liver and spleen. The lack of sequestration of mEGF-coated nanoprobes in the liver and spleen and the corresponding uptake of control nanoprobes at elevated levels in these organs suggest that the former are not recognized by the immune system. Further studies of cytokine and interleukin levels in the blood are required to confirm avoidance of an immune response.

The disulfide compound α-lipoic acid and its derivatives: A novel class of anticancer agents targeting mitochondria.

The endogenous disulfide α-lipoic acid (LA) is an essential mitochondrial co-factor. In addition, LA and its reduced counterpart dihydro lipoic acid form a potent redox couple with antioxidative functions, for which it is used as dietary supplement and therapeutic. Recently, it has gained attention due to its cytotoxic effects in cancer cells, which is the key aspect of this review. We initially recapitulate the dietary occurrence, gastrointestinal absorption and pharmacokinetics of LA, illustrating its diverse antioxidative mechanisms. We then focus on its mode of action in cancer cells, in which it triggers primarily the mitochondrial pathway of apoptosis, whereas non-transformed primary cells are hardly affected. Furthermore, LA impairs oncogenic signaling and displays anti-metastatic potential. Novel LA derivatives such as CPI-613, which target mitochondrial energy metabolism, are described and recent pre-clinical studies are presented, which demonstrate that LA and its derivatives exert antitumor activity in vivo. Finally, we highlight clinical studies currently performed with the LA analog CPI-613. In summary, LA and its derivatives are promising candidates to complement the arsenal of established anticancer drugs due to their mitochondria-targeted mode of action and non-genotoxic properties.

Gas-saturated solution process to obtain microcomposite particles of alpha lipoic acid/hydrogenated colza oil in supercritical carbon dioxide.

Alpha lipoic acid (ALA), an active substance in anti-aging products and dietary supplements, need to be masked with an edible polymer to obscure its unpleasant taste. However, the high viscosity of the ALA molecules prevents them from forming microcomposites with masking materials even in supercritical carbon dioxide (scCO2). Therefore, the purpose of this study was to investigate and develop a novel production method for microcomposite particles for ALA in hydrogenated colza oil (HCO). Microcomposite particles of ALA/HCO were prepared by using a novel gas-saturated solution (PGSS) process in which the solid-dispersion method is used along with stepwise temperature control (PGSS-STC). Its high viscosity prevents the formation of microcomposites in the conventional PGSS process even under strong agitation. Here, we disperse the solid particles of ALA and HCO in scCO2 at low temperatures and change the temperature stepwise in order to mix the melted ALA and HCO in scCO2. As a result, a homogeneous dispersion of the droplets of ALA in melted HCO saturated with CO2 is obtained at high temperatures. After the rapid expansion of the saturated solution through a nozzle, microcomposite particles of ALA/HCO several micrometers in diameter are obtained.

Therapeutic applications of lipoic acid: a patent review (2011 - 2014).

INTRODUCTION: Lipoic acid (LA), a naturally occurring 1,2-dithiolane analog that plays an essential role in mitochondrial bioenergetic reactions, has gained unprecedented attention as nutritional supplement and as therapeutic agent. Moreover, LA conjugates with other pharmacophores represent a promising approach toward the development of multifunctional drugs. AREAS COVERED: The reviewed patent applications from January 2011 to April 2014 include combinations of LA with other bioactive compounds as well as LA conjugates for the treatment of a wide range of clinical conditions. Additionally, some patents disclose methods to overcome the stability problems of LA. EXPERT OPINION: LA is currently in clinical use for the treatment of diabetic neuropathy, while small clinical trials using combinations of LA with known bioactive agents have been undertaken. The use of the LA is hampered by its instability and its rapid metabolism. Thus, formulations containing LA, in a form ensuring its stability and improving its bioavailability, can have important applications as medicines, nutritional supplements or cosmeceuticals. LA hybrids with other pharmacophores endowed with various activities, possess an enormous potential to promote human health and have been the subject of numerous publications and patent applications. Nevertheless, reliable in vivo data and clinical trials are necessary to prove these beneficial effects.

(R)-α-lipoyl-glycyl-L-prolyl-L-glutamyl dimethyl ester codrug as a multifunctional agent with potential neuroprotective activities.

The (R)-α-lipoyl-glycyl-L-prolyl-L-glutamyl dimethyl ester codrug (LA-GPE, 1) was synthesized as a new multifunctional drug candidate with antioxidant and neuroprotective properties for the treatment of neurodegenerative diseases. Physicochemical properties, chemical and enzymatic stabilities were evaluated, along with the capacity of LA-GPE to penetrate the blood-brain barrier (BBB) according to an in vitro parallel artificial membrane permeability assay for the BBB. We also investigated the potential effectiveness of LA-GPE against the cytotoxicity induced by 6-hydroxydopamine (6-OHDA) and H2O2 on the human neuroblastoma cell line SH-SY5Y by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assay. Our results show that codrug 1 is stable at both pH 1.3 and 7.4, exhibits good lipophilicity (log P=1.51) and a pH-dependent permeability profile. Furthermore, LA-GPE was demonstrated to be significantly neuroprotective and to act as an antioxidant against H2O2- and 6-OHDA-induced neurotoxicity in SH-SY5Y cells.

Age and gender dependent bioavailability of R- and R,S-α-lipoic acid: a pilot study.

Lipoic acid (LA) shows promise as a beneficial micronutrient toward improving elder health. Studies using old rats show that (R)-α-LA (R-LA) significantly increases low molecular weight antioxidants that otherwise decline with age. Despite this rationale for benefiting human health, little is known about age-associated alterations in absorption characteristics of LA, or whether the commercially available racemic mixture of LA (R,S-LA) is equally as bioavailable as the naturally occurring R-enantiomer. To address these discrepancies, a pilot study was performed to establish which form of LA is most effectively absorbed in older subjects relative to young volunteers. Young adults (average age=32 years) and older adults (average age=79 years) each received 500 mg of either R- or R,S-LA. Blood samples were collected for 3h after supplementation. After a washout period they were given the other chiral form of LA not originally ingested. Results showed that 2 out of 6 elder males exhibited greater maximal plasma LA and area under the curve for the R-form of LA versus the racemic mixture. The elder subjects also demonstrated a reduced time to reach maximal plasma LA concentration following R-LA supplementation than for the racemic mixture. In contrast, young males had a tendency for increased bioavailability of R,S-LA. Overall, bioavailability for either LA isoform was much more variable between older subjects compared to young adults. Plasma glutathione levels were not altered during the sampling period. Thus subject age, and potential for varied response, should be considered when determining an LA supplementation regimen.

Lipoic acid nanoparticles: effect of polymeric stabilizer on appetite suppression.

Alpha-lipoic acid (ALA), which is common in the human body, is efficacious in appetite suppression. However, its typical formulations of salt or micronized crystals cannot satisfy the desired bioavailability requirements for appetite suppression due to low absorption and a short plasma half-life. Herein, we describe a new ALA nanoparticulate formulation produced by nano-comminution using polymeric stabilizers, such as hydroxypropyl cellulose, Pluronic F127, and polyvinylpyrrolidone. Nanoparticles of similar sizes did not show any remarkable differences in the in vitro release profiles. However, the in vivo results from food intake studies in mice demonstrated that the hydroxypropyl cellulose case had the largest improved efficacy among the three polymeric stabilizer cases. Compared to the nanosuspension formulations, the powder formulations of nanoparticles had improved efficacy in reducing food intake for six hours, possibly because of the delayed release kinetics. Therefore, the ALA powder formulation of nanoparticles is a candidate to replace the current formulations to achieve proper appetite suppression.

Study of alpha-lipoic acid penetration in the human aqueous after topical administration.

PURPOSE: To determine the concentration of alpha-lipoic acid in the aqueous and investigate if its topical instillation can increase quantities. METHODS: A total of 70 patients selected to undergo cataract surgery were randomly divided into two groups. Group 1 was used as a control group; for the patients in Group 2, a single instillation of alpha-lipoic acid eye drops (1%) was administered. Immediately before surgery, an aliquot of 40-120 microL of aqueous was aspirated. The individual aspirations were combined to constitute pools representing time intervals with respect to administration. The levels of alpha-lipoic acid in the aqueous were measured using gas chromatography/mass spectrometry. RESULTS: Pool 0 was created by combining the samples of aqueous obtained from the patients in Group 1, the control group; and the level of alpha-lipoic acid was 27.5 + 2.6 ng/mL. In the other pools, the time interval between the administration of the eye drops and sampling was, respectively, 23 min, 53 min, 72 min, 93 min and 114 min; and the level of alpha-lipoic acid was 33.0 + 10.8 ng/mL, 52.0 + 2.5 ng/mL, 86.7 + 2.5 ng/mL, 91.2 + 2.5 ng/mL and 80.3 + 2.5 ng/mL. CONCLUSION: Our study demonstrates the presence of alpha-lipoic acid in the aqueous and indicates that its concentration increases after it is administered in the form of eye drops, reaching maximum values after around 93 min. The concentrations that are achieved in the anterior chamber allow us to theorize the possibility of exploiting the antioxidant properties of alpha-lipoic acid.

Antioxidant properties of an endogenous thiol: Alpha-lipoic acid, useful in the prevention of cardiovascular diseases.

In the past few years, a growing interest has been given to the possible antioxidant functions of a natural acid, synthesized in human tissues: alpha-lipoic acid (ALA). Both the oxidized (disulfide) and reduced (dithiol: dihydrolipoic acid, DHLA) forms of ALA show antioxidant properties. ALA administered in the diet accumulates in tissues, and a substantial part is converted to DHLA via a lipoamide dehydrogenase. Commercial ALA is usually a racemic mixture of the R and S forms. Chemical studies have indicated that ALA scavenges hydroxyl radicals, hypochlorous acid, and singlet oxygen. ALA exerts antioxidant effects in biological systems not only through direct ROS quenching but also via transition metal chelation. ALA has been shown to possess a number of beneficial effects both in the prevention and treatment of diabetes in experimental conditions. ALA presents beneficial effects in the management of symptomatic diabetic neuropathy and has been used in this context in Germany for more than 30 years. In cardiovascular disease, dietary supplementation with ALA has been successfully employed in a variety of in vivo models: ischemia-reperfusion, heart failure, and hypertension. More mechanistic and human in vivo studies are needed to determine whether optimizing the dietary intake of ALA can help to decrease cardiovascular diseases. A more complete understanding of cellular biochemical events that influence oxidative damage is required to guide future therapeutic advances.

Uptake and reduction of alpha-lipoic acid by human erythrocytes.

OBJECTIVES: The reducing capacity of erythrocytes has been used clinically as to estimate resistance to oxidant stress. In this work we targeted the antioxidant capacity of pyridine nucleotide disulfide reductases of these cells by measuring their ability to reduce the disulfide alpha-lipoic acid. METHODS: Erythrocyte reduction of alpha-lipoic acid and related disulfides was measured as reduction of 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) outside the cells. RESULTS: Lipoic acid-dependent DTNB reduction by human erythrocytes required d-glucose and consumed NADPH, but not NADH. This activity was inhibited by carmustine and phenylarsine oxide, as expected if alpha-lipoic acid is reduced by the glutathione and thioredoxin reductase systems. Reduction of hydroxyethyl disulfide, which provides an estimate of total erythrocyte disulfide reduction capacity, was similar to that of alpha-lipoic acid. Erythrocytes incubated with alpha-lipoic acid also reduced extracellular ferricyanide, although rates of dehydroascorbate reduction were several-fold greater, probably because intracellular GSH can recycle ascorbate but not alpha-lipoic acid in erythrocytes. CONCLUSION: These results show that alpha-lipoic acid-dependent DTNB reduction provides a simple method to selectively assess the capacity of pyridine nucleotide disulfide reductases of human erythrocytes. When coupled with other non-destructive assays, such as reduction of hydroxyethyl disulfide and ferricyanide, this assay provides a comprehensive approach to assessing erythrocyte reducing capacity in a variety of clinical conditions associated with oxidant stress.

Bioavailability of selenium from the selenotrisulphide derivative of lipoic acid.

BACKGROUND/PURPOSE: Selenium is a required micronutrient in mammals, needed for the activity of enzymes that contain selenocysteine at their active site. Several isoenzymes of glutathione peroxidase and thioredoxin reductase contain selenocysteine and thus the nutritional status of selenium in tissues can have significant impact on the steady state level of reactive oxygen species. The aims of this study were to evaluate the bioavailability of selenium derived from the selenotrisulfide derivative of lipoic acid (LASe) and determine the ability of this compound to be absorbed into skin. METHODS: Bioavailability of selenium derived from LASe was determined using a keratinocyte cell model (HaCat). Efficiency of utilization of selenium was assessed by following the decrease in the incorporation of radiolabeled selenite (75Se) in the presence of increasing concentration of selenium compounds. Percutaneous absorption of LASe was measured by determining selenium levels in full thickness biopsy of skin using a Yorkshire pig model. RESULTS: LASe was efficiently absorbed topically into pig skin, a good model of human skin. In a keratinocyte cell line LASe was an efficient source of selenium for selenoprotein synthesis, demonstrating that LASe is a good candidate as a topical selenium micronutrient. Both L-selenomethionine and selenate were found to be poor sources of selenium for selenoprotein synthesis in the skin cell model and L-selenomethionine was poorly absorbed into pig skin. CONCLUSION: These results indicate that stable selenotrisulfides, such as LASe, are good candidates for testing as topical selenium supplements.

Inhibition of oxygen radical formation by methylene blue, aspirin, or alpha-lipoic acid, prevents bacterial-lipopolysaccharide-induced fever.

Phagocytic cells contain NADPH oxidase that they use for host defense by catalyzing the production of superoxide. Bacterial lipopolysaccharide (LPS) has been found to stimulate NADPH oxidase in mobile and sessile macrophages and microglia. It also evokes fever in homeothermic animals and men, a reaction mediated by central nervous system (CNS) activities. The purpose of the present study was to determine whether reactive oxygen species are involved in LPS-induced fever. In rabbits we found that plasma hydroperoxide levels increased and catalase activity decreased 15 min after LPS injection and that fever started with a similar latency, while plasma levels of tumor necrosis factor-alpha (TNFalpha) increased 30 min after the injection. Treating rabbits with methylene blue or aspirin did not affect TNFalpha secretion but prevented the LPS-induced rise of hydroperoxides and the inactivation of catalase, abolishing fever. Incubation of human blood with nitroblue tetrazolium and LPS increased the number of formazan-positive neutrophils from 10 +/- 5 to 52 +/- 9%. Adding LPS to blood preincubated with either methylene blue, alpha-lipoic acid, or aspirin respectively decreased the number of formazan-positive neutrophils to 0.9 +/- 0.8, 0.8 +/- 0.9, or 2.0 +/- 0.9%, disclosing the antioxidant capacity of these drugs. Systemic application of 80 mg/kg alpha-lipoic acid elicited heat-loss reactions within 15 min and decreased core temperature by 2.2 +/- 0.3 degrees C within 2 h. Alpha-lipoic acid applied 45 min after LPS induced antipyresis within 15 min, and this antipyresis was associated with a decrease of elevated hydroperoxide levels and restoration of catalase activity. Our results show that fever is prevented when the production of reactive oxygen species is blocked and that an elevated body temperature returns to normal when oxygen radical production decreases. Estimation of plasma dihydrolipoic acid (DHLA) levels following injection of 80 mg/kg alpha-lipoic acid in afebrile and febrile rabbits revealed that this acid is converted into DHLA, which in afebrile rabbits increased the plasma DHLA concentration from 2.22 +/- 0.26 microg/ml to peak values of 8.60 +/- 2.28 microg/ml DHLA within 30 min and which in febrile rabbits increased it from 0.84 +/- 0.22 microg/ml to peak values of 3.90 +/- 0.94 microg/ml within 15 min. Methylene blue, aspirin, and alpha-lipoic acid, which all cross the blood-brain barrier, seem to act not only on peripheral tissues but also on the CNS. Brain structures that have been shown to sense oxidative stress are vicinal thiol groups attached to the NMDA subtype of glutamate receptor. Their reduction by thiol-reducing drugs like dithiothreitol or DHLA has been found to increase glutamate-mediated neuronal excitability, while the opposite effect has been observed after their oxidation. Because we found that systemic application of alpha-lipoic acid in the afebrile state elicits hypothermia and in the febrile state is antipyretic, we think this type of NMDA receptor is involved in thermoregulation and that oxidation of its thiol groups induces fever. It appears that temperature homeostasis can be maintained only if the redox homeostasis of the brain is guaranteed.

Photoprotection by antioxidants against UVB-radiation-induced damage in pig skin organ culture.

Topically applied antioxidants constitute an important group of protective agents against skin damage induced by ultraviolet radiation. The current study was performed to investigate whether a recently developed ex vivo pig skin model was suitable for short-term studies of the mechanism(s) of UVB-radiation-induced skin damage; the protective effect of topical application of alpha-tocopherol, l-ascorbic acid, alpha-lipoic acid, glutathione ethylester and N-acetylcysteine was tested. Increasing doses of the antioxidants were applied topically on ex vivo pig skin explants and allowed to penetrate for 60 min. Epidermal antioxidant bioavailability was measured before and 60 min after exposure to an ultraviolet B (UVB) radiation of 7.5 kJ/m2. Cell viability (trypan blue dye exclusion) and apoptosis were measured 48 h later in isolated keratinocytes. UVB-radiation-induced epidermal lipid peroxidation was determined immediately after exposure of the skin to a UVB dose of 28 kJ/m2. All antioxidants tested became bioavailable in pig skin epidermis, and none of them were depleted after UVB-radiation exposure. Increasing doses of the antioxidants tested decreased UVB-radiation-induced cell death and apoptosis. The highest doses of antioxidants prevented UVB-radiation-induced lipid peroxidation; alpha-lipoic acid only tended to decrease lipid peroxidation. In conclusion, a single topical dose of the above antioxidants on ex vivo pig skin can reduce UVB-radiation-induced oxidative stress and lipid peroxidation and thereby reduce apoptotic stimuli and cell death. Furthermore, the ex vivo pig skin model was a useful tool for testing compounds for their antioxidant activity.

Kinetic study of cutaneous and subcutaneous distribution following topical application of [7,8-14C]rac-alpha-lipoic acid onto hairless mice.

To diminish oxidative injury, topically applied antioxidants must reach susceptible cells. alpha-Lipoic acid is a potent thiol antioxidant that might be useful for skin protection; therefore, its skin penetration kinetics were assessed. The cutaneous and subcutaneous distributions of [7,8-14C]rac-alpha-lipoic acid were studied in anesthetized hairless mice after application of a 5% solution in propylene glycol for 0.5 to 4 hr. The mice were killed; then the skin was washed, and the stratum corneum was removed by 10 cellophane tape strippings. A punch biopsy of the frozen, stripped skin was sectioned, and amounts of [14C]-alpha-lipoic acid were determined in strippings and slices of epidermis (4 x 5 microns), dermis, and subcutaneous fat (10 x 10 microns, 20 x 20 microns). The rate of [14C]-alpha-lipoic acid absorption into skin was constant by 30 min (0.10 +/- 0.01 nmol/cm2/min); maximum skin concentrations were reached by 2 hr. The [14C]-alpha-lipoic acid penetration kinetics into the first layer of the stratum corneum predicted its penetration through the stratum corneum and subsequent percutaneous absorption (r2 = 0.96, P < 0.02). Cutaneous absorption of unlabeled alpha-lipoic acid and its reduction to the more potent antioxidant form, dihydrolipoic acid, were also demonstrated, using HPLC analysis with electrochemical detection. In conclusion, alpha-lipoic acid topically applied to skin penetrated readily, and was reduced to dihydrolipoic acid. Thus, alpha-lipoic acid could potentiate skin antioxidant protection.

Design, synthesis, and pharmacokinetic evaluation of a chemical delivery system for drug targeting to lung tissue.

We espouse the application of a novel chemical delivery system (CDS) approach to a delivery mechanism for drug targeting to lung tissue using the 1,2-dithiolane-3-pentyl moiety of lipoic acid as the "targetor moiety". The synthesis and the physicochemical and pharmacokinetic evaluation of a CDS modeling the lipoyl and other ester derivatives of chlorambucil (an antineoplastic agent) and cromolyn (a bischromone used in antiasthma prophylaxis) as compared with their respective parent drugs are described. The chlorambucil CDS was synthesized by esterifying the alcohol derivative of lipoic acid with chlorambucil using dicyclohexylcarbodiimide as the coupling agent. The cromolyn CDS was prepared by a multistep synthetic procedure culminating in the reaction of the alkyl bromide derivative of lipoic acid with the disodium salt of the bischromone compound. All the esters were highly lipophilic unlike the parent compounds. The in-vitro kinetic and in-vivo pharmacokinetic studies showed that the respective CDSs were sufficiently stable in buffer and biological media, hydrolyzed rapidly into the respective active parent drugs, and significantly enhanced delivery and retention of the active compound to lung tissue in comparison with the underivatized parent compounds used in conventional therapy.