Clinical Pharmacokinetics and Safety of a 10% Aminolevulinic Acid Hydrochloride Nanoemulsion Gel (BF-200 ALA) in Photodynamic Therapy of Patients Extensively Affected With Actinic Keratosis: Results of 2 Maximal Usage Pharmacokinetic Trials.

The nanoemulsion-based 10% aminolevulinic acid (ALA) hydrochloride gel BF-200 ALA optimizes epidermal penetration of its active ingredient and is approved for topical photodynamic therapy (PDT) for the treatment of actinic keratosis in the United States and Europe. To characterize systemic absorption from dermal application during PDT, ALA and its key active metabolite protoporphyrin IX (PpIX) were analyzed in 2 maximal usage pharmacokinetic trials (MUsT) in patients severely affected with actinic keratosis. The primary objective of both MUsTs was to assess baseline-adjusted plasma concentration-time curves for ALA and PpIX after a single PDT treatment applying either 2 g (1 tube) of BF-200 ALA on the face (MUsT-1) or applying 6 g (3 tubes) of BF-200 ALA on the face/scalp or body periphery (MUsT-2), to 20 or 60 cm2 , respectively. All PDTs were performed using red light at around 635 nm wavelength. Safety and tolerability were documented along with pharmacokinetics. In both MUsTs, ALA plasma concentrations were transiently increased to a maximum concentration at about 2.5 to 3.3 times above endogenous baseline with time to maximum concentration at ≈3 hours after dosing. Plasma levels subsequently returned to baseline within 10 hours after dosing. Overall baseline-adjusted mean area under the baseline-adjusted plasma concentration-time curve from time zero to the last sampling time point at which the concentration was at or above the lower limit of quantification ranged from 142.8 to 146.2, indicating that a similar, minor fraction of topical ALA is systemically absorbed under both dosing regimens. Systemic PpIX exposure after administration of either dose of BF-200 ALA was equally minimal. Application site skin reactions were treatment area size-related, albeit transient and consistent with the known safety profile of BF-200 ALA.

Differential expression of Cathepsin S and X in the spinal cord of a rat neuropathic pain model.

BACKGROUND: Ample evidence suggests a substantial contribution of cellular and molecular changes in the spinal cord to the induction and persistence of chronic neuropathic pain conditions. While for a long time, proteases were mainly considered as protein degrading enzymes, they are now receiving growing interest as signalling molecules in the pain pathology. In the present study we focused on two cathepsins, CATS and CATX, and studied their spatiotemporal expression and activity during the development and progression of neuropathic pain in the CNS of the rat 5th lumbar spinal nerve transection model (L5T). RESULTS: Immediately after the lesion, both cathepsins, CATS and CATX, were upregulated in the spinal cord. Moreover, we succeeded in measuring the activity of CATX, which was substantially increased after L5T. The differential expression of these proteins exhibited the same spatial distribution and temporal progression in the spinal cord, progressing up to the medulla oblongata in the late phase of chronic pain. The cellular distribution of CATS and CATX was, however, considerably different. CONCLUSION: The cellular distribution and the spatio-temporal development of the altered expression of CATS and CATX suggest that these proteins are important players in the spinal mechanisms involved in chronic pain induction and maintenance.

BF-1--a novel selective 5-HT2B receptor antagonist blocking neurogenic dural plasma protein extravasation in guinea pigs.

Serotonin 5-HT2B receptor antagonists have been proposed as migraine prophylactic drugs, but previously available 5-HT2B receptor antagonists displayed multiple monoaminergic side effects and had to be withdrawn from the market. Here, we set out to identify a novel antagonist with high affinity and selectivity towards 5-HT2B receptors. To test the affinity of new compounds towards various receptors, we generated a broad series of cells functionally coupling human monoaminergic receptors to luciferase. Using the cell lines we revealed pimethixene (1-methyl-4-(9H-thioxanthen-9-ylidene)piperidine) as highly potent, albeit non-selective 5-HT2B receptor antagonist and optimized its chemical structure to create highly potent and selective 5-HT2B receptor antagonists. We selected the methoxythioxanthene BF-1 for further analysis. In comparison to pimethixene, it lacked high affinities to 5-HT1A, 5-HT2A, 5-HT2C, histamine H1, dopamine D1 and D2 as well as muscarinic M1 and M2 receptors. BF-1 was tested as potential migraine prophylactic drug by blocking meta-chlorophenylpiperazine, (mCPP) or BW723C86 (5-((thiophen-2-yl)methoxy)-α-methyltryptamine) induced neurogenic dural plasma protein extravasation in a guinea pig model that may resemble a migraine attack. BF-1 was significantly more potent in this assay compared to the well know non-selective 5-HT2B antagonists, methysergide ((6aR,9R)-N-[(2S)-1-Hydroxybutan-2-yl]-4,7-dimethyl-6,6a,8,9-tetrahydroindolo[4,3-fg]quinoline-9-carboxamide) or pizotifen (4-(1-methyl-4-piperidylidine)-9,10-dihydro-4H-benzo-[4,5]cyclohepta[1,2]-thiophene). Therefore, we propose BF-1 as a new compound that may be developed for prophylactic migraine treatment without the typical monoaminergic side effects.

Sgk1, a cell survival response in neurodegenerative diseases.

Serum and glucocorticoid-regulated kinase 1 (sgk1) belongs to a family of serine/threonine kinases that is under acute transcriptional control by serum and glucocorticoids. An expanding set of receptors and cellular stress pathways has been shown to enhance sgk1 expression, which is implicated in the regulation of ion channel conductance, cell volume, cell cycle progression, and apoptosis. Recent evidence for the involvement of sgk1 in the early pathogenesis of MPTP-induced Parkinson's disease (PD) prompted us to investigate in more detail its expression and role in animal models of different neurodegenerative diseases. Here, we show that transcription of sgk1 is increased in several animal models of PD and a transgenic model of amyotrophic lateral sclerosis (ALS). The upregulation of sgk1 strongly correlates with the occurrence of cell death. Furthermore, we provide evidence that the Forkhead transcription factor FKHRL1 and some of the voltage-gated potassium channels are physiological substrates of sgk1 in vivo. Using a small interfering RNA approach to silence sgk1 transcripts in vitro, we give evidence that sgk1 exerts a protective role in oxidative stress situations. These findings underline a key role for sgk1 in the molecular pathway of cell death, in which sgk1 seems to exert a protective role.

Dorso-ventral polarity of the zebrafish embryo is distinguishable prior to the onset of gastrulation.

We describe a set of observations on developing zebrafish embryos and discuss the main conclusions they allow:(1) the embryonic dorso-ventral polarity axis is morphologically distinguishable prior to the onset of gastrulation; and (2) the involution of deep layer cells starts on the prospective dorsal side of the embryo. An asymmetry can be distinguished in the organization of the blastomeres in the zebrafish blastula at the 30% epiboly stage, in that one sector of the blastoderm is thicker than the other. Dye-labelling experiments with DiI and DiO and histological analysis allow us to conclude that the embryonic shield will form on the thinner side of the blastoderm. Therefore, this side corresponds to the prospective dorsal side of the embryo. Simultaneous injections of dyes on the thinner side of the blastoderm and on the opposite side show that involution of deep layer cells during gastrulation starts at the site at which the embryonic shield will form and extends from here to the prospective ventral regions of the germ ring.

Neurulation in the anterior trunk region of the zebrafish Brachydanio rerio.

We have studied the process of neurulation within the anterior trunk region of the zebrafish by means of serial sectioning of staged embryos and labelling cells by applications of the dye Dil and intracellular injections of fluoresceine dextran amine. The first morphological manifestation of the prospective neural plate is a dorsomedial ectodermal thickening which becomes visible immediately after gastrulation. Within 1-2 h, by the time somatogenesis begins, two bilaterally symmetrical thickenings have appeared more laterally, which eventually fuse with the medial thickening to form the neural keel. The central canal forms next by separation of the cells on either side of the midline of the neural keel, beginning ventrally at the 17-somite stage and progressing towards dorsal levels. By means of fluorescent dye labelling in the late gastrula, we found that both the medial and lateral thickenings contribute to the nerve cord. The medial thickening was found to contain, exclusively, neural progenitor cells from the 90-100% epiboly stage on, whereas the adjacent regions contained a mixture of neural and epidermal progenitor cells, as well as prospective neural crest cells. Between the 90-100% epiboly and 2-somite stages, this heterogeneity of developmental capabilities is resolved into territories, with epidermogenic and neurogenic cells clearly separated from each other. To achieve this segregation into neural and epidermal anlagen, cells from the lateral thickenings have to move over a distance of roughly 400 µm within 1-2 h. Epidermal overgrowth of the nerve cord occurs during the morphogenetic movements that accompany nerve cord formation.

Ecdysone and a dietary alkaloid interact in the development of the pheromone gland of a male moth (Creatonotos, Lepidoptera: Arctiidae).

Hair-covered scent organs of the male arctiid moth Creatonotos produce and dissipate the volatile pheromone hydroxydanaidal. The biosynthesis of this substance depends quantitatively upon the uptake of pyrrolizidine alkaloids (PA) with the larval foodplant. The size of the tubular, eversible scent organ (corema) is also positively correlated with the ingested amount of the same alkaloid, which acts like a specific growth factor. After an assessment of the corema normogenesis by Rick-Wagner (PhD thesis, University of Cologne, 1986) we injected PA into PA-free raised larvae, prepupae, and pupae. We found that the PA competence (sensitivity) of the corema anläge terminates with the first prepupal day. Ecdysone titer determinations (radioimmunoassay) are in agreement with those in other moth species. Ligated (ecdysone-free) pupal abdomina never developed imaginal structures, with or without earlier PA application. Ecdysone injection into ligated pupal abdomina of PA-fed specimens initiated the development of imaginal structures and also of coremata of more than (ecdysone-free) control size. Pupal abdomina without PA pre-treatment only developed very small coremata. With these experiments we have separated and identified two morphogenetic control agents of corema development: the dietary PA specifies the size of the organ whereas ecdysone induces the anlage to proliferate within these PA-dependent ranges and to differentiate adult structures, as it does with other imaginal anlagen.