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Automated sediment toxicity testing and biomonitoring has grown rapidly. This study tested the suitability of the marine amphipod Corophium volutator (Pallas, 1766) for sediment biomonitoring using the Multispecies Freshwater Biomonitor (MFB). Two experiments were undertaken to (1) characterize individual behaviors of C. volutator using the MFB and (2) examine behavioral changes in response to sediment spiked with the pesticide Bioban. Four behaviors were visually identified (walking, swimming, grooming and falling) and characterized in the MFB as different patterns of locomotor activity (0-2 Hz range). Ventilation was not visually observed but was detected by the MFB (2-8 Hz). No clear diel activity patterns were detected. The MFB detected an overall increase in C. volutator locomotor activity after Bioban addition to the sediments (56, 100, 121 mg kg(-1)). C. volutator was more active (both locomotion and ventilation) in the water column than the spiked sediment. C. volutator appears a sensitive and appropriate species for behavioral sediment toxicity assessment and biomonitoring.

The challenge of predicting the metabolism or toxicity of a drug in humans has been approached using in vivo animal models, in vitro systems, high throughput genomics and proteomics methods, and, more recently, computational approaches. Understanding the complexity of biological systems requires a broader perspective rather than focusing on just one method in isolation for prediction. Multiple methods may therefore be necessary and combined for a more accurate prediction. In the field of drug metabolism and toxicology, we have seen the growth, in recent years, of computational quantitative structure-activity relationships (QSARs), as well as empirical data from microarrays. In the current study we have further developed a novel computational approach, MetaDrug, that 1) predicts metabolites for molecules based on their chemical structure, 2) predicts the activity of the original compound and its metabolites with various absorption, distribution, metabolism, excretion, and toxicity models, 3) incorporates the predictions with human cell signaling and metabolic pathways and networks, and 4) integrates networks and metabolites, with relevant toxicogenomic or other high throughput data. We have demonstrated the utility of such an approach using recently published data from in vitro metabolism and microarray studies for aprepitant, 2(S)-((3,5-bis(trifluoromethyl)benzyl)-oxy)-3(S)phenyl-4-((3-oxo-1,2,4-triazol-5-yl)methyl)morpholine (L-742694), trovofloxacin, 4-hydroxytamoxifen, and artemisinin and other artemisinin analogs to show the predicted interactions with cytochromes P450, pregnane X receptor, and P-glycoprotein, and the metabolites and the networks of genes that are affected. As a comparison, we used a second computational approach, MetaCore, to generate statistically significant gene networks with the available expression data. These case studies demonstrate the combination of QSARs and systems biology methods.

Between the stereoisomers of amide-type local anaesthetics differences have been noticed with respect to pharmacokinetics and side effects, but not regarding local anaesthetic capacity. Therefore, only S-(-)-ropivacaine has been introduced into clinical practice and with bupivacaine both the racemate and the S-(-)-enantiomer (levobupivacaine) are available by now. Based on this background, the aim of the present study was to evaluate if there are also dissimilarities to be found both in the toxicity and in the effectiveness of the enantiomers of two newly synthesized chiral fomocaine alkylmorpholine derivatives, OW3 and OW13, with an additional C2-chain in 2- or an additional C3-chain in 3-position at the morpholine ring, respectively. For this purpose, in vitro the interaction capacity with cytochrome P450 (CYP)-mediated monooxygenase and oxidase functions was investigated using rat liver 9000 g supernatants or microsomes. In vivo LD50, paresis of the N. ischiadicus and surface and conduction anaesthesia (cornea, N. ischiadicus) were tested in rats. The enantiomers of both OW3 and OW13 caused a concentration dependent inhibition of all CYP-mediated model reactions investigated. With all model reactions the (-)-enantiomer of OW3 was less effective than the (+)-form, whereas the opposite was the case with OW13. Also toxicity was lower with the (-)-enantiomer of OW3 and with the (+)-form of OW13 than with the respective counterparts. With both derivatives no clear-cut dissimilarities were noticed in the local anaesthetic capacity of the enantiomers. None of the four compounds caused paresis. Thus, similar to amide-type local anaesthetics, also with the enantiomers of chiral fomocaine alkylmorpholine derivatives differences in pharmacokinetic properties and toxicity could be demonstrated.

Fomocaine (CAS 56583-43-8) is a basic ether-type local anaesthetic used in dermatological practice for surface anaesthesia. For many years, modifications of the fomocaine molecule have been pursued, e.g. to improve its affinity to the sodium channel and also in view of possible new (systemic) applications. In the present study fomocaine and eight fomocaine derivatives with an additional alkyl chain in 2- or 3-position of different length (C1 up to C4), or with a branched C3 chain in 3-position, respectively, at the morpholine ring were evaluated in vitro for possible structure-activity relationships with respect to the interactions with cytochrome P450 (CYP) mediated monooxygenase and oxidase functions using rat liver 9000 g supernatants or microsomes. Results were compared to in vivo data from rats on toxicity (LD50), paresis of the N. ischiadicus and surface and conduction anaesthesia (cornea, N. ischiadicus). In general, the influence of the derivatives on the CYP system was less than that of fomocaine, showing a further decline with enlarging chain length. Toxicity of the derivatives was comparable to that of fomocaine and lower only with the compound with a C4 alkyl chain in 2-position. The derivatives caused a stronger surface anaesthesia than fomocaine, exhibiting an additional increase with enlarging chain length. No clear-cut structure-activity relationships were observed with respect to paresis of the N. ischiadicus and to conduction anaesthesia. Especially the derivatives having a C2 or C4 chain in 2- or a C3 chain in 3-position, respectively, may be of interest for further investigations. In comparison to fomocaine they caused a stronger surface anaesthesia combined with a lower interaction capacity with the CYP system.

Although the precise role for the tuberous sclerosis complex-2 tumor suppressor gene (Tsc2) in tumor suppression is not clear, many studies have implicated Tsc2 in the regulation of cell differentiation, cell cycle control, GTPase activity, transcription, polycystin-1 localization, and translation initiation. We propose that Tsc2 also increases susceptibility to apoptosis, and that this functional role may contribute to the tumor suppressor activity of Tsc2. We previously characterized the apoptotic response of a Tsc2-null renal tumor cell line (ERC-18) to the tumor promoter okadaic acid (OKA). In the present study, we expressed Tsc2 in ERC-18 cells and compared the effect of Tsc2 expression on apoptotic induction. Tsc2 expression increased the susceptibility of ERC-18 cells to apoptosis induced by OKA and the phosphatidylinositol-3' kinase inhibitor, LY294002. In addition, Tsc2 expression abrogated OKA-induced cell detachment of ERC-18 cells. These results indicate that the OKA-induced, caspase-independent detachment previously observed in ERC-18 cells is Tsc2-dependent, and may support an additional role for the Tsc2 in regulating cell adhesion.

New derivatives of daunorubicin (DRB), doxorubicin (DOX), and epidoxorubicin (EDOX) with an amidine group bonded to C-3' of daunosamine moiety with either morpholine or hexamethyleneimine ring attached to the amidine group are studied in this paper. We have shown that all of these newly synthesized anthracycline derivatives inhibit human leukemia K562 cell line proliferation but only some of them induce erythroid differentiation when used at subtoxic concentrations. Morpholine derivative of DOX has the greatest potential to inhibit proliferation and to induce differentiation in vitro. The correlation between these two cellular processes was also significant for other tested compounds. In cell cycle analysis, we have demonstrated that those anthracycline derivatives that exert the greatest cytostatic potential caused G(2)/M arrest, which in turn, might contribute to the development of a differentiating phenotype. The concentrations of the compounds used in the study are pharmacologically relevant. These new potent inducers of differentiation might be exploited as anticancer drugs for treatment of leukemia by differentiation therapy.

The effect on hepatic drug metabolising enzymes was evaluated for three representative structures and that were selected from a series of substituted oxazine derivatives designed to possess particular pharmacological properties such as analgesic, antioxidant and hypolipidemic activity. In addition, since xenobiotic metabolism, reactive oxygen and nitrogen species, atherosclerosis and inflammation are interrelated and mutually affected, the effects of and on acute inflammation in vivo and lipoxygenase activity in vitro were also investigated. It was found that treatment of rats with caused induction of cytochrome P450, enhancement of the metabolism of aminopyrine in vitro and of zoxazolamine and hexobarbital in vivo. Compound appeared to induce particularly erythromycin N-demethylation, while, a nitric ester, reduced the catalytically active cytochrome P450, although it increased the metabolism of specific cytochrome P450 substrates, i.e. 4-nitrophenol and erythromycin. Compounds and with strong hypolipidemic and antioxidant properties, reduced acute inflammatory response in two inflammation models and inhibited lipoxygenase activity in vitro. These results are helpful in optimising the biological profile as well as the potential applications of substituted oxazines.

Pesticides represent an important threat for natural populations. While their effects are assessed on short terms acute exposure, some of their harmful consequences may only become apparent when combined with other stressors, notably natural ones, such as predation. Here, we investigated in a laboratory experiment how exposure to a common fungicide (fenpropimorph) would affect the responses to predation in the common frog Rana temporaria. The concentrations of fungicide we used were comparable to those found in nature (0, 2, or 11 microg/L). The higher concentration of fungicide reduced tadpole activity late in the experiment, and only 7% of the tadpoles reached metamorphosis. In the lower concentration, the ability to respond adaptively to predator presence was not affected, but the costs (delayed metamorphosis, smaller relative body size) of this response were increased. Our results highlightthe need to investigate sublethal effects of pesticides on organismal performance if assessment of pesticides real impact is to be obtained.

Forward genetic screens in zebrafish have been used to identify mutations in genes with important roles in organogenesis. One of these mutants, small heart, develops a diminutive and severely malformed heart and multiple developmental defects of the brain, ears, eyes, and kidneys. Using a positional cloning approach, we identify that the mutant gene encodes the zebrafish Na+/K+-ATPase alpha1B1 protein. Disruption of Na+/K+-ATPase alpha1B1 function via morpholino "knockdown" or pharmacological inhibition with ouabain phenocopies the mutant phenotype, in a dose-dependent manner. Heterozygosity for the mutation sensitizes embryos to ouabain treatment. Our findings present novel genetic and morphological details on the function of the Na+/K+-ATPase alpha1B1 in early cardiac morphogenesis and the pathogenesis of the small heart malformation. We demonstrate that the reduced size of the mutant heart is caused by dysmorphic ventricular cardiomyocytes and an increase in ventricular cardiomyocyte apoptosis. This study provides a new insight that Na+/K+-ATPase alpha1B1 is required for maintaining ventricular cardiomyocyte morphology and viability.

Canertinib, a water-soluble, orally available analog of PD-169414 (Pfizer Inc), is an EGFR tyrosine kinase inhibitor under development by Pfizer Inc as a potential treatment for cancer.

The genotoxicity of endogenously formed N-nitrosamines from secondary amines and sodium nitrite (NaNO(2)) was evaluated in multiple organs of mice, using comet assay. Groups of four male mice were orally given dimethylamine, proline, and morpholine simultaneously with NaNO(2). The stomach, colon, liver, kidney, urinary bladder, lung, brain, and bone marrow were sampled 3 and 24 h after these compounds had been ingested. Although secondary amines and the NaNO(2) tested did not yield DNA damage in any of the organs tested, DNA damage was observed mainly in the liver following simultaneous oral ingestion of these compounds. The administration within a 60 min interval also yielded hepatic DNA damage. It is considered that DNA damage induced in mouse organs with the coexistence of amines and nitrite in the acidic stomach is due to endogenously formed nitrosamines. Ascorbic acid reduced the liver DNA damage induced by morpholine and NaNO(2). Reductions in hepatic genotoxicity of endogenously formed N-nitrosomorpholine by tea polyphenols, such as catechins and theaflavins, and fresh apple, grape, and orange juices were more effective than was by ascorbic acid. In contrast with the antimutagenicity of ascorbic acid in the liver, ascorbic acid yielded stomach DNA damage in the presence of NaNO(2) (in the presence and absence of morpholine). Even if ascorbic acid acts as an antimutagen in the liver, nitric oxide (NO) formed from the reduction of NaNO(2) by ascorbic acid damaged stomach DNA.

To investigate the possible long-term consequences of gestational exposure to cannabinoids on cognitive functions, pregnant rats were administered with the CB1 receptor agonist WIN 55,212-2 (WIN), at a dose (0.5 mgkg) that causes neither malformations nor overt signs of toxicity. Prenatal WIN exposure induced a disruption of memory retention in 40- and 80-day-old offspring subjected to a passive avoidance task. A hyperactive behavior at the ages of 12 and 40 days was also found. The memory impairment caused by the gestational exposure to WIN was correlated with alterations of hippocampal long-term potentiation (LTP) and glutamate release. LTP induced in CA3-CA1 synapses decayed faster in brain slices of rats born from WIN-treated dams, whereas posttetanic and short-term potentiation were similar to the control group. In line with LTP shortening, in vivo microdialysis showed a significant decrease in basal and K(+)-evoked extracellular glutamate levels in the hippocampus of juvenile and adult rats born from WIN-treated dams. A similar reduction in glutamate outflow was also observed in primary cell cultures of hippocampus obtained from pups born from mothers exposed to WIN. The decrease in hippocampal glutamate outflow appears to be the cause of LTP disruption, which in turn might underlie, at least in part, the long-lasting impairment of cognitive functions caused by the gestational exposure to this cannabinoid agonist. These findings could provide an explanation of cognitive alterations observed in children born from women who use marijuana during pregnancy.

The synthesis of 5-(6-methyl-2-substituted 4-pyrimidinyloxymethyl)-2,3-dihydro-1,3,4-oxadiazole-2-thiones and their 3-morpholinomethyl derivatives and the results of anti-inflammatory activity in vivo are described. Most of the tested compounds exhibited anti-inflammatory activity and some of them were more active than acetylsalicylic acid.

In order to continue the research of the rutin semisynthetic derivatives we obtained some new morpholine and piperidine derivatives. Melting points, solubility, yield, C, H, N elemental analysis and UV spectra characterised all of these compounds. The toxicity was established by intraperitoneal administration at mice; the values of LD50 are between 743.75 and 856.25 mg/kg (medium toxicity). Pharmacological assays included the following parameters: leukocytes formula, NBT test, and serum complement activity. The new derivatives of rutin have a very good solubility in water, a pH closed to physiological value and immunosuppressive activity: peripheric blood PMN and lymphocytes are decreased; fagocytic capacity of peripheric blood PMN is also decreased; serum complement activity is not changed.

Fomocaine (CAS 56583-43-8) is a local anaesthetic (LA) with long lasting surface effect and low toxicity. Nevertheless, it is not optimal yet. Therefore, 7 newly synthesised derivatives, 4 diethanolamines (OE 6000, OE 7000, OE 8000, OE 9000) and 3 morpholines (OE 500, OE 1000, OE 5000) were compared with procaine-HCl (CAS 51-05-8) and tetracaine-HCl (CAS 136-47-0) in rats. Based on standard methods for the testing of LA effects and using two methods for characterising side effects and toxicity of LA (paresis of the N. ischiadicus, LD50) it can be concluded that: a) The very good surface anaesthesia caused by especially fomocaine and tetracaine could be stated but concerning conduction anaesthesia procaine is better qualified. b) Concerning conduction anaesthesia, diethanolamine derivatives are more potent compared to morpholine derivatives. c) Surface anaesthesia shows a different picture: the effect of fomocaine is in between. d) The paresis of the N. ischiadicus as a first sign of toxic side effects indicated that low effect is combined with short paresis. e) Compared to the LD50 of fomocaine, the toxicity of OE 500 and OE 5000 is only one half. On the basis of the experiments with fomocaine derivatives, distinct structure-activity relationships could be demonstrated for fomocaine derivatives concerning a) LA effects and b) toxicity. Altogether OE 9000 could be a promising candidate for systemic use.

Cerebellar granule neurons grown in high potassium undergo rapid apoptosis when switched to medium containing 5 mm potassium, a stimulus mimicking deafferentation. This cell death can be blocked by genetic deletion of Bax, a member of the pro-apoptotic Bcl-2 family, cycloheximide an inhibitor of macromolecular synthesis or expression of dominant-negative c-jun. These observations suggest that Bax activation is the result of c-jun target gene(s) up-regulation following trophic withdrawal. Candidate genes include the BH3-only Bcl-2 family members Dp5 and Bim. The molecular mechanisms underlying granule cell neuronal apoptosis in response to low potassium were investigated using CEP-1347 (KT7515), an inhibitor of the MLK family of JNKKK. CEP-1347 provided protection of potassium-serum-deprived granule cells, but such neuroprotection was not long term. The incomplete protection was not due to incomplete blockade of the JNK signaling pathway because c-jun phosphorylation as well as induction of c-jun RNA and protein were completely blocked by CEP-1347. Following potassium-serum deprivation the JNKK MKK4 becomes phosphorylated, an event blocked by CEP-1347. Cells that die in the presence of CEP-1347 activate caspases; and dual inhibition of caspases and MLKs has additive, not synergistic, effects on survival. A lack of synergism was also seen with the p38 inhibitor SB203580, indicating that the neuroprotective effect of the JNK pathway inhibitor cannot be explained by p38 activation. Activation of the JNK signaling pathway seems to be a key event in granule cell apoptosis, but these neurons cannot survive long term in the absence of sustained PI3 kinase signaling.

Cisapride and mosapride are structurally and pharmacologically related prokinetic agents. In contrast to mosapride, cisapride causes embryonic lethality in teratology studies, and has been related to fatal cardiac arrhythmia in the adult. The arrhythmogenic potential of cisapride is linked to its potential to inhibit a specific ion channel (I(Kr)) as a side effect. Mosapride lacks I(Kr)-blocking properties. The aims of this study were (1) to compare the effects of cisapride and mosapride on embryonic heart rhythm in vitro and (2) to investigate if cisapride in vivo, has potential to induce hypoxia-related teratogenic effects as has been shown for selective I(Kr)-blockers. Cisapride induced severe embryonic bradycardia (approximately 60% decrease), and arrhythmia in 94% of the cultured rat embryos at 1000 ng/ml. Mosapride did not induce any bradycardia or arrhythmia up to 2000 ng/ml. In vivo, single dose administration of cisapride to rats on gestational day (GD) 13 caused digital reductions (8/108 fetuses, 4/9 litters) at 75 mg/kg and high incidence of embryonic death (55-82%) at 100-200 mg/kg. Identical developmental toxic effects have been described after temporary interruption of oxygen supply, and after single dose administration of selective I(Kr)-blockers, on the same GD. The results support the idea that all potent I(Kr)-blocking agents have the potential to cause embryolethality and teratogenicity, and that the adverse effects are mediated via hypoxic episodes due to embryonic arrhythmia., (Copyright 2002 Elsevier Science Inc.)

ErbB2 is overexpressed in 25-30% of breast and ovarian cancers, correlates with poor prognosis and lower survival and has also been associated with chemoresistance. We have established an isogenic pair of human ovarian cells that differ only in the expression of erbB2 protein in order to elucidate the role of the protein in determining cellular sensitivity to various drugs and agents. These included cisplatin and paclitaxel, the main drugs used in the treatment of ovarian cancer, and also various signal transduction inhibitors affecting the ras and P13K pathways. Transfection of erbB2 resulted in cells stably overexpressing the protein and showing increased motility compared to the empty vector control cells. In cells overexpressing erbB2, the most notable effect on chemosensitivity was that of significantly increased (5-fold) sensitivity to the heat shock protein 90 (HSP90) molecular chaperone inhibitor geldanamycin. In contrast, erbB2-overexpressing cells showed statistically significant resistance to cisplatin, the P13K inhibitor LY294002 and the tyrosine kinase inhibitor emodin. No significant difference in growth inhibition was observed after exposure to paclitaxel, two additional HSP90 inhibitors radicicol and 17AAG, the cyclin-dependent kinase inhibitor flavopiridol, the epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor PD153035, the mek inhibitor U0126 or the famesyl transferase inhibitor R115777. Exposure of cells to geldanamycin, 17AAG, emodin, LY294002 and cisplatin led to depletion of erbB2 in the transfected cells. These data suggest that erbB2 status in ovarian cancr may contribute to chemosensitivity, in some cases leading to increased sensitivity (as with geldanamycin) but in other cases leading to resistance (as with cisplatin).

Since morpholino oligos were first introduced as a means to inhibit gene function in embryos, in the Spring of 2000, they have been tested in a range of model organisms, including sea urchin, ascidian, zebrafish, frog, chick, and mouse. This review surveys the results of these studies and examines the successes and limitations of the approach for targeting maternal and zygotic gene function. The evidence so far suggests that, with careful controls, morpholinos provide a relatively simple and rapid method to study gene function., ((C)2002 Elsevier Science (USA).)

Sanofi-Synthélabo (formerly Sanofi) is developing SR-121463, a vasopressin V, receptor antagonist, as a potential treatment for cardiovascular indications such as congestive heart failure (CHF) and hypertension [330073], [341858]. By September 2001, it had entered phase IIa trials for these indications [421268]. SR-121463 was in phase I clinical trials for CHF and hypertension in June 2001 [359231], [413342]. It was also being evaluated for the potential treatment of glaucoma but its development has been discontinued for this indication [367094]. In October 1999, Lehman Brothers predicted a 5% chance of the compound reaching market, with a launch anticipated in 2004 and potential peak sales of $100 million in 2012 [346267].

Exposure of Japanese medaka (Oryzias latipes) embryos and adult males to sublethal concentrations of two commonly used fungicides, Acrobat MZ and Tattoo C, had varying effects on development and adult male behavior. During embryolarval assays, medaka exposed to Acrobat exhibited decreased heart rates throughout embryonic development and were smaller at hatching than controls. Differences in hatching size were not apparent with Tattoo although some decrease in heart rate was observed at higher concentrations. Tail lesions and abnormal spinal development was a problem common to embryos exposed to Acrobat. Neither time to hatch nor early growth was affected by exposure to either fungicide. Adult males that had been exposed to high concentrations of Acrobat or Tattoo over a 28-day period were less likely to approach females during behavioral trials. The absence of other behavioral effects, particularly those related to reproductive performance, may have been due to females being sexually unreceptive during behavioral trials. No other behavioral modifications were apparent for adult males exposed to either fungicide. Information about developmental and behavioral responses to sublethal concentrations of pesticides is important for establishing environmental guidelines concerning their use.

Cetethyl Morpholinium Ethosulfate is a quaternary salt used as an antistatic agent and as a surfactant in several hair care products. The concentration at which this ingredient is used is unknown, although data reported in 1984 indicated a maximum concentration of 1%. In an inhalation toxicity study, the approximate lethal concentration of Cetethyl Morpholinium Ethosulfate was 0.403 mg/mm3. This ingredient was shown to be a severe ocular irritant in an animal study. No other safety test data on this ingredient were available. These data were clearly insufficient to support the safety of Cetethyl Morpholinium Ethosulfate in cosmetics. Data available on Morpholine were summarized, but these data themselves were insufficient to support safety. The data needed in order to complete the safety assessment of Cetethyl Morpholinium Ethosulfate include: methods of manufacture and impurities, especially nitrosamines; current concentration of use; skin penetration; if there is significant skin penetration, then both a 28-day dermal toxicity study to assess general skin and systemic toxicity and a reproductive and developmental toxicity study are needed; two genotoxicity studies, at least one in a mammalian system, if positive, then a 2-year dermal carcinogenisis study using National Toxicology Program (NTP) methods may be needed; ultraviolet (UV) absorption data, if significantly absorbed, then photosensitization data are needed; dermal irritation and sensitization; and ocular toxicity, if available.

The aim of the present study was to examine the chemopreventive efficacy of S-methylcysteine (SMC) on rat hepatocarcinogenesis induced by concurrent administration of sodium nitrite (NaNO(2)) and morpholine (Mor) using a medium-term rat liver carcinogenesis bioassay (Ito test). Administration of SMC caused significant reduction in the areas of glutathione S-transferase placental form positive foci along with a significant decrease of hepatocyte 5-bromo-2'-deoxyuridine (BrdU) labeling indices. These results demonstrated potent chemopreventive effects of SMC against hepatocarcinogenesis due to concurrent administration of Mor and NaNO(2). SMC could thus be an effective chemopreventive agent for decreasing the risk of carcinogenicity from environmental precursors of N-nitroso compounds.

Delta9-Tetrahydrocannabinol, the main active component of marijuana, induces apoptosis of transformed neural cells in culture. Here, we show that intratumoral administration of Delta9-tetrahydrocannabinol and the synthetic cannabinoid agonist WIN-55,212-2 induced a considerable regression of malignant gliomas in Wistar rats and in mice deficient in recombination activating gene 2. Cannabinoid treatment did not produce any substantial neurotoxic effect in the conditions used. Experiments with two subclones of C6 glioma cells in culture showed that cannabinoids signal apoptosis by a pathway involving cannabinoid receptors, sustained ceramide accumulation and Raf1/extracellular signal-regulated kinase activation. These results may provide the basis for a new therapeutic approach for the treatment of malignant gliomas.

The synthesis, pharmacology and toxicology of four morpholine derivatives from 1-(2-arylmorpholino)-3-phenyl-3-propanonoxime and the synthesis of two anilides are described. The structures of the synthesized derivatives were proved by IR, 1H NMR and occasionally with 13C NMR. The acute toxicity of the compounds in mice was determined. A comparative pharmacological study of the in vivo effect on the central nervous system was realised by the following screening tests: pentobarbital induced sleeping time, locomotor activity and behaviour despair test for antidepressive activity. The most active compound was 1-(2-phenylmorpholino)-3-phenyl-3-propanonoxime (2b) which showed low toxicity and antidepressive activity at a dose of 1/10 LD50.

N-methylmorpholine, which is used as a catalyst in polyurethane foams producing, in solvents, stabilizing agents, and corrosion inhibitors, was administered to female rats by gavage at 100, 200, 600, and 900 mg/kg during organogenesis. It did not exhibit selective toxicity toward the developing conceptus. This compound administered to pregnant females was fetotoxic and teratogenic in the presence of maternal toxicity. N-methylmorpholine induced anophthalmia, internal hydrocephalus, and hydronephrosis but only at one dose which was also maternotoxic. Teratogenesis Carcinog. Mutagen. 19:369-376, 1999., (Copyright 1999 Wiley-Liss, Inc.)

The synthesis of nine original morpholine derivatives, i.e. 2-aryl-4-(3-arylpropyl)morpholines, is described. The structure of all synthesised derivatives was proved by IR and 1H-NMR, and some of them by 13C-NMR. Acute toxicity studies of the compounds were performed on mice. A comparative pharmacological study of the in vivo effects on the central nervous system was undertaken using the screening tests: hexobarbital induced sleeping time: locomotor activity; and behaviour despair (for antidepressive activity). The most active compound 4-(2-benzoylethyl)-2-phenyl-3-methyl) morpholine 4e was studied for MAO-A and MAO-B inhibition in vitro in rat brain mitochondria preparations.

4-week repeated dose toxicity study with 4-week recovery test of landiolol hydrochloride (ONO-1101), a novel ultra short acting beta-blocker, was conducted in beagle dogs. ONO-1101 was administered intravenously to dogs of both sexes at a dose level of 0 (control), 12.5, 25 or 50 mg/kg/day. No deaths occurred throughout the treatment period. Transitory licking chops, vomiting, nausea, diarrhea and soft feces were observed occasionally in both sexes dosed 25 and 50 mg/kg/day and the incidence seemed dose-dependent. However, those incidence declined in the course of the treatment period. Hematology showed a decrease in red blood cell count, hematocrit and hemoglobin value in both sexes receiving 25 and 50 mg/kg/day. ONO-1101 did not effect on body weight, food consumption, respiratory rate, pulse, rectal temperature, heart rate, blood pressure, electrocardiography, renal or hepatic function, ophthalmology, urinalysis, occult blood in feces, blood chemistry, organ weights, necropsy and microscopic findings at any doses. These results indicate that the no-adverse-effect level of ONO-1101 in dogs is 12.5 mg/kg/day for both sexes in this study.

A perinatal and postnatal study of landiolol hydrochloride (ONO-1101), a novel ultra short acting beta-blocker, was conducted in Sprague-Dawley(SD) rats. ONO-1101 was administered intravenously at a dose level of 0 (control), 25, 50 or 100 mg/kg/day from day 17 of gestation to day 20 after parturition to examine the effects on pregnancy, delivery, lactation and the effects on postnatal growth and development of offspring. In the 100 mg/kg/day group, hypoactivity, reddish lacrimation, clonic convulsion and bradypnea/apnea were observed after administration and 5 animals died in the treatment period, and body weight on day 21 and food consumption on day 14-21 of dam at weaning were lower than control group. In the 50 mg/kg/day group, reddish lacrimation was occasionally seen in some animals. ONO-1101 had no effects on pregnancy, delivery, lactation and necropsy findings or organ weights of dams. In the 100 mg/kg/day group, viability of offspring on day 4 after birth decreased and body weight gain of the suckling suppressed, but those changes recovered after weaning. On the skeletal examination of offspring culled on day 4 after birth, decrease in the mean number of osiffied phalanges of hindpaw and increase in the incidence of unossified talus bone were seen in the 100 mg/kg/day group, however, no delay of ossification was found form the weanling. There were no influence of ONO-1101 on external differentiation, functional, behavioral or learning abilities or reproductive performance in offspring. From the above results, it is estimated that the no-toxic dose level of ONO-1101 under these experimental conditions is 50 mg/kg/day for dam and offspring.

4-week repeated dose toxicity study with 4-week recovery test of landiolol hydrochloride (ONO-1101), a novel ultra short acting beta-blocker, was conducted in Sprague-Dawley (SD) rats. ONO-1101 was administered intravenously to rats of both sexes at a dose level of 0 (control), 12.5, 25, 50 or 100 mg/kg/day. In the 100 mg/kg/day group, bradypnea or dyspnea was seen in all animals, pale in ear, eye and foot, tremor, reddish lacrimation and loss of righting reflex were also observed in some animals right after administration, and then those signs disappeared within 1 min after administration. During the treatment period, 3/20 animals of each sex in the 100 mg/kg/day showed clonic convulsion and died within 2 min after administration. No clinical changes were seen in the 50 mg/kg/day group or lower. Histopathological findings showed atrophy of the submaxillary gland in females and vessel-wall thickening and perivascular fibrosis of the injection site (tail) in both sexes at 100 mg/kg/day, however those changes were reversible. ONO-1101 did not effect on body weight, food consumption, ophthalmology, urinalysis, hematology, blood chemistry, organ weights or necropsy at any doses. These results indicate that the no-adverse-effect level of ONO-1101 in rats is 50 mg/kg/day for both sexes in this study.

A teratogenicity study of landiolol hydrochloride (ONO-1101), a novel ultra short acting beta-blocker, was conducted in Sprague-Dawley (SD) rats. ONO-1101 was administered intravenously at a dose level of 0 (control), 25, 50 or 100 mg/kg/day to pregnant rats from day 7 to 17 of gestation, and effects of ONO-1101 on dams, fetuses and their offspring, were examined. In the 100 mg/kg/day group, hypoactivity, bradypnea, reddish lacrimation, clonic convulsion and loss of righting reflex were observed and 2 animals died. Food consumption in the 100 mg/kg/day group decreased during the treatment period. No drug-related changes were observed in dams for their body weights, necropsy findings or organ weights. Decrease in placental weight was seen in the 100 mg/kg/day group, but no effect was found in fetal weight. ONO-1101 had no effects on delivery and lactation. On day 4 after birth, viability of offspring were decrease in the 50 or 100 mg/kg/day group, and body weight of males were decreased in the 100 mg/kg/day group, but no change caused by the treatment was observed in growth of offspring thereafter. On skeletal examination in offsprings culled on day 4 after birth, increase in the incidence of unossificated talus were seen in the 50 or 100 mg/kg/day group. But no drug-related anomalies were observed in external, skeletal or visceral findings in fetuses. It was not also found any influence of ONO-1101 on external differentiations, functional, behavioral or learning abilities or reproductive performance in offspring. From the above results, it is estimated that the no-toxic dose level of ONO-1101 under these experimental conditions is 50 mg/kg/day for dams, and 25 mg/kg/day for their offspring.

Single dose toxicity studies of landiolol hydrochloride (ONO-1101), a novel ultra short acting beta-blocker, were conducted in Sprague-Dawley (SD) rats and beagle dogs. ONO-1101 was administered intravenously at a dose level of 37.5, 75, 150 or 300 mg/kg to rats of both sexes and 25, 50 or 100 mg/kg to male dogs. In the rat study, 5/6 males in the 150 mg/kg group and all animals in the 300 mg/kg group died during or right after administration. Survivors in the 150 mg/kg group showed temporal hypoactivity, bradypnea, dyspnea, tremor, loss of righting reflex and reddish lacrimation up to 5 min after injection. One male in the 150 mg/kg group had a tendency of suppression on body weight gain. No effects on clinical signs and body weight gain were seen in the 75 mg/kg group or lower. Necropsy findings showed only red tear in the majority of the decedents. In the dog study, all animals died within 6 min after administration in the 100 mg/kg group, showed ataxic gait, rolling and tachypnea followed by bradypnea and gasping/apnea. Incontinence of urine, defecation and vocalization were also seen in each one of two animals before death. Temporal hypoactivity was seen 1 min after administration in the 50 mg/kg group. No clinical signs were seen in the 25 mg/kg group. ONO-1101 did not affect bodyweight or food consumption. Necropsy findings of the decedents showed no abnormalities. It is indicated that the minimum lethal doses are 150 mg/kg in rats and 100 mg/kg in dogs.

A teratogenicity study of landiolol hydrochloride (ONO-1101), a novel ultra short acting beta-blocker, was conducted in KAR:NZW rabbits. ONO-1101 was administered intravenously at a dose level of 0 (control), 25, 50 or 100 mg/kg/day to pregnant rabbit from day 6 to 18 of gestation to examine the effects on dams and fetuses. Death occurred on 3 animals in the 100 mg/kg/day group and one animal in the 50 mg/kg/day group during the treatment period. Hypoactivity, bradypnea/apnea and clonic convulsion after administration was observed in animal dosed 100 mg/kg/day. No maternal effects were seen on body weight, food consumption, necropsy findings or organ weights. External, skeletal and visceral findings revealed no adverse effects of ONO-1101 on fetuses. From the above results, it is estimated that the no-toxic dose level of ONO-1101 under these experimental conditions is 25 mg/kg/day for dams and 100 mg/kg for fetuses.

A fertility study of landiolol hydrochloride (ONO-1101), a novel ultra short acting beta-blocker, was conducted in Sprague-Dawley (SD) rats. ONO-1101 was administered intravenously to males from the 64th day before mating until necropsy, and to females from 15th day before mating until day 7 of gestation, at a dose level of 0 (control), 25, 50 or 100 mg/kg/day. On day 20 of gestation, all dams were sacrificed and their fetuses were examined. In the 100 mg/kg/day group, hypoactivity, clonic convulsion, bradypnea/apnea and redish lacrimation were observed after administration in both sexes, and 3 males and 2 females died. Reddish lacrimation was occasionally seen in males at late stage of the treatment period in 50 mg/kg/day group. In the 100 mg/kg/day group, body weight gain suppressed in females from the premating through the gestation period, and food consumption decreased in females during the premating period, and mean thymus weight decreased in males. ONO-1101 did not affect estrous cycle, copulatory or fertility in both sexes or external, skeletal or visceral features of the fetuses. From the above results, it is estimated that the no-toxic dose level of ONO-1101 under these experimental conditions is 50 mg/kg/day for general toxicity in parents, and 100 mg/kg/day for the reproductive performance in parents and for the development of fetuses.

The effects of the synthetic cannabinoid receptor agonist WIN 55,212-2 on dopamine receptor-mediated alleviation of akinesia were evaluated in the reserpine-treated rat model of parkinsonism. The dopamine D2 receptor agonist quinpirole (0.1 mg/kg, ip) caused a significant alleviation of the akinesia. This effect was significantly reduced by coinjection with the cannabinoid receptor agonist WIN 55,212-2 (0.1 and 0.3 mg/kg). The simultaneous administration of the cannabinoid receptor antagonist SR 141716A (3 mg/kg, ip) with quinpirole and WIN 55,212-2 blocked the effect of WIN 55,212-2 on quinpirole-induced alleviation of akinesia. The selective dopamine D1 receptor agonist chloro-APB (SKF82958, 0.1 mg/kg) alleviated akinesia in a significant manner. WIN 55,212-2 (0.1-1 mg/kg, ip) did not affect the antiakinetic effect of chloro-APB. Combined injection of both D1 and D2 dopamine receptor agonists (both at either 0.1 or 0.02 mg/kg) resulted in a marked synergism of the antiakinetic effect. WIN 55,212-2 (0.1-1 mg/kg) significantly reduced the antiakinetic effect of combined injections of quinpirole and chloro-APB at both 0.1 and 0.02 mg/kg. The effect of 0.3 mg/kg WIN 55,212-2 on combined D1 and D2 agonist-induced locomotion (0.02 mg/kg) was blocked by SR 141761A (3 mg/kg). Neither WIN 55,212-2 alone (0.1 and 0.3 mg/kg) nor SR 141716A (3 and 30 mg/kg) alone had an antiparkinsonian effect. These results suggest that cannabinoids may modulate neurotransmission in the pathway linking the striatum indirectly to basal ganglia outputs via the lateral globus pallidus and the subthalamic nucleus.

For carcinogenic risk assessment of combinations of N-nitroso precursors in man, the effects of feeding methylurea (MU) or morpholine (Mor) plus sodium nitrite (NaNO2) were investigated using a multi-organ carcinogenesis model. In experiment 1, to initiate multiple organs, groups of 10 or 20 male F344 rats were treated with 6 carcinogens targeting different organs. Starting a week after completion of this initiation phase, animals were given 0.1% MU or 0.5% Mor in their food and/or 0.15% NaNO2 in their drinking water for 23 weeks. The induction of tumors and/or preneoplastic lesions in the forestomach and esophagus was significantly increased in the group receiving MU plus NaNO2. The numbers and areas of liver glutathione S-transferase placental form (GST-P)-positive foci were significantly elevated with MU or Mor plus NaNO2. Experiment 2 was conducted to assess formation of N-nitroso compounds in the stomach, and to detect DNA adduct generation in target organs by immunohistochemical staining. Groups of 5 or 14 animals were starved overnight, then given 0.4% MU or 2.0% Mor in the diet, or basal diet alone for 1 h. Then NaNO2 or distilled water was given intragastrically. The mean gastric N-methyl-N-nitrosourea yield in the MU plus NaNO2 group was 7700 micrograms at 2 h after combined administration. The mean N-nitrosomorpholine yield in the group given Mor plus NaNO2 was 6720 micrograms. Immunohistochemically, N7-methyldeoxyguanosine-positive nuclei were evident in the forestomach epithelium at 8 h after the combination treatment with MU plus NaNO2.

Oral administration of the HIV protease inhibitor L-689,502 caused cholestasis and hepatocyte injury in rats and dogs. These changes occurred rapidly, with elevations in serum transaminase observed as early as 6 hr after oral dosing in dogs. The acute phase of this hepatotoxic response was characterized in more detail in rats. Following intravenous administration, bile flow was decreased in a dose-dependent manner with greater than 90% decrease in less than 30 min at a dose of 5 mg/kg. The decrease in bile flow was associated with a decrease in erythritol clearance. The decrease in bile flow was not due to disruption of biliary tight junctions. Sucrose clearance was not increased and biliary bile acid concentrations in treated animals were not different from controls. Unlike control animals, bile flow was not stimulated by infusion of the bile acid tauroursodeoxycholic acid in animals treated with L-689,502. These cholestatic effects may be due, in part, to direct hepatocyte injury. Histological examination of perfusion-fixed livers 30 min after L-689,502 administration revealed periportal changes including hepatocyte vacuolation and occasional single cell necrosis. On a subcellular level, the nucleus and mitochondria were intact in less-severely affected cells. However, extensive vacuolation with multilamellar inclusions was pronounced in these cells. In addition, canalicular ectasia was also observed which was consistent with the cholestatic changes that were seen. In summary, L-689,502 is a potent, rapid acting hepatotoxin in dogs and rats. The mechanism by which this agent induces cholestasis is novel compared to other well-characterized cholestatic agents such as alpha-naphtylisothiocyanate and ethinyl estradiol., (Copyright 1997 Academic Press.)

1. The sigma-drug binding site of guinea-pig liver is carried by a protein which shares significant amino acid sequence similarities with the yeast sterol C8-C7 isomerase (ERG2 protein). Pharmacologically-but not structurally-the sigma 1-site is also related to the emopamil binding protein, the mammalian sterol C8-C7 isomerase. We therefore investigated if sterol C8-C7 isomerase inhibitors are high affinity ligands for the (+)-[3H]-pentazocine labelled sigma 1-binding site. 2. Among the compounds which bound with high affinity to native hepatic and cerebral as well as to yeast expressed sigma 1-binding sites were the agricultural fungicide fenpropimorph (Ki 0.005 nM), the antihypocholesterinaemic drugs triparanol (Ki 7.0 nM), AY-9944 (Ki, 0.46 nM) and MDL28,815 (Ki 0.16 nM), the enantiomers of the ovulation inducer clomiphene (Ki 5.5 and 12 nM, respectively) and the antioestrogene tamoxifen (Ki 26 nM). 3. Except for tamoxifen these affinities are essentially identical with those for the [3H]-ifenprodil labelled sterol C8-C7 isomerase of S. cerevisiae. This demonstrates that sigma 1-binding protein and yeast isomerase are not only structurally but also pharmacologically related. Because of its affiliations with yeast and mammalian sterol isomerases we propose that the sigma 1-binding site is localized on a sterol isomerase related protein, involved in postsqualene sterol biosynthesis.

The hypothesis is offered predicting that Gaucher patients could be treated with a drug that slows the synthesis of glucosylceramide, the lipid that accumulates in this disorder. The present therapeutic approach involves augmenting the defective enzyme, glucosylceramide beta-glucosidase, with exogenous beta-glucosidase isolated from human tissue. This spectacularly expensive mode of treatment should be replaceable with a suitable enzyme inhibitor that simply slows formation of the lipid and matches the rate of synthesis with the rate of the defective, slowly working beta-glucosidase. Several drugs that possess this ability are available, the best known of which is 1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), a designer inhibitor that resembles the synthase's substrate and product. PDMP has been found to be effective in mice, rats, fish, and a wide variety of cultured cells. Its use, at suitable dosages, seems to be harmless, although long-term tests have not been made. The lack of suitable animal models of Gaucher disease has made it difficult to test the hypothesis adequately, but PDMP does rapidly lower the levels of glucosylceramide in normal animal tissues and the animals evidently do well with the lowered levels of glucosylceramide and its more complex glycolipid metabolites.

MF-268 bitartrate [(3a S, 8a R)-1,2,3,3a,8,8a-hexahydro-1,3a,8-trimethylpyrrolo[2,3-b]indol- 5-ol[8-(cis2,-6-dimethyl-morpholin-4-yl)octyl]-carbamate L-bitartrate hydrate; Mediolanum Farmaceutici, Milan, Italy] is a pseudo-reversible carbamate-type cholinesterase inhibitor (ChEI) which interacts with the catalytic and regulatory anionic site of the enzyme. Its effects on extracellular levels of acetylcholine (ACh), norepinephrine (NE), dopamine (DA), and serotonin (5-HT, 5-hydroxytryptamine) were studied in rat cortex by using a microdialysis technique coupled with high-performance liquid chromatography-electrochemical detection (HPLC-ECD). Conscious, freely moving rats were systemically [per os (p.o.) and subcutaneously (s.c.)] administered MF-268 with no ChEI in the probe. Cholinesterase inhibition in brain was assayed in parallel experiments. Oral administration of MF-268 (0.5, 2.0, and 5.0 mg/kg) produced a significant increase of extracellular ACh in cortex; the maximal increase was 300% [not significant (n.s.)], 460% and 1,200%, respectively. Maximal cholinesterase (ChE) inhibition was 2.3% (n.s.) at 9 hr and 9.7% (P < .05) at 12 hr after the 2.0 and 5.0 mg/kg doses, respectively. Norepinephrine and DA levels were increased 180% and 100% after the 5.0 mg/kg dose, respectively; 100% and 60% after the 2.0 mg/kg dose, respectively; and 70% for both amines after the 0.5 mg/kg dose, respectively. The elevation lasted at least 5 hr with the 2.0 and 5.0 mg/kg doses. There were no major changes in 5-HT levels at these three doses. Subcutaneous administration (0.5 and 2.0 mg/kg) produced a maximal 360% (5.5 hr) and 2,500% (5 hr) increase in extracellular ACh, respectively. Maximal ChE inhibition was 13% (0.5 mg/kg) and 41% (2.0 mg/kg). Neither 0.5 nor 2.0 mg/kg produced a consistent modification of NE. Only a transient increase in DA was seen with the 0.5 mg/kg dose. There were no changes in 5-HT levels at these two doses. MF-268-treated animals showed slight cholinergic side effects (chewing, tremor) at both doses. MF-268 administered intracortically through the microdialysis probe at a concentration of 50 microM induced a 5,900% increase in ACh levels at 6 hr. This effect started 30 min after injection and continued throughout the period of administration. MF-268 produced a significant decrease in NE levels (-44%) starting at 30 min, and a slight but significant increase in DA levels of 45% at 2.5 hr. A significant increase of 5-HT (58%) was also observed starting at 4 hr. Slight symptoms of cholinergic toxicity were observed during intracortical administration.

The relationship between different chemical modifications on morpholinylanthracyclines and their ability to overcome multidrug resistance (MDR) has been evaluated testing all compounds in vitro on LoVo and LoVo/DX human colon adenocarcinoma cells and in vivo disseminated P388 and P388/DX murine leukemias. Results obtained led us to the following conclusions: 1) the insertion of the morpholinyl or the methoxymorpholinyl group on position 3' of the sugar moiety confers the ability to overcome MDR in vitro and in vivo; conversely, 4' morpholinyl compounds are effective on MDR cells only in vitro and result inactive in vivo on DX-resistant leukemia; 2) all chemical modifications performed on 3' morpholinyl or methoxymorpholinyl derivatives, that is substitutions on the aglycone or on position 2 of the morpholino ring, do not interfere with the activity of the compounds: all derivatives present in fact the same efficacy on sensitive and resistant models. It is concluded that position 3' in the sugar moiety plays a crucial role in the ability of morpholinyl-anthracyclines to overcome MDR.

Montirelin hydrate (NS-3) is a new drug for the treatment of disturbance of consciousness. The single dose toxicity studies of its degradation product and impurity (CNK-603) and its metabolite (CNK-6004) were conducted in Slc: ddY mice. The compounds were administered intravenously to male and female mice. Deaths occurred in both sexes of mice receiving more than 125 mg/kg of CNK-603. There were no treatment-related effects on survival in both sexes of mice receiving doses up to 2,000 mg/kg of CNK-6004. Approximate lethal dose of CNK-603 was 125 mg/kg and lethal dose of CNK-6004 was more than 2,000 mg/kg. Decrease in locomotor activity was observed in mice receiving the two compounds. Tremor, prone position and dyspnea were seen in mice receiving CNK-603. The body weight showed no changes attributable to the dosing of the two compounds in mice. In autopsies, congestion and hemorrhage in the lung were observed in dead mice given CNK-603. There were no remarkable changes in mice given CNK-6004. These results show that the lethal dose of CNK-603 is over 4 times lower than that of montirelin hydrate, and that CNK-6004 is less toxic than montirelin hydrate.

Neutrophils contain a multicomponent NADPH oxidase system that is involved in the production of microbicidal oxidants. Stimulation of human neutrophils with the peptide FMLP activates this respiratory burst enzyme to produce superoxide and also has been shown to result in activation of phosphatidylinositol (Ptdlns) 3-kinase. Treatment of human neutrophils with 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002), a potent and specific inhibitor of Ptdlns 3-kinase, resulted in complete inhibition of Ptdlns 3-kinase activity as well as in inhibition of superoxide production in FMLP-treated neutrophils in suspension; FMLP-stimulated oxidant production in adherent cells was also abolished. Treatment of human neutrophils with PMA resulted in production of superoxide without activation of Ptdlns 3-kinase; LY294002 did not block superoxide production in neutrophils exposed to PMA. In addition, LY294002 did not inhibit cellfree NADPH oxidase activation, CD11b-dependent adhesion, actin polymerization in response to FMLP, or FMLP-induced calcium flux. These results suggest that the signal transduction pathway of the FMLP-receptor involves activation of Ptdlns 3-kinase, which is required for subsequent superoxide production induced by the chemotactic peptide. Furthermore, Ptdlns 3-kinase may be located directly upstream of protein kinase C or other protein kinases, which in turn activate the NADPH oxidase system.

The albino experiments have shown that morpholide has an anticonvulsant effect in electroshock-, corazole-, thiosemicarbazide-, bicuculline-, strychnine-, arecoline-, nicotine-, tremorine-induced convulsions and tremors. The agent has a wider range of pharmacological effects than phenobarbital. There is a higher anticonvulsant effect of morpholide combined with phenobarbital, hexamidine and diazepam. It is suggested that GABA-ergic mechanisms contribute to the anticonvulsant action of morpholide.

Morpholinoalkyl esters (HCl salts) of diclofenac (1) were synthesized and evaluated in vitro and in vivo for their potential use as prodrugs for oral delivery. Prodrugs were freely soluble in simulated gastric fluid (SGF) and pH 7.4 phosphate buffer and showed a minimum of a 2000-fold increase in solubility over the parent drug. All prodrugs were more lipophilic than 1 as indicated by n-octanol/pH 7.4 buffer partition coefficients, but less lipophilic than 1 in terms of n-octanol/SGF partition coefficients. Potentiometrically determined ionization constants (pKas) were in the range of 7.52 to 8.40 at 25 degrees C. The chemical and enzymatic hydrolyses of prodrugs were evaluated in SGF/pH 7.4 phosphate buffer and rat plasma, respectively, at 37 degrees C. All prodrugs were quantitatively hydrolyzed to 1 by either chemical and/or enzymatic means. An increase in carbon chain length rendered the prodrugs more stable at pH 7.4, but less stable in SGF. In general, the esters were hydrolyzed rapidly in rat plasma at 37 degrees C, the half-lives of hydrolysis being in the range of 4.85 to 23.49 min. Based on in vitro results, prodrug 2 was chosen to evaluate solid-state stability, bioavailability, and in vivo ulcerogenicity. At elevated temperatures, the solid-state decomposition of 2 followed biphasic kinetics, with rapid decomposition occurring initially. The extent, but not the rate, of absorption was significantly greater in rats for prodrug 2 than 1 following single dose oral administration. Prodrug 2 was significantly less irritating to gastric mucosa than 1 following single and chronic oral administration in rats.

A proposed weak point in cancer cells is their need to synthesize novel or rare glucosphingolipids. It is further proposed that cancer patients be treated with a drug that slows the synthesis of glucosylceramide, the precursor of a large family of glucosphingolipids. Experimental data are furnished for chemotherapeutic and biochemical effects of PDMP, an analog of glucosylceramide and its precursor, ceramide. Promising results were obtained in the treatment of mice carrying Ehrlich ascites carcinoma cells and rats carrying C6 glioma cells. PDMP was found to be oxidized by cytochrome P-450, but this process could be blocked in vivo with piperonyl butoxide or cimetidine. A high level of blood glucose was found to elevate the size of rat kidneys and their content of UDP-glucose and its product, glucosylceramide. The excessive growth could be blocked by PDMP, which competes with UDP-glc for binding to glucosylceramide synthase. It is suggested that cancer patients be maintained at a low glucose level in order to slow the synthesis of glucosylceramide by tumor cells. Metabolic changes produced by PDMP in cultured cells, besides a rapid deletion of glucosphingolipids, were accumulation of the precursors (ceramide and sphingosine), loss of protein kinase C, and accumulation of diacylglycerol. It is suggested that many of the cellular changes produced by PDMP, such as loss of cell binding, are owing to existence of glucosylceramide-based "islands" floating in the outer cell surface; the islands may contain growth factor receptors and adhesion factors. An inhibitor that blocks sphingolipid synthesis, such as cycloserine, may prove to be a useful adjuvant for therapy with PDMP.

Fifty chemicals were tested for mutagenic activity in post-meiotic and meiotic germ cells of male Drosophila melanogaster using the sex-linked recessive lethal (SLRL) assay. As in the previous studies in this series, feeding was chosen as the first route of administration. If the compound failed to induce mutations by this route, injection exposure was used. One gaseous chemical (1,3-butadiene) was tested only by inhalation. Those chemicals that were mutagenic in the sex-linked recessive lethal assay were further tested for the ability to induce reciprocal translocations. Eleven of the 50 chemicals tested were mutagenic in the SLRL assay. These included bis(2-chloroethyl) ether, 1,4-butanediol diglycidyl ether, 1-chloro-2-propanol, dimethyl methylphosphonate, dimethyl morpholinophosphoramidate, dimethyloldihydroxyethylene urea, 2,2-dimethyl vinyl chloride, hexamethylphosphoramide, isatin-5-sulfonic acid (Na salt), isopropyl glycidyl ether, and urethane. Five of these, including 1,4-butanediol diglycidyl ether, 2,2-dimethyl vinyl chloride, hexamethylphosphoramide, isopropyl glycidyl ether, and urethane, also induced reciprocal translocations.