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Analysis of exposure margins in developmental toxicity studies for detection of human teratogens

The draft Step 2 ICH S5(R3) guideline includes an exposure-based endpoint as an option for selecting the high-dose in reproductive and developmental toxicity studies. To help determine an appropriate exposure margin for embryofetal developmental toxicity testing, a retrospective analysis was undertaken to determine what threshold would have been sufficient to detect hazards to embryofetal development in rats and rabbits for 18 known and 4 presumed human teratogens. The analysis showed that using a high dose that provided at least a 6-fold exposure margin in the developmental toxicity studies would have been sufficient to detect the teratogenic hazard with relevance for humans for all these therapeutics. With regards to human risk assessment practices for developmental toxicity, the analysis showed that, after excluding lenalidomide and pomalidomide data in rats, all available AUC margins at the NOAEL for the induction of malformations or embryofetal lethality were <4-fold of the exposure at the MRHD for all 22 therapeutics. These data support the proposed general approach of increased level of concern for human risk when exposure margins of the NOAEL to the MRHD are <10-fold, reduced concern when the exposure margins are 10- to 25-fold, and minimal concern when the exposure margin is > 25-fold.

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Reference:
Oxazolidine – Wikipedia,
Oxazolidine | C3H1339NO – PubChem

 

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Identification of a New Zinc Binding Chemotype by Fragment Screening

The discovery of a new zinc binding chemotype from screening a nonbiased fragment library is reported. Using the orthogonal fragment screening methods of native state mass spectrometry and surface plasmon resonance a 3-unsubstituted 2,4-oxazolidinedione fragment was found to have low micromolar binding affinity to the zinc metalloenzyme carbonic anhydrase II (CA II). This affinity approached that of fragment sized primary benzenesulfonamides, the classical zinc binding group found in most CA II inhibitors. Protein X-ray crystallography established that 3-unsubstituted 2,4-oxazolidinediones bound to CA II via an interaction of the acidic ring nitrogen with the CA II active site zinc, as well as two hydrogen bonds between the oxazolidinedione ring oxygen and the CA II protein backbone. Furthermore, 3-unsubstituted 2,4-oxazolidinediones appear to be a viable starting point for the development of an alternative class of CA inhibitor, wherein the medicinal chemistry pedigree of primary sulfonamides has dominated for several decades.

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Revisiting the SAR of the Antischistosomal Aryl Hydantoin (Ro 13-3978)

The aryl hydantoin 1 (Ro 13-3978) was identified in the early 1980s as a promising antischistosomal lead compound. However, this series of aryl hydantoins produced antiandrogenic side effects in the host, a not unexpected outcome given their close structural similarity to the antiandrogenic drug nilutamide. Building on the known SAR of this compound series, we now describe a number of analogs of 1 designed to maximize structural diversity guided by incorporation of substructures and functional groups known to diminish ligand-androgen receptor interactions. These analogs had calculated polar surface area (PSA), measured LogD7.4, aqueous kinetic solubility, and estimated plasma protein binding values in ranges predictive of good ADME profiles. The principal SAR insight was that the hydantoin core of 1 is required for high antischistosomal activity. We identified several compounds with high antischistosomal efficacy that were less antiandrogenic than 1. These data provide direction for the ongoing optimization of antischistosomal hydantoins.

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2-CYANOPYRROLOPYRIMIDINES AND PHARMACEUTICAL USES THEREOF

The invention relates to pyrrolo pyrimidines of formula (I), wherein Y represents-(CH2)t-O-or-(CH2)r-S-, p is 1 or 2, r is 1, 2 or 3, t is 1, 2 or 3, or Y is-(CH2)j-or-CH=CH-, j is 1 or 2; p is 1 or 2, or Y is-(CH2)f-, f is 1 or 2, p is 1, and the further radicals and symbols have the meaning as defined herein; their preparation, their use as pharmaceuticals, pharmaceutical compositions containing them, the use of such a compound for the manufacture of a pharmaceutical preparation for the treatment of neuropathic pain and to a method for the treatment of such a disease in animals, especially in humans.

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Pregnant rats treated with dimethadione (DMO), the N-demethylated metabolite of the anticonvulsant trimethadione, produce offspring having a 74% incidence of congenital heart defects (CHD); however, the incidence of CHD has high inter-litter variability (40-100%) that presents a challenge when studying the initiating events prior to the presentation of an abnormal phenotype. We hypothesized that the variability in CHD incidence was the result of differences in maternal systemic concentrations or embryonic tissue concentrations of DMO. To test this hypothesis, dams were administered 300. mg/kg DMO every 12. h from the evening of gestational day (GD) 8 until the morning of GD 11 (six total doses). Maternal serum levels of DMO were assessed on GD 11, 12, 13, 14, 15, 18 and 21. Embryonic tissue concentrations of DMO were assessed on GD 11, 12, 13 and 14. In a separate cohort of GD 12 embryos, DMO concentrations and parameters of growth and development were assessed to determine if tissue levels of DMO were correlated with these endpoints. Embryos were exposed directly to different concentrations of DMO with whole embryo culture (WEC) and their growth and development assessed. Key findings were that neither maternal systemic concentrations nor tissue concentrations of DMO identified embryos that were sensitive or resistant to DMO in vivo. Direct exposure of embryos to DMO via WEC also failed to show correlations between embryonic concentrations of DMO with developmental outcomes in vitro. We conclude that neither maternal serum nor embryonic tissue concentrations of DMO predict embryonic outcome.

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Oxazolidine – Wikipedia,
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Determination of mean whole body intracellular pH in unanesthetized dogs

In order to determine mean whole body pHi in unanesthetized dogs, assumptions on the urinary and intestinal excretion (5,5 dimethyloxazolidine, 2,4 dione) DMO, the total loss of an injected single dose of [14C]DMO, and the distribution kinetics of DMO were tested experimentally. Urinary excretion of DMO was almost negligible. The best assumption on the total loss of DMO was based on the exponential decay observed over a period of 1 to 4 weeks. The biological half life of DMO -0.14 5 days, the time constant being -0.14 d-1. The extracellular distribution of DMO was considered to equal that of [3H]inulin. Between 1 and 7 hours after an injection of inulin in nephrectomized dogs the distribution volume increased linearly from 16% of the body weight after 2 hours to 21% after 6 hours. Based on these experimental results pHi was determined in 16 unanesthetized dogs. 120 min after the injection of DMO pHi was 7.05 and 430 min after the injection pHi was 7.11. It is concluded that the assumptions made allow the estimation of pHi in unanesthetized dogs over a period of 1 to 7 hours.

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Effects of antiepileptic drugs on GABA responses and on reduction of GABA responses by PTZ and DMCM on mouse neurons in cell culture

The mechanisms of action of antiepileptic drugs effective against generalized absence seizures (antiabsence AEDs) remain uncertain. Antiabsence AEDs are generally effective against seizures induced in experimental animals by pentylenetetrazol (PTZ) and methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM), drugs which reduce GABAergic inhibition. Thus, antiabsence AEDs have been suggested to enhance GABAergic inhibition. We studied the effects of several AEDs on GABA responses recorded from mouse spinal cord neurons grown in primary dissociated cell culture. Four antiabsence AEDs were included: ethosuximide (ESM), dimethadione (DMO), sodium valproate (VPA), and diazepam (DZP). Two experimental AEDs, CGS 9896 and ZK 91296, with anticonvulsant action against PTZ- or DMCM-induced seizures were also included. Possible effects of the antiabsence and experimental AEDs on PTZ- and DMCM-induced inhibition of GABA responses were also evaluated. PTZ and DMCM reversibly reduced GABA responses in a concentration-dependent manner. PTZ completely inhibited GABA responses at 10 mM (IC50 of 1.1 mM), whereas DMCM-induced inhibition of GABA responses reached a plateau level of 39% of control values at 1 muM (IC50 of 33 nM). ESM (1,200 muM), DMO (6 mM), VPA (200 muM), CGS 9896 (2 muM), and ZK 91296 (2 muM) did not alter GABA responses. DZP enhanced GABA responses in a concentration-dependent manner. The inhibition of GABA responses produced by PTZ 1 mM was unaltered by ESM (600 muM), DMO (6 mM), CGS 9896 (1 muM), or ZK 91296 (1 muM). Coapplication of VPA (200 muM) and PTZ (1 mM) slightly enhanced the PTZ effect. DZP (>10 nM), however, reversed the PTZ-induced reduction of GABA responses. The DMCM (250 nM) inhibition of GABA responses was unaltered by ESM (600 muM), DMO (2 mM), or VPA (200 muM). CGS 9896 (2 muM) and ZK 91296 (2 muM), however, antagonized the DMCM effect. DZP (>10 nM) significantly reversed the DMCM-induced inhibition of GABA responses. The lack of effect of VPA, ESM, and DMO on postsynaptic GABA responses suggests that direct enhancement of postsynaptic GABA action is not a common mechanism of action of antiabsence AEDs. The AEDs DZP, CGS 9896, and ZK 91296 all reversed DMCM, but not PTZ, reduction of GABA responses, suggesting that these AEDs blocked DMCM seizures by acting at benzodiazepine receptors. However, since only DZP enhanced GABA responses, it is unclear how CGS 9896 and ZK 91296 blocked PTZ seizures.

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Zebrafish Models of Neurodevelopmental Disorders: Past, Present, and Future

Zebrafish are increasingly being utilized as a model system to investigate the function of the growing list of risk genes associated with neurodevelopmental disorders. This is due in large part to the unique features of zebrafish that make them an optimal system for this purpose, including rapid, external development of transparent embryos, which enable the direct visualization of the developing nervous system during early stages, large progenies, which provide considerable tractability for performing high-throughput pharmacological screens to identify small molecule suppressors of simple behavioral phenotypes, and ease of genetic manipulation, which has been greatly facilitated by the advent of CRISPR/Cas9 gene editing technologies. This review article focuses on studies that have harnessed these advantages of the zebrafish system for the functional analysis of genes that are strongly associated with the following neurodevelopmental disorders: autism spectrum disorders (ASD), epilepsy, intellectual disability (ID) and schizophrenia. We focus primarily on studies describing early morphological and behavioral phenotypes during embryonic and larval stages resulting from loss of risk gene function. We highlight insights into basic mechanisms of risk gene function gained from these studies as well as limitations of studies to date. Finally, we discuss advances in in vivo neural circuit imaging in zebrafish, which promise to transform research using the zebrafish model by illuminating novel circuit-level mechanisms with relevance to neurodevelopmental disorders.

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SYNTHESIS OF 13-AZA-16-OXA-, 13-AZA-16-THIA- AND 13,16-DIAZASTEROIDAL COMPOUNDS THROUGH CYCLIZATION OF N-ACYLIMINIUM ION INTERMEDIATES

5,5-Dimethyl-2,4-oxazolidinedione, 2,4-thiazolidinedione and 5,5-dimethylhydantoin were coupled with 2-(3,4-dihydro-1-naphthyl)ethyl alcohol by an application of Mitsunobu’s method to give the corresponding N-substituted products (5)-(7), respectively.Reduction of 5 – 7 with diisobutylaluminium hydride, followed by cyclization of the reduction products with formic acid yielded the 13-aza-16-oxa- (11), 13-aza-16-thia- (12) and 13,16-diaza-1,3,5(10),9(11)-estratetraen-17-one (13), respectively.

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Synthesis and spectroscopic properties of nickel complexes of benzo-, 1,2-naphtho-, or 2,3-naphthoannulated beta-oxatetraazachlorins

The first beta-oxatetraazachlorin derivatives have been synthesized starting from phthalonitrile derivatives and 5,5-dimethyl-1,3-oxazolidine-2,4- dione instead of the previously employed tetramethylsuccinonitrile. Absorption and magnetic circular dichroism (MCD) spectral properties are similar to those obtained for the corresponding tetramethyl-substituted TAC derivatives, confirming the presence of the low-symmetry aromatic structures.

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