DETQ, an allosteric potentiator of the dopamine D1 receptor, was tested in therapeutic models that were known to respond to D1 agonists. Because of a species difference in affinity for DETQ, all ...rodent experiments used transgenic mice expressing the human D1 receptor (hD1 mice). When given alone, DETQ reversed the locomotor depression caused by a low dose of reserpine. DETQ also acted synergistically with L-DOPA to reverse the strong hypokinesia seen with a higher dose of reserpine. These results indicate potential as both monotherapy and adjunct treatment in Parkinson's disease. DETQ markedly increased release of both acetylcholine and histamine in the prefrontal cortex, and increased levels of histamine metabolites in the striatum. In the hippocampus, the combination of DETQ and the cholinesterase inhibitor rivastigmine increased ACh to a greater degree than either agent alone. DETQ also increased phosphorylation of the AMPA receptor (GluR1) and the transcription factor CREB in the striatum, consistent with enhanced synaptic plasticity. In the Y-maze, DETQ increased arm entries but (unlike a D1 agonist) did not reduce spontaneous alternation between arms at high doses. DETQ enhanced wakefulness in EEG studies in hD1 mice and decreased immobility in the forced-swim test, a model for antidepressant-like activity. In rhesus monkeys, DETQ increased spontaneous eye-blink rate, a measure that is known to be depressed in Parkinson's disease. Together, these results provide support for potential utility of D1 potentiators in the treatment of several neuropsychiatric disorders, including Parkinson's disease, Alzheimer's disease, cognitive impairment in schizophrenia, and major depressive disorder.
•The dopamine D1 potentiator DETQ was tested in humanized D1 mice and rhesus monkeys.•Actions of DETQ were dependent on endogenous dopaminergic tone.•DETQ displayed a behavioral profile consistent with central D1 receptor activation.•Neurochemical actions of DETQ support potential pro-cognitive effects.•D1 potentiators show promise for Parkinson's disease and other CNS disorders.
ABSTRACT
Fanconi anemia (FA) is a rare inherited disorder caused by pathogenic variants in one of 19 FANC genes. FA patients display congenital abnormalities, and develop bone marrow failure, and ...cancer susceptibility. We identified homozygous mutations in four FA patients and, in each case, only one parent carried the obligate mutant allele. FANCA and FANCP/SLX4 genes, both located on chromosome 16, were the affected recessive FA genes in three and one family respectively. Genotyping with short tandem repeat markers and SNP arrays revealed uniparental disomy (UPD) of the entire mutation‐carrying chromosome 16 in all four patients. One FANCA patient had paternal UPD, whereas FA in the other three patients resulted from maternal UPD. These are the first reported cases of UPD as a cause of FA. UPD indicates a reduced risk of having another child with FA in the family and has implications in prenatal diagnosis.
We report evidence for UPD as a cause of Fanconi anemia(FA), a rare, mostly recessive disorder. We discovered patients with homozygous pathogenic variants in FANCA and FANCP/SLX4, where only one of their parents was a carrier. Using high density SNP arrays, and short tandem repeat markers (inverted triangles), we identified these patients lack a contribution of the entire chromosome 16 from one of their parents, and present homozygosity of the inherited mutant region from the other parent.
Context:
Targeted secretion inhibitors (TSIs), a new class of recombinant biotherapeutic proteins engineered from botulinum toxin, represent a novel approach for treating diseases with excess ...secretion. They inhibit hormone secretion from targeted cell types through cleavage of SNARE (soluble N-ethylmaleimide-sensitive factor-activating protein receptor) proteins. qGHRH-LHN/D is a TSI targeting pituitary somatotroph through binding to the GHRH-receptor and cleavage of the vesicle-associated membrane protein (VAMP) family of SNARE proteins.
Objective:
Our objective was to study SNARE protein expression in pituitary adenomas and to inhibit GH secretion from somatotropinomas using qGHRH-LHN/D.
Design:
We analyzed human pituitary adenoma analysis for SNARE expression and response to qGHRH-LHN/D treatment.
Setting:
The study was conducted in University Hospitals.
Patients:
We used pituitary adenoma samples from 25 acromegaly and 47 nonfunctioning pituitary adenoma patients.
Outcome:
Vesicle-SNARE (VAMP1–3), target-SNARE (syntaxin1, SNAP-23, and SNAP-25), and GHRH-receptor detection with RT-qPCR, immunocytochemistry, and immunoblotting. Assessment of TSI catalytic activity on VAMPs and release of GH from adenoma cells.
Results:
SNARE proteins were variably expressed in pituitary samples. In vitro evidence using recombinant GFP-VAMP2&3 or pituitary adenoma lysates suggested sufficient catalytic activity of qGHRH-LHN/D to degrade VAMPs, but was unable to inhibit GH secretion in somatotropinoma cell cultures.
Conclusions:
SNARE proteins are present in human pituitary somatotroph adenomas that can be targeted by TSIs to inhibit GH secretion. qGHRH-LHN/D was unable to inhibit GH secretion from human somatotroph adenoma cells. Further studies are required to understand how the SNARE proteins drive GH secretion in human somatotrophs to allow the development of novel TSIs with a potential therapeutic benefit.
Starting from a novel, achiral 1,3,4-benzotriazepine-based CCK2 receptor antagonist, a process of optimization has afforded further compounds of this type that maintain the nanomolar affinity for ...recombinant, human CCK2 receptors and high selectivity over CCK1 receptors observed in the initial lead but display more potent inhibition of pentagastrin-stimulated gastric acid secretion in vivo. Moreover, this has largely been achieved without altering their potency at wild-type canine and rat receptors, as judged by their displacement of 125I-BH-CCK-8S in a radioligand binding assay and by their activity in an isolated, perfused rat stomach bioassay, respectively. 2-(5-Cyclohexyl-1-(2-cyclopentyl-2-oxo-ethyl)-2-oxo-1,2-dihydro-3H-1,3,4-benzotriazepin-3-yl)-N-(3-(5-oxo-2,5-dihydro- 1,2,4oxadiazol-3-yl)-phenyl)-acetamide (47) was identified as the most effective compound stemming from this approach, proving to be a potent inhibitor of pentagastrin-stimulated gastric acid secretion in rats and dogs by intravenous bolus as well as by enteral administration.
A 1,3,4-benzotriazepine was identified as a suitable lead in our effort toward obtaining a non-peptide parathyroid hormone-1 receptor (PTH1R) antagonist. A process of optimization afforded ...derivatives displaying nanomolar PTH1R affinity, a representative example of which behaved as a PTH1R antagonist in cell-based cyclic adenosine monophosphate (cAMP) assays, with selectivity over PTH2 receptors.
Novel, achiral 1H-1,3,5-benzotriazepine-2,4(3H,5H)-diones have been prepared and structurally characterized. These compounds are potent CCK(2) receptor antagonists that display a high degree of ...selectivity over CCK(1) receptors.
Studies have shown that measurement of thermodynamic parameters (enthalpy, DeltaH degrees and entropy, DeltaS degrees ) can allow discrimination of agonists and antagonists (e.g. Weiland, G.A., ...Minneman, K.P., Molinoff, P.B., 1979. Fundamental difference between the molecular interactions of agonists and antagonists with the beta-adrenergic receptor. Nature, 281, 114.). Recently, we found that agonists and antagonists were not thermodynamically-distinguished at cholecystokinin (CCK)2-receptors in rat cerebral cortex. However, in this study, the possibility that thermodynamic discrimination at CCK2-receptors exists but that it was not detected, could not be excluded because radioligand binding studies and functional assays were performed in different rat tissues. Therefore, we have repeated these studies using the recombinant CCK2 short isoform (CCK2S)-receptor expressed in NIH3T3 cells, so that ligand affinity (pKI) and intrinsic activity (alpha) measurements could be made in exactly the same receptor system. CCK-8S but not R-L-365,260, S-L-365,260, JB95008, JB93242 or PD134,308 expressed intrinsic activity in an IP assay. The pKD of 3H-JB93182 decreased with increasing temperature. pKI values for antagonists (R-L-365,260, S-L-365,260, JB95008) and agonists (pentagastrin, CCK-8S) were higher at 4 than at 30 degrees C. There was no effect of temperature on pKI values for the antagonists, PD134,308 and JB93242. Therefore, CCK2-receptor agonists and antagonists at human CCK2S-receptors cannot be discriminated by thermodynamic analysis.
A series of 1,3,4-benzotriazepine-based CCK2 antagonists have been devised by consideration of the structural features that govern CCK receptor affinity and the receptor subtype selectivity of ...1,4-benzodiazepine-based CCK2 antagonists. In contrast to the latter compounds, these novel 1,3,4-benzotriazepines are achiral, yet they display similar affinity for CCK2 receptors to the earlier molecules and are highly selective over CCK1 receptors.
Novel, potent, and selective non-imidazole histamine H3 receptor antagonists have been prepared based on the low-affinity ligand dimaprit (pK I 7.32 ± 0.12, pK B 5.93 ± 0.17). Detailed ...structure−activity studies have revealed that N-(4-chlorobenzyl)-N-(6-pyrrolidin-1-ylhexyl)guanidine (pK I 8.38 ± 0.21, pK B 8.39 ± 0.13), 30, and N-(4-chlorobenzyl)-N-(7-pyrrolidin-1-ylheptyl)guanidine (pK I 8.78 ± 0.12, pK B 8.38 ± 0.10), 31, exhibit high affinity for the histamine H3 receptor. Antagonists 30 and 31 demonstrate significant selectivity over the other histamine, H1 and H2, receptor subtypes and a 100-fold selectivity in the σ1 binding assay. Compounds 30 and 31 are the most potent, selective non-imidazole histamine H3 receptor antagonists reported in the literature to date.
The systematic optimization of the structure of a novel 2,4,5-trisubstituted imidazole-based cholecystokinin-2 (CCK2) receptor antagonist afforded analogues with nanomolar receptor affinity. These ...compounds were now comparable in their potency to the bicyclic heteroaromatic-based compounds 5 (JB93182) and 6 (JB95008), from which the initial examples were designed using a field-point based molecular modeling approach. They were also orally active as judged by their inhibition of pentagastrin stimulated acid secretion in conscious dogs, in contrast to the bicyclic heteroaromatic-based compounds, which were ineffective because of biliary elimination. Increasing the hydrophilicity through replacement of a particular methylene group with an ether oxygen, as in 3-{5-(adamantan-1-yloxymethyl)-2-cyclohexyl-1H-imidazole-4-carbonylamino}benzoic acid (53), had little effect on the receptor affinity but significantly increased the oral potency. Comparison of the plasma pharmacokinetics and the inhibition of pentagastrin-stimulated acid output following bolus intraduodenal administration of both 53 and 6 indicated that 53 was well absorbed, had a longer half-life, and was not subject to the elimination pathways of the earlier series.
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