Establishment and characterization of multiple patient-derived organoids from a case of advanced endometrial cancer.

Patient-derived organoids (PDOs) retain the original tumor's characteristics to a large degree and allow direct evaluation of the drug sensitivity, thereby emerging as a valuable resource for both basic and preclinical researches. Whereas most past studies stereotypically adopted a single PDO as an avatar of the patient, it remains to be investigated whether this assumption can be justified even for the tumor with spatial diversity. To address this issue, we established and characterized multiple PDOs originating from various sites of a patient with advanced uterine carcinosarcoma (UCS). Specifically, cancer cells were separately sampled from three sites; resected UCS tumor tissue, the peritoneal lavage fluid, and an intra-uterine brushing of the tumor. The three derived PDOs were morphologically undistinguishable, displaying typical carcinoma organoids-like appearance, but two of them proliferated at a faster rate. The primary tumor harbored mutations in TP53 and STK11 along with amplifications in CCNE1, ERBB2, and KRAS. These two mutations and the CCNE1 amplification were detected in all PDOs, while either KRAS or ERBB2 amplification was selectively observed in each PDO in a mutually exclusive manner. Observed intra-tumor heterogeneity in HER2 expression was differentially reproduced in the PDOs, which mirrored each PDO's sensitivity to HER2 inhibitors. Inter-PDO heterogeneity was also evident in sensitivity to standard cytotoxic agents. Lastly, a drug screening identified four candidate reagents commonly effective to all PDOs. Collectively, we showed that multiple PDOs could help reproduce the spatial diversity of a tumor and serve as a valuable resource in UCS research in many respects.

Role of serine racemase in behavioral sensitization in mice after repeated administration of methamphetamine.

BACKGROUND: The N-methyl-D-aspartate (NMDA) receptors play a role in behavioral abnormalities observed after administration of the psychostimulant, methamphetamine (METH). Serine racemase (SRR) is an enzyme which synthesizes D-serine, an endogenous co-agonist of NMDA receptors. Using Srr knock-out (KO) mice, we investigated the role of SRR on METH-induced behavioral abnormalities in mice. METHODOLOGY/PRINCIPAL FINDINGS: Evaluations of behavior in acute hyperlocomotion, behavioral sensitization, and conditioned place preference (CPP) were performed. The role of SRR on the release of dopamine (DA) in the nucleus accumbens after administration of METH was examined using in vivo microdialysis technique. Additionally, phosphorylation levels of ERK1/2 proteins in the striatum, frontal cortex and hippocampus were examined using Western blot analysis. Acute hyperlocomotion after a single administration of METH (3 mg/kg) was comparable between wild-type (WT) and Srr-KO mice. However, repeated administration of METH (3 mg/kg/day, once daily for 5 days) resulted in behavioral sensitization in WT, but not Srr-KO mice. Pretreatment with D-serine (900 mg/kg, 30 min prior to each METH treatment) did not affect the development of behavioral sensitization after repeated METH administration. In the CPP paradigm, METH-induced rewarding effects were demonstrable in both WT and Srr-KO mice. In vivo microdialysis study showed that METH (1 mg/kg)-induced DA release in the nucleus accumbens of Srr-KO mice previously treated with METH was significantly lower than that of the WT mice previously treated with METH. Interestingly, a single administration of METH (3 mg/kg) significantly increased the phosphorylation status of ERK1/2 in the striatum of WT, but not Srr-KO mice. CONCLUSIONS/SIGNIFICANCE: These findings suggest first, that SRR plays a role in the development of behavioral sensitization in mice after repeated administration of METH, and second that phosphorylation of ERK1/2 by METH may contribute to the development of this sensitization as seen in WT but not Srr-KO mice.

Levels of D-serine in the brain and peripheral organs of serine racemase (Srr) knock-out mice.

D-Serine, an endogenous co-agonist of the N-methyl-D-aspartate (NMDA) receptor, plays an important role in mammalian brain neurotransmission, via the NMDA receptor. D-Serine is synthesized from L-serine by the pyridoxal-5' phosphate-dependent enzyme serine racemase (SRR), and D-serine is metabolized by D-amino acid oxidase (DAAO). In this study, we measured levels of the neurotransmission related amino acids, d-serine, L-serine, glycine, glutamine and glutamate in the frontal cortex, hippocampus, striatum and cerebellum as well as in peripheral tissues of blood, heart, pancreas, spleen, liver, kidney, testis, epididymis, heart, lung, muscle and eyeball, in wild-type (WT) and Srr-knockout (Srr-KO) mice. Levels of D-serine in the frontal cortex, hippocampus, and striatum of Srr-KO mice were significantly lower than in WT mice, while levels in the cerebellum stayed the same. In contrast, levels of L-serine, glycine, glutamine and glutamate remained the same in all tested brain regions. In vivo microdialysis using free-moving mice showed that extracellular levels of D-serine in the hippocampus of Srr-KO mice were significantly lower than in WT mice while the other amino acid levels remained the same between mice. In peripheral organs, levels of D-serine in the kidney, testis, and muscle of Srr-KO mice were significantly lower than in WT mice. Tissue levels of the other tested amino acids in peripheral organs were not altered. These results suggest that SRR is the major enzyme responsible for D-serine production in the mouse forebrain, and that other pathways of d-serine production may exist in the brain and peripheral organs.

Effects of aripiprazole on MK-801-induced prepulse inhibition deficits and mitogen-activated protein kinase signal transduction pathway.

Based on NMDA hypofunction hypothesis for negative symptoms and cognitive deficits in schizophrenia, MK-801-induced animal models of schizophrenia may help us understand the different effects between typical and atypical antipsychotics. On the other hand, the mitogen-activated protein kinase (MAPK) signaling pathways may participate in antipsychotic actions. The aim of this study was to investigate the effects of aripiprazole on MK-801-induced prepulse inhibition (PPI) disruption and MAPK phosphorylation in mice. To clarify the effects of aripiprazole on MK-801-induced PPI disruption, we measured PPI of 51 ddY male mice after aripiprazole was administered 15 min prior to the injection of MK-801, and measured activation of cytosol and nuclear MAPK phosphorylation by western blotting. Aripiprazole (4.0 mg/kg) significantly reversed the MK-801 (0.15 mg/kg)-induced PPI deficits. Pretreatment of aripiprazole (40 mg/kg) had a tendency to suppress MK-801 (1.0 mg/kg)-induced pMEK/MEK (Ser218/222) activation. In addition, aripiprazole treatment showed a significant decrease of pERK/ERK. Our data suggested that aripiprazole may reverse MK-801-induced PPI deficits through regulation of MAPK phosphorylation in the same way as the atypical antipsychotic drug, clozapine.

Effects of quetiapine on phencyclidine-induced cognitive deficits in mice: a possible role of alpha1-adrenoceptors.

Accumulating evidence suggests that alpha(1)-adrenoceptors may be involved in the mechanisms of action of some antipsychotic drugs. The present study was undertaken to examine the effects of quetiapine, an atypical antipsychotic drug with alpha(1)-adrenoceptor antagonism, on cognitive deficits in mice after repeated administration of the NMDA receptor antagonist phencyclidine (PCP). Subsequent subchronic (14 days) administration of quetiapine (1.0, 10, or 30 mg/kg, p.o.) attenuated PCP (10 mg/kg/day for 10 days)-induced cognitive deficits in mice, in a dose dependent manner. Furthermore, PCP (10 mg/kg)-induced cognitive deficits were also significantly ameliorated by subsequent subchronic (14 days) administration of the selective alpha(1)-adrenoceptor antagonist prazosin (1.0 mg/kg/day, p.o.). Moreover, Western blot analysis revealed that levels of two subtypes (alpha(1A) and alpha(1B)) of alpha(1)-adrenoceptors were significantly lower in the brains of PCP-treated mice than in those of saline-treated mice. These findings suggest that repeated PCP administration could decrease the density of alpha(1)-adrenoceptors in mouse brain, and that subsequent subchronic administration of quetiapine might ameliorate PCP-induced cognitive deficits via alpha(1)-adrenoceptors. Therefore, it is likely that antagonism at alpha(1)-adrenoceptors is involved in the mechanism underlying quetiapine's psychopharmacological action.

Phencyclidine-induced cognitive deficits in mice are ameliorated by subsequent subchronic administration of donepezil: role of sigma-1 receptors.

This study was undertaken to examine the effects of two acetylcholinesterase inhibitors (donepezil and physostigmine) on cognitive deficits in mice after repeated administration of the NMDA receptor antagonist phencyclidine (PCP). In the novel object recognition test, PCP (10 mg/kg/day for 10 days)-induced cognitive deficits were significantly improved by subsequent subchronic (14 days) administration of donepezil (1.0 mg/kg/day), but not donepezil (0.1 mg/kg/day). Furthermore, the effect of donepezil (1.0 mg/kg/day) on PCP-induced cognitive deficits was significantly antagonized by co-administration of the selective sigma-1 receptor antagonist NE-100 (1.0 mg/kg/day), suggesting the role of sigma-1 receptors in the active mechanisms of donepezil. In contrast, PCP-induced cognitive deficits were not improved by subsequent subchronic (14 days) administration of physostigmine (0.25 mg/kg/day). Moreover, repeated administration of PCP significantly caused the reduction of sigma-1 receptors in the hippocampus. The present study suggests that agonistic activity of donepezil at sigma-1 receptors plays a role in the active mechanisms of donepezil on PCP-induced cognitive deficits in mice. Therefore, it is likely that donepezil would be potential therapeutic drugs for the treatment of the cognitive deficits in schizophrenia.

Fluorescence response of a 4-trifluoroacetylaminophthalimide to iodide ions upon 254 nm irradiation in MeCN.

The title trifluoroacetylaminophthalimide derivative produced a violet fluorescence (lambdaFLmax 392 nm) in MeCN, and it displayed a green emission (lambdaFLmax 506 nm) after irradiation at 254 nm in the presence of iodide ions. The corresponding amidate ion of the trifluoroacetamide was identified as the green fluorescence emitter. The deprotonation reaction may be caused by proton-abstracting solvated electrons generated by a photochemical charge--transfer-to-solvent process from I(-) to MeCN.

Glycine and D: -serine, but not D: -cycloserine, attenuate prepulse inhibition deficits induced by NMDA receptor antagonist MK-801.

RATIONALE: Several agents that stimulate the glycine site of N-methyl-D: -aspartate (NMDA) receptors have been reported to moderately improve both negative symptoms and cognitive dysfunctions in patients with schizophrenia. However, differences in efficacy have also been reported, and further comparative pharmacological studies are still needed. OBJECTIVES: We aimed to explore the effects of two glycine site agonists of the NMDA receptor, glycine and D: -serine, and a partial agonist, D: -cycloserine, on prepulse inhibition (PPI) deficits induced by a NMDA receptor antagonist, MK-801, in mice. Furthermore, we performed in vivo microdialysis and additional PPI measurements using a selective glycine site antagonist to verify if the beneficial effects observed after the systemic administration of glycine were due to glycine itself via its activity at the glycine site. RESULTS: High doses of glycine (1.6 g/kg) and D: -serine (1.8 and 2.7 g/kg) significantly attenuated MK-801-induced PPI deficits. In contrast, D: -cycloserine did not show any amelioration of MK-801-induced PPI deficits at doses ranging from 7.5 mg/kg to 60 mg/kg. The selective glycine site antagonist, L-701,324 (10 mg/kg), antagonized the effect of glycine on MK-801-induced PPI deficits. Furthermore, in vivo microdialysis demonstrated that intraperitoneal injection of glycine significantly increased glycine and L: -serine levels, but decreased D: -serine levels in the prefrontal cortex. CONCLUSIONS: The findings of the present study suggest that glycine and D: -serine but not D: -cycloserine could attenuate PPI deficits associated with NMDA receptor hypofunction via NMDA glycine sites in the brain.