Identification of novel selective P2Y 6 receptor antagonists by high-throughput screening assay

Ito, Masaaki; Egashira, Shinichiro; Yoshida, Kazuki; Mineno, Tomoko; Kumagai, Kazuo; Kojima, Hirotatsu; Okabe, Takayoshi; Nagano, Tetsuo; Ui, Michio; Matsuoka, Isao

doi:10.1016/j.lfs.2017.05.017

Cited by 25 publications

(11 citation statements)

References 23 publications

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“…Moreover, we calculated z'-factor, a measure of robustness of assay, and we found a value of 0.645, which indicates that our protocol was adequate for use. Ito and coworkers (2017) performed a high-throughput screening assay to identify novel antagonists for P2Y6R using P2Y6-1321N1 cells and obtained a z'-factor of 0.80 [30]. Valentin et al (2011) used primary cells (Huvec) to perfume high-throughput screening based on calcium measurement and obtained a z'-factor of 0.60 [41].…”

Section: Discussionmentioning

confidence: 99%

“…They found that some flavonoids could inhibit P2Y2R and suggest the use of their structures to develop new compounds with antagonistic activity [29]. Recently, Ito et al applied a protocol of intracellular calcium measurement in a high-throughput screening campaign to search novel antagonists for P2Y6 receptor and found one compound, TIM-38, with potential activity [30].…”

Section: Introductionmentioning

confidence: 99%

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New Insights in Purinergic Therapy: Novel Antagonists for Uridine 5′-Triphosphate-Activated P2Y Receptors from Brazilian Flora

Ferreira

Soares‐Bezerra

Silveira

et al. 2019

Journal of Medicinal Food

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P2Y2 and P2Y4 receptors are physiologically activated by UTP and are widely expressed in many cell types in humans. They promote an increase in intracellular calcium via PLCβ/ IP3 and act on ion flux and water secretion. P2Y2 plays an important role in inflammation and proliferation of tumor cells, which could be attenuated with the use of antagonists. However, little is known about the physiological functions related to P2Y4 due to the lack of selective ligands for these receptors, which can be solved through the search of novel compounds with antagonistic activity. In the present study, we have applied a methodology of calcium measurement to identify new antagonist candidates for these receptors. Firstly, we established optimal conditions for calcium assay using J774.G8, a murine macrophage cell line, which expresses functional P2Y2 and P2Y4 receptors. J774.G8 cells were loaded with 2 μM of Fluo-4 to test its sensitivity in responding calcium stimuli. ATP and ionomycin, known as inductors of intracellular calcium rise, were used to stimulate cells. The EC50 obtained were 11 μM and 103 nM, respectively. Subsequently, investigation of P2Y2 and P2Y4 expression was performed. These cells responded with EC50 of 1.021 μM to the UTP stimulation. Screening assays were performed and a total of 100 extracts from Brazilian natural products were tested. JA2, RA3, and RB3 extracts stood out for their ability to inhibit UTP-induced responses without causing cytotoxicity and presented IC50 of 32.32 μg/mL, 14.99 μg/mL, and 12.98 μg/mL, respectively. Collectively, our results point to the discovery of potential antagonists candidates from natural products for UTPactivated receptors.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

New Insights in Purinergic Therapy: Novel Antagonists for Uridine 5′-Triphosphate-Activated P2Y Receptors from Brazilian Flora

Ferreira

Soares‐Bezerra

Silveira

et al. 2019

Journal of Medicinal Food

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“…285,286 Recently, a novel selective hP2Y 6 R antagonist TIM-38 was reported with low potency (IC 50 ¼ 4.3 mM). 287 TIM-38 could be a useful pharmacological tool and a starting point for the development of therapeutic agents against P2Y 6 receptor-implicated disease. Activation of P2Y 6 R by either its endogenous ligand UDP or selective agonist MRS-2693 has shown to promote production of proinflammatory cytokines IL-6 and IL-8 and contribute to phagocytosis of neurons.…”

Section: P2y 6 Receptor and Functions In The Cnsmentioning

confidence: 99%

Purinergic Receptors of the Central Nervous System: Biology, PET Ligands, and Their Applications

Zarrinmayeh

Territo

2020

Mol Imaging

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Purinergic receptors play important roles in central nervous system (CNS). These receptors are involved in cellular neuroinflammatory responses that regulate functions of neurons, microglial and astrocytes. Based on their endogenous ligands, purinergic receptors are classified into P1 or adenosine, P2X and P2Y receptors. During brain injury or under pathological conditions, rapid diffusion of extracellular adenosine triphosphate (ATP) or uridine triphosphate (UTP) from the damaged cells, promote microglial activation that result in the changes in expression of several of these receptors in the brain. Imaging of the purinergic receptors with selective Positron Emission Tomography (PET) radioligands has advanced our understanding of the functional roles of some of these receptors in healthy and diseased brains. In this review, we have accumulated a list of currently available PET radioligands of the purinergic receptors that are used to elucidate the receptor functions and participations in CNS disorders. We have also reviewed receptors lacking radiotracer, laying the foundation for future discoveries of novel PET radioligands to reveal these receptors roles in CNS disorders.

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“…16–17,18,19,20,21,22,23,24,25,26 Also, a few P2Y 6 R antagonists have reported, but none approaching nM affinity. 2,27,28 Both mononucleotides (mainly UDP and its analogues, e.g. 1 – 5 , 8 , 9a , 10 , 11 and 13 ) and dinucleotides (mainly dinucleoside triphosphates, e.g.…”

Section: Introductionmentioning

confidence: 99%

“…36 The P2Y 6 R can induce both vasoconstrictor and vasodilator effects, 4 which might complicate the use of its ligands therapeutically. Thus, there is a need for more potent and selective P2Y 6 R ligands, both agonists and antagonists, 27 as receptor probes and potential drug candidates.…”

Section: Introductionmentioning

confidence: 99%

Pyrimidine nucleotides containing a (S)-methanocarba ring as P2Y₆ receptor agonists

et al. 2017

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Both agonists and antagonists of the UDP-activated P2Y6 receptor (P2Y6R) have been proposed for therapeutic use, in conditions such as cancer, inflammation, neurodegeneration and diabetes. Uracil nucleotides containing a South-bicyclo[3.1.0]hexane ((S)-methanocarba) ring system in place of the ribose ring were synthesized and shown to be potent P2Y6R agonists in a calcium mobilization assay. The (S)-methanocarba modification was compatible with either a 5-iodo or 4-methoxyimino group on the pyrimidine, but not with a α,β-methylene 5´-diphosphate. (S)-Methanocarba dinucleotide potency was compatible with a N4-methoxy modification on the proximal nucleoside that is assumed to bind at the P2Y6R similarly to UDP; (N)-methanocarba was preferred on the distal nucleoside moiety. This suggests that the distal dinucleotide P2Y6R binding site prefers a ribose-like group that can attain a (N) conformation, rather than (S). Dinucleotide binding was modeled by homology modeling, docking and molecular dynamics simulations, which suggested the same ribose conformational preferences found empirically.

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Identification of novel selective P2Y 6 receptor antagonists by high-throughput screening assay

Cited by 25 publications

References 23 publications

New Insights in Purinergic Therapy: Novel Antagonists for Uridine 5′-Triphosphate-Activated P2Y Receptors from Brazilian Flora

New Insights in Purinergic Therapy: Novel Antagonists for Uridine 5′-Triphosphate-Activated P2Y Receptors from Brazilian Flora

Purinergic Receptors of the Central Nervous System: Biology, PET Ligands, and Their Applications

Pyrimidine nucleotides containing a (S)-methanocarba ring as P2Y₆ receptor agonists

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Identification of novel selective P2Y 6 receptor antagonists by high-throughput screening assay

Cited by 25 publications

References 23 publications

New Insights in Purinergic Therapy: Novel Antagonists for Uridine 5′-Triphosphate-Activated P2Y Receptors from Brazilian Flora

New Insights in Purinergic Therapy: Novel Antagonists for Uridine 5′-Triphosphate-Activated P2Y Receptors from Brazilian Flora

Purinergic Receptors of the Central Nervous System: Biology, PET Ligands, and Their Applications

Pyrimidine nucleotides containing a (S)-methanocarba ring as P2Y6 receptor agonists

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Product

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About

Pyrimidine nucleotides containing a (S)-methanocarba ring as P2Y₆ receptor agonists