A2A Adenosine Receptor Antagonists in Neurodegenerative Diseases

Background: Alzheimer's disease (AD) is the most common form of dementia worldwide, with approximately 6 million American cases in 2020. The clinical signs of AD include cognitive dysfunction, apathy, anxiety and neuropsychiatric signs, and pathogenetic mechanisms that involve amyloid peptide-β extracellular accumulation and tau hyperphosphorylation. Unfortunately, current drugs to treat AD can provide only symptomatic relief but are not disease-modifying molecules able to revert AD progression. The endogenous modulator adenosine, through A2A receptor activation, plays a role in synaptic loss and neuroinflammation, which are crucial for cognitive impairment and memory damage. Objective: In this review, recent advances covering A2A adenosine receptor antagonists will be extensively reviewed, providing a base for the rational design of future A2A inhibitors. Method: Herein, the literature on A2A adenosine receptors and their role in synaptic plasticity and neuroinflammation as well as the effects of A2A antagonism in animal models of AD and in humans are reviewed. Furthermore, current chemical and structure-based strategies are presented. Results : Caffeine, the most widely consumed natural product stimulant and an A2A antagonist, improves human memory. Similarly, synthetic A2A receptor antagonists, as described in this review, may provide a means to fight AD. Conclusion: This review highlights the clinical potential of A2A adenosine receptor antagonists as a novel approach to treat patients with AD.

Antioxidant and Antiinflammatory Effects of Epilobium parviflorum, Melilotus officinalis and Cardiospermum halicacabum Plant Extracts in Macrophage and Microglial Cells

Background: We investigated the phenolic content characterizing different plant extracts from Epilobium parviflorum, Cardiospermum halicacabum, and Melilotus officinalis, their antioxidant, antiinflammatory effects, and their mechanism of action. Methods: plant samples were macerated in 40% ethanol or hot/ cold glycerate and assessed for polyphenols content. The antioxidant activity was investigated by DPPH radical scavenging assay and H2DCFDA test in LPS-stimulated RAW264.7 macrophages and N9 microglial cells. MTS experiments and antiinflammatory properties verified cellular toxicity through NO assay. Interaction with A2A adenosine receptors was evaluated through binding assays using [3H]ZM241385 radioligand. Results: Polyphenols were present in 40% ethanol plant extract, which at 0.1–10 µg/µL achieved good antioxidant effects, with a DPPH radical scavenging rate of about 90%. In LPS-stimulated cells, these plant extracts, at 1μg/μL, did not affect cell vitality, displayed significant inhibition of H2DCFDA and NO production, and inhibited ZM 241385 binding in CHO cells transfected with A2A receptors. RAW 264.7 and N9 cells presented a density of them quantified in 60 ± 9 and 45 ± 5 fmol/mg of protein, respectively. Conclusion: Epilobium parviflorum, Cardiospermum halicacabum, and Melilotus officinalis extracts may be considered a source of agents for treating disorders related to oxidative stress and inflammation.

Locating ligand binding sites in G-protein coupled receptors using combined information from docking and sequence conservation

GPCRs (G-protein coupled receptors) are the largest family of drug targets and share a conserved structure. Binding sites are unknown for many important GPCR ligands due to the difficulties of GPCR recombinant expression, biochemistry, and crystallography. We describe our approach, ConDockSite, for predicting ligand binding sites in class A GPCRs using combined information from surface conservation and docking, starting from crystal structures or homology models. We demonstrate the effectiveness of ConDockSite on crystallized class A GPCRs such as the beta2 adrenergic and A2A adenosine receptors. We also demonstrate that ConDockSite successfully predicts ligand binding sites from high-quality homology models. Finally, we apply ConDockSite to predict the ligand binding sites on a structurally uncharacterized GPCR, GPER, the G-protein coupled estrogen receptor. Most of the sites predicted by ConDockSite match those found in other independent modeling studies. ConDockSite predicts that four ligands bind to a common location on GPER at a site deep in the receptor cleft. Incorporating sequence conservation information in ConDockSite overcomes errors introduced from physics-based scoring functions and homology modeling.

Adenosine receptors in GtoPdb v.2021.2

Adenosine receptors (nomenclature as agreed by the NC-IUPHAR Subcommittee on Adenosine Receptors [110]) are activated by the endogenous ligand adenosine (potentially inosine also at A3 receptors). Crystal structures for the antagonist-bound [153, 313, 221, 61], agonist-bound [375, 203, 204] and G protein-bound A2A adenosine receptors [49] have been described. The structures of an antagonist-bound A1 receptor [128] and an adenosine-bound A1 receptor-Gi complex [86] have been resolved by cryo-electronmicroscopy. Another structure of an antagonist-bound A1 receptor obtained with X-ray crystallography has also been reported [57]. caffeine is a nonselective antagonist for adenosine receptors, while istradefylline, a selective A2A receptor antagonist, is on the market for the treatment of Parkinson's disease.

Combined Therapy of A1AR Agonists and A2AAR Antagonists in Neuroinflammation

Alzheimer’s, Parkinson’s, and multiple sclerosis are neurodegenerative diseases related by neuronal degeneration and death in specific areas of the central nervous system. These pathologies are associated with neuroinflammation, which is involved in disease progression, and halting this process represents a potential therapeutic strategy. Evidence suggests that microglia function is regulated by A1 and A2A adenosine receptors (AR), which are considered as neuroprotective and neurodegenerative receptors, respectively. The manuscript’s aim is to elucidate the role of these receptors in neuroinflammation modulation through potent and selective A1AR agonists (N6-cyclopentyl-2′- or 3′-deoxyadenosine substituted or unsubstituted in 2 position) and A2AAR antagonists (9-ethyl-adenine substituted in 8 and/or in 2 position), synthesized in house, using N13 microglial cells. In addition, the combined therapy of A1AR agonists and A2AAR antagonists to modulate neuroinflammation was evaluated. Results showed that A1AR agonists were able, to varying degrees, to prevent the inflammatory effect induced by cytokine cocktail (tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and interferon (IFN)-γ), while A2AAR antagonists showed a good ability to counteract neuroinflammation. Moreover, the effect achieved by combining the two most effective compounds (1 and 6) in doses previously found to be non-effective was greater than the treatment effect of each of the two compounds used separately at maximal dose.

Advantages in Wound Healing Process in Female Mice Require Upregulation A2A-Mediated Angiogenesis under the Stimulation of 17β-Estradiol

Estrogenic steroids and adenosine A2A receptors promote the wound healing and angiogenesis processes. However, so far, it is unclear whether estrogen may regulate the expression and pro-angiogenic activity of A2A receptors. Using in vivo analyses, we showed that female wild type (WT) mice have a more rapid wound healing process than female or male A2A-deficient mice (A2AKO) mice. We also found that pulmonary endothelial cells (mPEC) isolated from female WT mice showed higher expression of A2A receptor than mPEC from male WT mice. mPEC from female WT mice were more sensitive to A2A-mediated pro-angiogenic response, suggesting an ER and A2A crosstalk, which was confirmed using cells isolated from A2AKO. In those female cells, 17β-estradiol potentiated A2A-mediated cell proliferation, an effect that was inhibited by selective antagonists of estrogen receptors (ER), ERα, and ERβ. Therefore, estrogen regulates the expression and/or pro-angiogenic activity of A2A adenosine receptors, likely involving activation of ERα and ERβ receptors. Sexual dimorphism in wound healing observed in the A2AKO mice process reinforces the functional crosstalk between ER and A2A receptors.