Progress in the discovery of selective, high affinity A2B adenosine receptor antagonists as clinical candidates

The selective, high affinity A2B adenosine receptor (AdoR) antagonists that were synthesized by several research groups should aid in determining the role of the A2B AdoR in inflammatory diseases like asthma or rheumatoid arthritis (RA) and angiogenic diseases like diabetic retinopathy or cancer. CV Therapeutics scientists discovered the selective, high affinity A2B AdoR antagonist 10, a 8-(4-pyrazolyl)-xanthine derivative [CVT-6883, Ki(hA2B) = 22 nM; Ki(hA1) = 1,940 nM; Ki(hA2A) = 3,280; and Ki(hA3) = 1,070 nM] that has favorable pharmacokinetic (PK) properties (t1/2 = 4 h and F > 35% rat). Compound 10 demonstrated functional antagonism at the A2B AdoR (KB = 6 nM) and efficacy in a mouse model of asthma. In two phase 1 clinical trials, CVT-6883 was found to be safe, well tolerated, and suitable for once daily dosing. A second compound 20, 8-(5-pyrazolyl)-xanthine, has been nominated for development from Baraldi’s group in conjunction with King Pharmaceuticals that has favorable A2B AdoR affinity and selectivity [Ki(hA2B) = 5.5 nM; Ki(hA1) > 1,000 nM; Ki(hA2A) > 1,000; and Ki(hA3) > 1,000 nM], and it has been demonstrated to be a functional antagonist. A third compound 32, a 2-aminopyrimidine, from the Almirall group has high A2B AdoR affinity and selectivity [Ki(hA2B) = 17 nM; Ki(hA1) > 1,000 nM; Ki(hA2A) > 2,500; and Ki(hA3) > 1,000 nM], and 32 has been moved into preclinical safety testing. Since three highly selective, high affinity A2B AdoR antagonists have been nominated for development with 10 (CVT-6883) being the furthest along in the development process, the role of the A2B AdoR in various disease states will soon be established

P2Y2 and P2Y6 receptor activation elicits intracellular calcium responses in human adipose-derived mesenchymal stromal cells

Adipose tissue contains self-renewing multipotent cells termed mesenchymal stromal cells. In situ, these cells serve to expand adipose tissue by adipogenesis, but their multipotency has gained interest for use in tissue regeneration. Little is known regarding the repertoire of receptors expressed by adipose-derived mesenchymal stromal cells (AD-MSCs). The purpose of this study was to undertake a comprehensive analysis of purinergic receptor expression. Mesenchymal stromal cells were isolated from human subcutaneous adipose tissue and confirmed by flow cytometry. The expression profile of purinergic receptors was determined by quantitative real-time PCR and immunocytochemistry. The molecular basis for adenine and uracil nucleotide-evoked intracellular calcium responses was determined using Fura-2 measurements. All the known subtypes of P2X and P2Y receptors, excluding P2X2, P2X3 and P2Y12 receptors, were detected at the mRNA and protein level. ATP, ADP and UTP elicited concentration-dependent calcium responses in mesenchymal cells (N = 7–9 donors), with a potency ranking ADP (EC50 1.3 ± 1.0 μM) > ATP (EC50 2.2 ± 1.1 μM) = UTP (3.2 ± 2.8 μM). Cells were unresponsive to UDP (< 30 μM) and UDP-glucose (< 30 μM). ATP responses were attenuated by selective P2Y2 receptor antagonism (AR-C118925XX; IC50 1.1 ± 0.8 μM, 73.0 ± 8.5% max inhibition; N = 7 donors), and UTP responses were abolished. ADP responses were attenuated by the selective P2Y6 receptor antagonist, MRS2587 (IC50 437 ± 133nM, 81.0 ± 8.4% max inhibition; N = 6 donors). These data demonstrate that adenine and uracil nucleotides elicit intracellular calcium responses in human AD-MSCs with a predominant role for P2Y2 and P2Y6 receptor activation. This study furthers understanding about how human adipose-derived mesenchymal stromal cells can respond to external signalling cues

Recent improvements in the development of A2B adenosine receptor agonists

Adenosine is known to exert most of its physiological functions by acting as local modulator at four receptor subtypes named A1, A2A, A2B and A3 (ARs). Principally as a result of the difficulty in identifying potent and selective agonists, the A2B AR is the least extensively characterised of the adenosine receptors family. Despite these limitations, growing understanding of the physiological meaning of this target indicates promising therapeutic perspectives for specific ligands. As A2B AR signalling seems to be associated with pre/postconditioning cardioprotective and anti-inflammatory mechanisms, selective agonists may represent a new therapeutic group for patients suffering from coronary artery disease. Herein we present an overview of the recent advancements in identifying potent and selective A2B AR agonists reported in scientific and patent literature. These compounds can be classified into adenosine-like and nonadenosine ligands. Nucleoside-based agonists are the result of modifying adenosine by substitution at the N6-, C2-positions of the purine heterocycle and/or at the 5′-position of the ribose moiety or combinations of these substitutions. Compounds 1-deoxy-1-{6-[N′-(furan-2-carbonyl)-hydrazino]-9H-purin-9-yl}-N-ethyl-β-D-ribofuranuronamide (19, hA1Ki = 1050 nM, hA2AKi = 1550 nM, hA2B EC50 = 82 nM, hA3Ki > 5 μM) and its 2-chloro analogue 23 (hA1Ki = 3500 nM, hA2AKi = 4950 nM, hA2B EC50 = 210 nM, hA3Ki > 5 μM) were confirmed to be potent and selective full agonists in a cyclic adenosine monophosphate (cAMP) functional assay in Chinese hamster ovary (CHO) cells expressing hA2B AR. Nonribose ligands are represented by conveniently substituted dicarbonitrilepyridines, among which 2-[6-amino-3,5-dicyano-4-[4-(cyclopropylmethoxy)phenyl]pyridin-2-ylsulfanyl]acetamide (BAY-60–6583, hA1, hA2A, hA3 EC50 > 10 μM; hA2B EC50 = 3 nM) is currently under preclinical-phase investigation for treating coronary artery disorders and atherosclerosis

Vaccinomics and Personalized Vaccinology: Is Science Leading Us Toward a New Path of Directed Vaccine Development and Discovery?

As is apparent in many fields of science and medicine, the new biology, and particularly new high-throughput genetic sequencing and transcriptomic and epigenetic technologies, are radically altering our understanding and views of science. In this article, we make the case that while mostly ignored thus far in the vaccine field, these changes will revolutionize vaccinology from development to manufacture to administration. Such advances will address a current major barrier in vaccinology—that of empiric vaccine discovery and development, and the subsequent low yield of viable vaccine candidates, particularly for hyper-variable viruses. While our laboratory's data and thinking (and hence also for this paper) has been directed toward viruses and viral vaccines, generalization to other pathogens and disease entities (i.e., anti-cancer vaccines) may be appropriate

Integration of P2Y receptor-activated signal transduction pathways in G protein-dependent signalling networks

The role of nucleotides in intracellular energy provision and nucleic acid synthesis has been known for a long time. In the past decade, evidence has been presented that, in addition to these functions, nucleotides are also autocrine and paracrine messenger molecules that initiate and regulate a large number of biological processes. The actions of extracellular nucleotides are mediated by ionotropic P2X and metabotropic P2Y receptors, while hydrolysis by ecto-enzymes modulates the initial signal. An increasing number of studies have been performed to obtain information on the signal transduction pathways activated by nucleotide receptors. The development of specific and stable purinergic receptor agonists and antagonists with therapeutical potential largely contributed to the identification of receptors responsible for nucleotide-activated pathways. This article reviews the signal transduction pathways activated by P2Y receptors, the involved second messenger systems, GTPases and protein kinases, as well as recent findings concerning P2Y receptor signalling in C6 glioma cells. Besides vertical signal transduction, lateral cross-talks with pathways activated by other G protein-coupled receptors and growth factor receptors are discussed

Role of biomechanics in the understanding of normal, injured, and healing ligaments and tendons

Ligaments and tendons are soft connective tissues which serve essential roles for biomechanical function of the musculoskeletal system by stabilizing and guiding the motion of diarthrodial joints. Nevertheless, these tissues are frequently injured due to repetition and overuse as well as quick cutting motions that involve acceleration and deceleration. These injuries often upset this balance between mobility and stability of the joint which causes damage to other soft tissues manifested as pain and other morbidity, such as osteoarthritis

P2X7 receptor: Death or life?

The P2X7 plasma membrane receptor is an intriguing molecule that is endowed with the ability to kill cells, as well as to activate many responses and even stimulate proliferation. Here, the authors give an overview on the multiplicity and complexity of P2X7-mediated responses, discussing recent information on this receptor. Particular attention has been paid to early and late signs of apoptosis and necrosis linked to activation of the receptor and to the emerging field of P2X7 function in carcinogenesis