New Strategies for the Synthesis of A 3 Adenosine Receptor Antagonists Pier Giovanni Baraldi, a, * Andrea Bovero, a Francesca Fruttarolo, a Romeo Romagnoli, a Mojgan Aghazadeh Tabrizi, a Delia Preti, a Katia Varani, b Pier Andrea Borea b and Allan R. Moorman c a Dipartimento di Scienze Farmaceutiche, Universita ` di Ferrara, Via Fossato di Mortara, 17/19 44100 Ferrara, Italy b Dipartimento di Medicina Clinica e Sperimentale-Sezione di Farmacologia, Universita ` di Ferrara, Via Fossato di Mortara, 17/19 44100 Ferrara, Italy c King Pharmaceuticals Research and Development, 4000 CentreGreen Way, Suite 300, Cary, NC 27513, USA Received 4 March 2003; accepted 18 July 2003 Abstract—NewA 3 adenosinereceptorantagonistsweresynthesizedandtestedathumanadenosinereceptorsubtypes.Anadvanced syntheticstrategypermittedustoobtainalargeamountofthekeyintermediate 5 that was then submitted to alkylation procedures in order to obtain the derivatives 6–8. These compounds were then functionalised into ureas at the 5-position (compounds 9–11, 18 and 19) to evaluate their affinity and selectivity versus hA 3 adenosine receptor subtype; in particular, compounds 18 and 19 dis- played a value of affinity of 4.9 and 1.3nM, respectively. Starting from 5, the synthetic methodologies employed permitted us to perform a rapid and a convenient divergent synthesis. A further improvement allowed the regioselective preparation of the N 8 - substituted compound 7. This method could be used as an helpful general procedure for the design of novel A 3 adenosine receptor antagonists without the difficulty of separating the N 8 -substituted pyrazolo[4,3-e]1,2,4-triazolo[1,5-c]pyrimidines from the corresponding N 7 -isomers. # 2003 Elsevier Ltd. All rights reserved. Introduction Adenosine, an endogenous modulator of a wide range of biological functions in the nervous, cardiovascular, 1 renal, and immune systems, interacts with at least four cell surface receptor subtypes classified as A 1 ,A 2A ,A 2B and A 3 . These receptor subtypes belong to the super- family of G protein-coupled receptors and have been cloned from several animal species. 2 In particular, the A 3 adenosine receptor subtype, which is distributed in different organs (lung, liver, heart, kidney, and, in low density, in the brain) 3 exerts its action through the modulation of two second messengers systems: stimula- tion of phospholipases C 4 and D 5 and inhibition of adenylate cyclase. 6 The potential therapeutic applica- tions of activating or antagonizing this receptor subtype have been investigated in recent years and in particular, antagonists for A 3 receptor promise to be useful for the treatment of inflammation and in regulation of cell growth. 7,8 Recently, our research group reported a large series of pyrazolo-triazolo-pyrimidines bearing substituted phe- nylcarbamoyl residues at the amino group at the 5- position (Chart 1, compound a) as highly potent and selective antagonists of the human A 3 adenosine recep- tor. 9,10 Among these, Compound b (5-[(phenyl)amino]- carbonyl]amino-8-methyl-2-(2-furyl)-pyrazolo[4,3- e]- 1,2,4-triazolo[1,5-c] pyrimidine (Chart 1) showed highly favorable binding affinity and selectivity for the human A 3 adenosine receptor. 11 Unfortunately, the major problem within this class of compounds is the typical low water solubility that has limited the in vivo pharmacological screening. Starting from these experimental observations, we decided to introduce oxygenated functions like b-hydroxyethyl, acetic, and diethyloxyethyl groups at the 8-position of the pyrazole nitrogen (Fig.1) with the aim of increasing the water solubility of the final compounds obtained 0968-0896/$ - see front matter # 2003 Elsevier Ltd. All rights reserved. doi:10.1016/S0968-0896(03)00484-X Bioorganic & Medicinal Chemistry 11 (2003) 4161–4169 *Corresponding author. Tel.: +39-0532-291293; fax: +39-0532- 291296; e-mail: pgb@dns.unife.it