Synthesis and structure-activity relationships of a new set of 1,2,4-triazolo[4,3-a]quinoxalin-1-one derivatives as adenosine receptor antagonists

In a previous paper (Colotta V. et al., J. Med. Chem. 2000, 43, 1158), we reported the synthesis and the binding activity of some 4-oxo (A) and 4-amino (B) substituted 1,2,4-triazolo[4,3-a]quinoxalin-1-ones, bearing different substituents on the appended 2-phenyl ring (region 1), some of which were potent and selective A(1) or A(3) antagonists. To further investigate the SAR in this class of antagonists, in the present paper some 2-phenyl-1,2,4-triazolo[4,3-a]quinoxalin-1-one derivatives of both series A and B, bearing simple substituents on the benzofused moiety (region 2), are reported. The binding data at bovine A(1) (bA(1)) and A(2A)(bA(2A)) and at human A(3) (hA(3)) adenosine receptors (ARs) show that in series A (compounds 1, 4-11) the presence of substituents on the benzofused moiety is, in general, not advantageous for anchoring at all three AR subtypes, while within series B (compounds 12-21) it exerts a beneficial effect for both bA(1) and hA(3) AR affinities which span the low nanomolar range. In particular, among the 4-amino derivatives 12-21, the 8-chloro-6-nitro (compound 17) and the 6-nitro (compound 18) substitutions afford, respectively, the highest bA(1) and hA(3) AR affinity. Moreover, compound 18, additionally investigated in binding assays at human A(1) (hA(1)) receptors, shows a 183-fold selectivity for hA(3) versus hA(1) receptors. Finally, the SAR studies provide some new insights about the steric and lipophilic requirements of the hA(3) receptor binding pocket which accommodates the benzofused moiety of our 4-amino-triazoloquinoxalin-1-one derivatives.     

Comparison of inhibitory activity of isomeric triazolopyridine derivatives towards adenosine receptor subtypes or do similar structures reveal similar bioactivities?

The synthesis of an array of 8-amino-2-aryl-[1,2,4]triazolo[1,5-a]pyridine-6-carboxyl amide derivatives is described for the first time. A subset of 20 derivatives were compared to their isomeric 5-amino-2-aryl-[1,2,4]triazolo[1,5-a]pyridine-7-carboxyl amide counterparts with regard to their potential to inhibit the human adenosine 2a (hA2a) receptor and their selectivity against the human adenosine 1 (hA1) receptor. Based on the analysis of H-bond donor/acceptor capabilities of the isomeric triazolopyridine pairs it can be concluded that the H-bond donor strength of the free amino functionality is the main determinant for hA2a inhibitory activity and hA1 selectivity.     

Synthesis, ligand-receptor modeling studies and pharmacological evaluation of novel 4-modified-2-aryl-1,2,4-triazolo[4,3-a]quinoxalin-1-one derivatives as potent and selective human A3 adenosine receptor antagonists

The study of some 4-substituted-2-aryl-1,2,4-triazolo[4,3-a]quinoxalin-1-one derivatives, designed as hA(3) adenosine receptor antagonists, is reported. The new compounds bear on the four-position different acylamino, sulfonylamino, benzylureido and benzyloxy moieties, which have also been combined with a para-methoxy group on the 2-phenyl ring or with a nitro residue at the six-position. Many derivatives show high hA(3) adenosine receptor affinities and selectivities both versus hA(1) and hA(2A) receptors. The observed structure-affinity relationships of this class of antagonists have been exhaustively rationalized using the recently published ligand-based homology modeling (LBHM) approach. The selected 4-bismethanesulfonylamino-2-phenyl-1,2,4-triazolo[4,3-a]quinoxalin-1-one (13), which shows high hA(3) affinity (K(i)=5.5nM) and selectivity versus hA(1), hA(2A) (both selectivity ratios>1800) and hA(2B) (cAMP assay, IC(50)>10,000nM) receptors, was tested in an in vitro rat model of cerebral ischemia, proving to be effective in preventing the failure of synaptic activity, induced by oxygen and glucose deprivation in the hippocampus, and in delaying the occurrence of anoxic depolarization.     

Synthesis, adenosine receptor binding and 3D-QSAR of 4-substituted 2-(2'-furyl)-1,2,4-triazolo[1,5-a]quinoxalines

A collection of 25 2-(2'-furyl)-1,2,4-triazolo[1,5-a]quinoxalines incorporating different substitution patterns at position 4 have been synthesized and their binding affinity towards human adenosine receptors (hA(1), hA(2A), hA(2B) and hA(3)) was determined. The biological data show that several potent at hA(1), but lightly selective, adenosine ligands were identified. Moreover, these results confirmed the hypothesis that the structural modifications carried out on the 4-position of the tricyclic system produces a remarkable modification of the adenosine receptorial profile. A 3D-QSAR modelling study (GRIND/ALMOND methodology) performed on the hA(1) data gave further support to the pharmacological results, and it is presented as a useful tool for the future design of ligands with better pharmacological profiles.     

Multinuclear NMR spectroscopy and antitumor activity of novel platinum(II) complexes with 5,7-disubstituted-1,2,4-triazolo[1,5-a]pyrimidines

Novel platinum(II) complexes with 5,7-disubstituted-1,2,4-triazolo[1,5-a]pyrimidines have been synthesized and characterized by infrared and multinuclear magnetic resonance spectroscopic techniques (1H, 13C, 15N, 195Pt). The complexes are of two types: [PtCl2(L)2] and [PtCl2(NH3)(L)], where L=5,7-diphenyl-1,2,4-triazolo[1,5-a]pyrimidine (dptp) and 5,7-ditertbutyl-1,2,4-triazolo[1,5-a]pyrimidine (dbtp). Significant 15N NMR upfield shifts (92-95 ppm) were observed for N(3) atom indicating this nitrogen atom as a coordination site. The molecular structure suggest that Pt(II) ion has the square planar geometry with N(3) bonded 5,7-disubstituted-1,2,4-triazolo[1,5-a]pyrimidines, N-bonded second ligand (NH3 for cis-[PtCl2(NH3)(L)] or, respectively, 5,7-disubstituted-1,2,4-triazolo[1,5-a]pyrimidines for cis-[PtCl2L2]) and two cis chloride anions. The antiproliferative activity in vitro of complexes (1-4) have been tested against the cells of four human cell lines: SW707 rectal adenocarcinoma, A549 non-small cell lung carcinoma, T47D breast cancer and HCV29T bladder cancer. The results indicate a moderate antiproliferative activity of (4) against the cells of rectal, breast and bladder cancer and a marked and selective cytotoxic effect of (1-3) against the cells of all studied human cancer lines.     

Synthesis of 3-(alpha- and beta-D-arabinofuranosyl)-6-chloro-1,2,4-triazolo[4,3-b]pyridazine

Di-O-isopropylidene- and O-methanesulfonyl-protected 1-C-(6-chloro-1,2,4-triazolo[4,3-b]pyridazin-3-yl)pentitols were prepared in three to four steps from D-galactose, D-glucose, D-mannose, and 2,3:5,6-di-O-isopropylidene-alpha-D-mannofuranose. Acid-catalysed treatment of (1S)- and (1R)-1-C-(6-chloro-1,2,4-triazolo[4,3-b]-pyridazin-3-yl)-2,3:4,5-di-O-isopropylidene-1-O-methanesulfonyl-D-arabinitols in refluxing 1,2-dimethoxyethane furnished 3-(alpha- and beta-D-arabinofuranosyl)-6-chloro-1,2,4-triazolo[4,3-b]pyridazine, respectively. Several structures, including the structure of the 3-(beta-D-arabinofuranosyl)-6-chloro-1,2,4-triazolo[4,3-b]pyridazine, were also determined by single-crystal X-ray diffraction analysis.     

Synthesis of 4-amino-6-(hetero)arylalkylamino-1,2,4-triazolo[4,3-a]quinoxalin-1-one derivatives as potent A(2A) adenosine receptor antagonists

In previous papers (Colotta, V. et al. Arch. Pharm. Pharm. Med. Chem. 1999, 332, 39. Colotta, V. et al. J. Med. Chem. 2000, 43, 1158) we reported the synthesis and binding affinity at bovine (b) A₁ and A_2A and human (h) A₃ adenosine receptors (ARs) of the 4-amino-6-benzylamino-2-phenyl-1,2,4-triazolo[4,3-a]quinoxalin-1-one (compound A) which resulted in a potent and selective A_2A AR antagonist. Compound A provided the lead compound of a series of 6- or 8-(hetero)arylalkylamino-4-amino-2-phenyl-1,2,4-triazolo[4,3-a]quinoxalin-1-one derivatives (compounds 1–20) which are the object of this paper. Most of the newly synthesized compounds are inactive at hA₃ ARs while they possess both nanomolar bA_2A affinities and different degrees of bA_2A versus bA₁ selectivity. The binding data show that hydrophilic substituents on the benzyl moiety are the most profitable for bA_2A receptor affinity. Furthermore, their steric hindrance seems to play an important role for the bA_2A AR interaction, thus suggesting that the 6-aralkylamino moiety of these ligands interacts with a size-limited binding pocket of this AR subtype. Thus, the SAR studies provided us some new insights about the structural requirements of the bA_2A AR recognition site.     

1,2,4-triazolo[4,3-a]pyrimidines: a new kind of ligands. Structure of the silver(I) dimer with the 7-oxo derivative

This paper describes the silver dinuclear complex [Ag₂L₂(NO₃)₂] · 2H₂O, where L represents the bridging ligand 7,8-dihydro-7-oxo-1,2,4-triazolo[4,3-a]pyrimidine, this being the first example of a coordination compound of a 1,2,4-triazolo[4,3-a]pyrimidine derivative. As a difference with the most studied 1,2,4-triazolo[1,5-a]pyrimidine derivatives, the coordination takes place through the contiguous nitrogen atoms of the triazole ring, closing a six member Ag₂N₄ core with a higher intermetallic distance, 3.4791(3) Å. Linear coordination of silver is not possible in this geometry, so flat trigonal coordination involving also the nitrate counteranion is found instead.     

3H-[1,2,4]-Triazolo[5,1-i]purin-5-amine derivatives as adenosine A2A antagonists

A novel series of 3-substituted-8-aryl-[1,2,4]-triazolo[5,1-i]purin-5-amine analogs related to Sch 58261 was synthesized in order to identify potent adenosine A(2A) receptor antagonists with improved selectivity over the A(1) receptor, physiochemical properties, and pharmacokinetic profiles as compared to those of Sch 58261. As a result of structural modifications, numerous analogs with excellent in vitro binding affinities and selectivities were identified. Moreover, compound 27 displayed both superior in vitro and highly promising in vivo profiles.     

Synthesis and analgesic/anti-inflammatory evaluation of fused heterocyclic ring systems incorporating phenylsulfonyl moiety

A series of pyrazolo[1,5-a]pyrimidine, triazolo[1,5-a]pyrimidine, and pyrimido[1,2-a]benzimidazole ring systems incorporating phenylsulfonyl moiety were synthesized via the reaction of 3-(N,N-dimethylamino)-1-aryl-2-(phenylsulfonyl)prop-2-en-1-one derivatives 2a,b with appropriate nitrogen nucleophiles. The analgesic and anti-inflammatory activities of the newly synthesized compound were investigated in vivo. 3-Bromo-2-phenyl-6-(phenylsulfonyl)-7-(4-methylphenyl)pyrazolo[1,5-a]pyrimidine (5e) was found to have an excellent analgesic activity in comparison with indomethacin as a reference drug, while the highest anti-inflammatory effect was observed in the case of 2-(4-bromophenyl)-6-(phenylsulfonyl)-5-(4-methylphenyl)pyrazolo[1,5-a]pyrimidine (5d). From the structure-activity relationship (SAR) point of view, the analgesic/anti-inflammatory activity of pyrazolo[1,5-a]pyrimidine derivatives was found to be much higher than triazolo[1,5-a]pyrimidine and pyrimido[1,2-a]benzimidazole derivatives.     

2-cyanoaminopyrimidines as a class of antitumor agents that promote tubulin polymerization

A series of 4-chloro-2-cyanoamino-6-fluoroalkylamino-5-phenylpyrimidines was prepared as a result of our efforts to modify a series of [1,2,4]triazolo[1,5-a]pyrimidines that proved to be potent anticancer agents with a unique mechanism of tubulin inhibition. On the cyanoamino nitrogen, a methyl group is optimal for activity among alkyl groups introduced. The structure-activity relationship for the rest of the molecule resembles that of [1,2,4]triazolo[1,5-a]pyrimidines. A lead compound (5) retained in vitro potency compared with TTI-237. In the mechanism of action studies, it behaved in the same manner as TTI-237. In addition, it is also capable of overcoming multidrug resistance due to P-gp. These findings strongly suggest that this series of 2-cyanoaminopyrimidines binds at the same site and in the same binding mode as TTI-237. Further modifications of the 2-cyanoamino group are underway.     

2-(2-Furanyl)-7-phenyl[1,2,4]triazolo[1,5-c]pyrimidin-5-amine analogs: highly potent, orally active, adenosine A2A antagonists. Part 1

The structure-activity relationship of this novel class of compounds based on 2-(2-furanyl)-7-phenyl[1,2,4]-triazolo[1,5-c]pyrimidin-5-amine, 1, and its analogs was evaluated for their in vitro and in vivo adenosine A(2A) receptor antagonism. Several compounds displayed oral activity at 3 mg/kg in a rat catalepsy model. Specifically, compound 8g displayed an excellent in vitro profile, as well as a highly promising in vivo profile.     

Synthesis of novel D-ring fused 7'-aryl-androstano[17,16-d][1,2,4] triazolo[1,5-a]pyrimidines

The preparation of novel steroidal heterocycles containing the 7-aryl-substituted 1,2,4-triazolo[1,5-a]pyrimidine moiety fused to the 16,17-positions of the steroid nucleus is described. The Aldol reaction of 4-aza-androst-3,17-dione (1a) and dehydroepiandrosterone (DHEA, 1b) with aromatic aldehydes was catalyzed by KF/Al(2)O(3) to give the corresponding 3-oxo-4-aza-5α- and 3β-hydroxy-5-en-16-arylidene-17-ketosteroids (2a-r). Subsequently, the intermediates 2a-r reacted with dinucleophilic 3-amino-1,2,4-triazole in presence of t-BuOK to afford the title compounds (3a-r). All the synthesized heterosteroids are new and are currently being evaluated for their biological activities.     

Synthesis and structure-activity relationships of 1,2,4-triazolo[1,5-a]pyrimidin-7(3H)-ones as novel series of potent β isoform selective phosphatidylinositol 3-kinase inhibitors

A series of 1,2,4-triazolo[1,5-a]pyrimidin-7(3H)-ones with excellent enzyme inhibition, improved isoform selectivity, and excellent inhibition of downstream phosphorylation of AKT has been identified. Several compounds in the series demonstrated potent (～ 0.100 μM IC(50)) growth inhibition in a PTEN deficient cancer cell line.     

Synthesis and biological evaluation of 1-substituted-3-(6-methylpyridin-2-yl)-4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)pyrazoles as transforming growth factor-β type 1 receptor kinase inhibitors

A series of 1-substituted-3-(6-methylpyridin-2-yl)-4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)pyrazoles 14a-ae, 16a, 16b, and 21a-c has been prepared and evaluated for their ALK5 inhibitory activity in an enzyme assay and in a cell-based luciferase reporter assay. The 4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)-N-(4-methoxyphenyl)-3-(6-methylpyridin-2-yl)-1H-pyrazole-1-carbothioamide (14n) inhibited ALK5 phosphorylation with IC(50) value of 0.57 nM and showed 94% inhibition at 100 nM in a luciferase reporter assay using HaCaT cells permanently transfected with p3TP-luc reporter construct.     

Pyrrolo- and pyrazolo-[3,4-e][1,2,4]triazolo[1,5-c]pyrimidines as adenosine receptor antagonists

The discovery and development of adenosine receptor antagonists have represented for years an attractive field of research from the perspective of identifying new drugs for the treatment of widespread disorders such as inflammation, asthma and Parkinson's disease. The present work can be considered as an extension of our structure-activity relationship studies on the pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidine (PTP) nucleus, extensively investigated by us as a useful template, in particular, for the identification of A(2A) and A(3) adenosine receptor antagonists. In order to explore the role of the nitrogen at the 7-position, we performed a new synthetic strategy for the preparation of pyrrolo[3,4-e][1,2,4]triazolo[1,5-c]pyrimidine derivatives which can be considered as 7-deaza analogues of the parent PTPs. We also synthesised a novel series of pyrazolo[3,4-e][1,2,4]triazolo[1,5-c]pyrimidines as junction isomers of the reference compounds. In both cases we obtained some examples of potent antagonists (K(i) in the low nanomolar range) with variable selectivity profiles in relation to the nature of substituents introduced at the C(5)-, N(8)- and/or N(9)-positions. The pyrrolo-triazolo-pyrimidine derivative 9b appeared to be a potent A(3) adenosine receptor antagonist (K(i)=10 nM) with good selectivity over hA(1) (74-fold) and hA(2A) (20-fold) adenosine receptors combined with low activity at the hA(2B) subtype (IC(50)=906 nM). Moreover, some examples of high-affinity A(1)/A(2A) dual antagonists have been identified in both series. This work constitutes a new and important contribution for the comprehension of the interaction between PTPs and adenosine receptors.     

Synthesis and in vitro activity of novel 1,2,4-triazolo[4,3-a]pyrimidine oxazolidinone antibacterial agents. Part II

The synthesis and antibacterial activity of 1,2,4-triazolo[4,3-a]pyrimidine oxazolidinones is reported. Compound 3e with a 2,4-disubstituted thiophene ring was found to be a potent inhibitor of Gram-positive pathogens and was 4-16-fold more potent than Linezolid.     

Synthesis and pharmacological investigation of novel 1-substituted-4-phenyl-1,2,4-triazolo[4,3-a]quinazolin-5(4H)-ones as a new class of H1-antihistaminic agents

A series of novel 1-substituted-4-phenyl-1,2,4-triazolo[4,3-a]quinazolin-5(4H)-ones 7 were synthesized by the cyclization of 2-hydrazino-3-phenylquinazolin-4(3H)-one 6 with various one carbon donors. The starting material 2-hydrazino-3-phenylquinazolin-4(3H)-one 6, was synthesized from aniline 1 by a novel innovative route. When tested for their in vivo H(1)-antihistaminic activity on conscious guinea pigs all the test compounds protected the animals from histamine induced bronchospasm significantly, whereas the compound 1-methyl-4-phenyl-1,2,4-triazolo[4,3-a]quinazolin-5(4H)-one 7b (percentage protection 70.7%) was found to be equipotent with the reference standard chlorpheniramine maleate (percentage protection 71%). These compounds show negligible sedation ( approximately 5%) when compared to the reference standard (26%). Hence they could serve as prototype molecules for future development.     

Synthesis, characterization, crystal structures and in vitro antistaphylococcal activity of organotin(IV) derivatives with 5,7-disubstituted-1,2,4-triazolo[1,5-a]pyrimidine

New organotin(IV) complexes of 5,7-ditertbutyl-1,2,4-triazolo[1,5-a]pyrimidine (dbtp) and 5,7-diphenyl-1,2,4-triazolo[1,5-a]pyrimidine (dptp) with 1:1 and/or 1:2 stoichiometry were synthesized and investigated by X-ray diffraction, FT-IR and ¹¹⁹Sn Mössbauer in the solid state and by ¹H and ¹³C NMR spectroscopy, in solution. Moreover, the crystal and molecular structures of Et₂SnCl₂(dbtp)₂ and Ph₂SnCl₂(EtOH)₂(dptp)₂ are reported. The complexes contain hexacoordinated tin atoms: in Et₂SnCl₂(dbtp)₂ two 5,7-ditertbutyl-1,2,4-triazolo[1,5-a]pyrimidine molecules coordinate classically the tin atom through N(3) atom and the coordination around the tin atom shows a skew trapezoidal structure with axial ethyl groups. In Ph₂SnCl₂(EtOH)₂(dptp)₂ two ethanol molecules coordinate tin through the oxygen atom and the 5,7-diphenyl-1,2,4-triazolo[1,5-a]pyrimidine molecules are not directly bound to the metal center but strictly H-bonded, through N(3), to the OH group of the ethanol moieties; Ph₂SnCl₂(EtOH)₂(dptp)₂ has an all-trans structure and the C-Sn-C fragment is linear. On the basis of Mössbauer data, the 1:2 diorganotin(IV) complexes are advanced to have the same structure of Et₂SnCl₂(dbtp)₂, while Me₂SnCl₂(dptp)₂ to have a regular all-trans octahedral structure. A distorted cis-R₂ trigonal bipyramidal structure is assigned to 1:1 diorganotin(IV) complexes. The in vitro antibacterial activities of the synthesized complexes have been tested against a group of reference pathogen micro-organisms and some of them resulted active with MIC values of 5 μg/mL, most of all against staphylococcal strains, which shows their inhibitory effect.     

Studies on adenosine A2a receptor antagonists: comparison of three core heterocycles

Piperazine and (R)-2-(aminomethyl)pyrrolidine derivatives of [1,2,4]triazolo[1,5-a][1,3,5]triazine have recently been shown to be potent and selective adenosine A(2a) receptor antagonists. We have replaced the triazolotriazine core structure with two different heterocyclic cores. One of these, the one deriving from [1,2,4]triazolo[1,5-c]pyrimidine, appears to be particularly effective and selected analogs from this series have been shown to be orally active in a mouse catalepsy model of Parkinson's disease.