Synthesis and Biological Activity of a New Series of N6-Arylcarbamoyl, 2-(Ar)alkynyl-N6-arylcarbamoyl, and N6-Carboxamido Derivatives of Adenosine-5′-N-ethyluronamide as A1 and A3 Adenosine Receptor Agonists

Pier Giovanni Baraldi; Barbara Cacciari; Maria José Pineda de Las Infantas; Romeo Romagnoli; Giampiero Spalluto; Rosaria Volpini; Stefano Costanzi; Sauro Vittori; Gloria Cristalli; Neli Melman; Kyung-Sun Park; Xiao-duo Ji; Kenneth A Jacobson

doi:10.1021/jm980147p

. Author manuscript; available in PMC: 2022 Aug 10.

Published in final edited form as: J Med Chem. 1998 Aug 13;41(17):3174–3185. doi: 10.1021/jm980147p

Synthesis and Biological Activity of a New Series of N⁶-Arylcarbamoyl, 2-(Ar)alkynyl-N⁶-arylcarbamoyl, and N⁶-Carboxamido Derivatives of Adenosine-5′-N-ethyluronamide as A₁ and A₃ Adenosine Receptor Agonists

Pier Giovanni Baraldi ^1,^*,^†, Barbara Cacciari ^1,^†, Maria José Pineda de Las Infantas ^1,^†, Romeo Romagnoli ^1,^†, Giampiero Spalluto ^1,^†, Rosaria Volpini ^1,^‡, Stefano Costanzi ^1,^‡, Sauro Vittori ^1,^‡, Gloria Cristalli ^1,^‡, Neli Melman ^1,^§, Kyung-Sun Park ^1,^§, Xiao-duo Ji ^1,^§, Kenneth A Jacobson ^1,^§

PMCID: PMC9364910 NIHMSID: NIHMS1828399 PMID: 9703463

Abstract

A new series of 1-(6-amino-9H-purin-9-yl)-1-deoxy-N-ethyl-β-d-ribofuranuronamide-bearing N-arylureas or N-arylcarboxamido groups at the purine 6 position and N-arylureas combined with halogens or alkynyl chains at the 2 position have been synthesized and tested for affinity at A₁ and A_2A adenosine receptors in rat brain membranes and at cloned rat A₃ receptors expressed in CHO cells. The derivatives contained the 5′ substituent found in the potent, nonselective agonist 1-(6-amino-9H-purin-9-yl)-1-deoxy-N-ethyl-β-d-ribofuranuronamide (NECA). While the carboxamido derivatives (9–13) showed affinity for A₁ receptors, the urea derivatives (30–45) showed different degrees of affinity and selectivity for the A₃ adenosine receptor subtype. In particular the derivative bearing a p-sulfonamidophenyl-urea at the 6 position, 31 showed a high affinity (K_i = 9 nM) and selectivity for the A₃ receptors compared to that of the reference compound 1-[6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-β-d-ribofuranuronamide (IB-MECA). Furthermore, the importance of the stereochemistry in the interaction of these ligands at the rat A₃ adenosine receptors has been evaluated by introducing a chiral chain at the 6 position. The introduction of halogens or alkynyl chains at the purine 2 position of selected ureas did not give the expected enhancement of potency at A_2A and/or A₃ receptors but rather showed a dramatic reduction of A_2A affinity, resulting in compounds with good A_2A/A₃ selectivity. For example, the 2-(3-hydroxy-3-phenyl-1-propyn-1-yl)-6-(4-methoxyphenylurea) derivative 61 showed the capability to bind simultaneously to A₁ and A₃ receptor subtypes, excluding the A_2A receptor. Compound 31 was shown to be an agonist, 9-fold more potent than NECA, at A₃ receptors in rat RBL-2H3 mast cell membranes through stimulation of binding of [³⁵S]GTP-γ-S.

Introduction

Four subtypes of adenosine receptors (A₁, A_2A, A_2B, and A₃) have been cloned^1–4 and characterized pharmacologically with the use of potent agonists and antagonists. The A₁ and A₂ receptor subtypes are linked to inhibition and stimulation, respectively, of adenylyl cyclase.^5–7 A₃ receptors are coupled both to the stimulation of phospholipase C, as shown in rat RBL-2H3 mast cells⁸ and in rat brain slices,⁹ and to the inhibition of adenylyl cyclase.⁴

Many selective agents have been developed for the A₁^10–15 and A_2A^16–19 receptor subtypes. Some of these seem promising as potential therapeutic agents in the treatment of Parkinson’s disease,²⁰ cognitive deficits,²¹ schizophrenia,²² epilepsy^23a and renal failure.^23b Selective and/or high affinity agonists and antagonists for the A_2B receptor have not yet been reported. Since the cloning of the A₃ receptor from a rat brain cDNA library,⁴ recent efforts have been made in order to develop selective agonist ligands²⁴ as well as some selective antagonists.^25–28 The selective agonists, 1-[6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-β-d-ribofuranuronamide (IB-MECA) 1, shows a K_i value of 1.1 ± 0.3 nM at rat A₃ receptors and a 50-fold selectivity vs either A₁ or A_2A receptors.²⁹ The related agonist,[¹²⁵I]1-[6-[[(4-amino-3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-β-d-ribofuranuronamide ([¹²⁵I]AB-MECA),³⁰ has become a useful radioligand for the screening of new derivatives interacting with cloned A₃ receptors.

graphic file with name nihms-1828399-f0001.jpg

The A₃ receptor mediates many processes such as inflammation,³¹ hypotension,³² and mast cell degranulation.³³ Also, the A₃ receptor apparently has a role in the central nervous system. IB-MECA 1 induces behavioral depression³⁴ and, upon chronic administration, protects against cerebral ischemia.³⁵ Furthermore, at high concentrations A₃ selective agonists were also found to induce apoptosis in HL-60 human leukemia cells.³⁶

From the point of view of structure–activity relationships, the ribose moiety of adenosine has to be maintained unchanged, with the exception of the 5′ position, at which alkyluronamide groups are well tolerated.³⁷ Furthermore, some substitutions at the purine 6 and/or 2 positions modulates affinity and selectivity of adenosine agonists for the different receptor subtypes.²⁴ Alkyl or aryl derivatives at the 6 position of the purine nucleus are A₁ selective, and many derivatives at the 2 position bearing C, N, or O substituents are A_2A selective. 6-Benzyl analogues also containing the 5′-uronamide modification, such as 1, have been found to be potent and selective A₃ agonists.³⁸ We have recently reported a series of adenosine 5′-uronamide derivatives, substituted at the purine 6 position, of general formula 2, which have shown high affinity at adenosine A₃ receptors. In particular, NECA derivatives containing at the 6-positon the 3-chloro-, 2-chloro-, or 4-methoxyphenyl-carbamoyl moieties have shown affinity for A₃ adenosine receptors with 4–7 nM K_i values. However, while the selectivity with respect to A_2A receptors was high (A_2A/A₃ from 96 to 510), the discrimination between A₁ and A₃ receptors was very low (A₁/A₃ from 4- to 10-fold).³⁹

A new series of carboxamido and urea derivatives at the 6 position of NECA (9–13, 30–45, and 52–62) have been prepared to further explore the structural basis for the selectivity for A₃ vs A₁ receptors. In particular, we were interested in evaluating the influence of the length of the acyl chain at the adenine N position on the receptor affinity. We prepared some amide derivatives of NECA (9–13), which are structurally related to our previously reported series of urea derivatives.³⁹ Also, some new urea derivatives at purine 6 position of NECA (30–45) were synthesized to evaluate the physicochemical parameters involved in the interaction with A₃ adenosine receptors. Furthermore, we introduced a chiral center in compounds 33 and 34 to evaluate the stereoselectivity of binding of the substituent at 6 position.

graphic file with name nihms-1828399-f0002.jpg

On the basis of the observation that 2-(1-hexyn-1-yl)-1-(6-amino-9H-purin-9-yl)-1-deoxy-N-ethyl-β-d-ribofuranuronamide (HENECA), a potent A_2A adenosine agonist,^16,40 also showed high affinity at rat A₃ receptors,⁴¹ we combined ureido moieties at the purine 6 position, that were favorable in A₃ receptor binding, with the substitution at the 2 position. Using results from a previous work,³⁹ we introduced the A₃ receptor affinity enhancing 4-methoxyphenyl-and 3-chlorophenyl-urea groups at the purine 6 position, while substituting the 2 position with halogens (Cl, I) (52–55) and alkynyl groups (56–62).

Results and Discussion

Chemistry.

The preparation of N-acyl derivatives of NECA, compounds 9–13, was performed following the general synthetic strategy depicted in Scheme 1.

The amino group at the 6 position of NECA is not very reactive; therefore, it was necessary to protect the hydroxyl groups of the ribose moiety during selective acylation. Reaction of 2′,3′-O-isopropylidene-protected NECA (3) with the appropriate acyl chloride in the presence of a catalytic amount of triethylamine at reflux afforded the adducts 4–8 in a good yield, which were in turn converted into the desired compounds (9–13) by deprotection in aqueous 1 N HCl and dioxane at 65 °C. In a similar manner, derivatives 30–45 were prepared by nucleophilic reaction of the appropriate isocyanate with 3 and then by deprotection of the ribosyl moiety (Scheme 2).

When not commercially available, the isocyanate was prepared by reacting the corresponding substituted anilines using trichloromethylchloroformate, as described previously.⁴²

Using 4-nitrophenyl isocyanate or 5-chloropyridin-2-yl isocyanate under the conditions mentioned above, the bis adducts 44 and 45 were the only products isolated from the reaction mixture.

The 2-chloro- and the 2-iodo-6-arylcarbamoyl derivatives of NECA 52–55 were synthesized from 2′,3′-O-isopropylidene-2-chloro-NECA (46)⁴³ and 2′,3′-O-isopropylidene-2-iodo-NECA (47),⁴⁴ respectively (Scheme 3).

The reaction of 46 or 47 in dry THF with the appropriate isocyanate to afford the protected nucleosides 48–51 was carried out in a steel bomb at 110 °C for 24 h. The nucleosides were deprotected in 1 N HCl and dioxane at 65 °C to give the corresponding compounds 52–55. The synthesis of the (ar)alkynyl derivatives 56–62 was carried out by a modification of the palladium-catalyzed cross-coupling reaction.⁴⁰

The 2-iodo derivatives 54 and 55 in dry acetonitrile, DMF, and triethylamine reacted with cuprous iodide, bis(triphenylphosphine)palladium dichloride, and the appropriate terminal alkyne, at room temperature for several hours under an atmosphere of N₂, to produce the alkynyl derivatives 56–58 and 59–62, respectively (Scheme 4).

Biological Activity.

All the synthesized compounds (9–13, 30–45, and 52–62) were tested in radioligand binding assays for affinity at the rat brain A₁, A_2A, and A₃ receptors, and the results are summarized in Table 1.⁴⁵

Table 1.

Affinities of Synthesized Derivatives (9–13, 30–45, and 52–62) in Radioligand Binding Assays at Rat Brain A₁, A_2A, and A₃ Adenosine Receptors


compd	R	R¹	R²	K₁ (nM) or % inhibition			A₁/A₃	A_2A/A₃
compd	R	R¹	R²	K_i(A₁)^a	K_i(A_2A)^b	K_i(A₃)^c	A₁/A₃	A_2A/A₃
IB-MECA, 1				54 ± 5	56 ± 8	1.1 ± 0.3	49	51
C1-IB-MECA				820 ± 570	470 ± 365	0.33 ± 0.08	2500	1400
NECA^d				6.3	10.3	113	0.071	0.091
9	H	4-biphenyl	H	5.92 ± 1.04	3580 ± 850	979 ± 22	0.0060	3.7
10	H	2,4-Cl-Ph-CH₂	H	16.0 ± 3.3	26.6 ± 8.8	167 ± 7	0.095	0.16
11	H	4-CH₃O-Ph	H	86.4 ± 14.2	237 ± 46	837 ± 142	0.10	0.28
12	H	2-Cl-Ph	H	1170 ± 220	1960 ± 300	754 ± 132	1.6	2.6
13	H	Ph	H	252 ± 52	670 ± 211	824 ± 126	0.30	0.81
30	H	PhCH₂NH	H	171 ± 15	1793 ± 230	38.3 ± 6.4	4.5	47
31	H	4-SO₂NH₂-PhNH	H	453 ± 141	1180 ± 360	9.73 ± 0.75	47	120
32	H	4-CH₃CO-PhNH	H	72.7 ± 11.8	1050 ± 270	20.9 ± 7.51	3.5	50
33	H	(R)-α-phenylethyl-NH	H	433 ± 171	279 ± 171	16.3 ± 3.7	27	17
34	H	(S)-α-phenylethyl-NH	H	537 ± 33	2970 ± 930	319 ±134	1.7	9.3
35	H	5-Me-isoxazol-3-yl-NH	H	146 ± 39	884 ± 232	532 ± 240	0.27	1.7
36	H	l,3,4-thiadiazol-2-yl-NH	H	208 ± 42	917 ± 254	5550 ± 2800	0.037	0.16
37	H	4-n-C₃H₇O-PhNH	H	247 ± 43	255 ± 85	107 ± 34	2.3	2.4
38	H	Ph-CH₂CH₂NH	H	129 ± 44	2650 ± 640	149 ± 26	0.86	18
39	H	3,4-MeO-Ph-CH₂CH₂NH	H	1770 ± 490	1570 ± 220	411 ± 178	4.3	3.8
40	H	fur-2-yl-CH₂NH	H	419 ± 22	1860 ± 370	713 ± 91	0.58	2.6
41	H	4-(pyridin-2-yl-NHSO₂)PhNH	H	292 ± 76	740 ± 283	54.1 ± 10.8	5.4	14
42	H	4-(5-Me-isoxazol-3-yl-NHSO₂)PhNH	H	901 ± 62	977 ± 425	155 ± 5	5.8	6.3
43	H	4-(pyrimidin-2-yl-NHSO₂)PhNH	H	725 ± 141	16.8 ± 3.1	405 ± 12	1.7	0.041
44	4-NO₂-Ph-NH-CO	4-NO₂-Ph-NH	H	89.1 ± 8.8	2530 ± 200	168 ± 42	0.53	15
45	5-Cl-pyridin-2-yl-NH-CO	5-Cl-pyridin-2-yl-NH	H	761 ± 187	799 ±191	5700 ± 100	0.13	0.14
52	H	3-Cl-Ph-NH	Cl	90.0 ± 23.7	557 ± 112	77.6 ± 25	1.2	4.3
53	H	4-MeO-Ph-NH	Cl	22.0 ± 5.5	59.0 ± 5.3	17.1 ± 8.6	1.2	3.5
54	H	3-Cl-Ph-NH	I	103 ± 23	10 600 ± 3700	315 ± 63	0.33	34
55	H	4-MeO-Ph-NH	I	29.4 ± 12.1	260	251 ± 95	0.12	1.0
56	H	3-Cl-Ph-NH	nC₄H₉–C≡C	2040 ± 540	7130 ± 2570	581 ± 82	3.5	12
57	H	3-Cl-Ph-NH	Ph–C≡C	1330 ± 490	48700 ± 1400	611 ± 100	2.1	80
58	H	3-Cl-Ph-NH	PhCH(OH)–C≡C	814 ± 113	12 ± 3% (10⁻⁴)	696 ±149	1.2	>140
59	H	4-MeO-Ph-NH	nC₄H₉–C≡C	428 ± 124	2330 ± 480	211±104	2.0	11
60	H	4-MeO-Ph-NH	Ph–C≡C	4790 ± 760	29 ± 3% (10⁻⁴)	154 ± 46	31	>4000
61	H	4-MeO-Ph-NH	PhCH(OH)–C≡C	75.1 ± 20.6	12 ± 6% (10⁻⁴)	324 ± 65	0.23	>300
62	H	4-MeO-Ph-NH	Ph(CH₂)₃–C≡C	14400 ± 4000	1670 ± 420	89.5 ± 19	160	19

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Displacement of specific [³H]R–PIA binding (A₁) in rat brain membranes, expressed as K_i ± SEM in nM (n = 3–6).

Displacement of specific [³H]CGS 21680 binding (A_2A) in rat striatal membranes expressed as K_i ± SEM in nM (n = 3–6).

Displacement of specific [¹²⁵I]AB-MECA binding at rat A₃ receptors expressed in CHO cells, expressed as K_i ± SEM in nM (n = 3–6).

Values at A₁ and A_2A receptors are taken from Bruns et al.⁴⁵ K_i values at A₁ receptors are vs specific binding of [³H]-N⁶-cyclohexyladenosine ([³H]-CHA). K_i values at A_2A receptors are vs specific binding of [³H]-NECA in the presence of 50 nM CPA in rat striatal membranes.

In our previous studies, we demonstrated that the 6-substituted adenosine uronamides are full agonists in inhibiting adenylate cyclase via rat A₃ receptors.³⁹

From the binding assays, it appeared that the presence of an amide vs urea functionality at the 6 position (9–13) was generally detrimental in terms of affinity at rat A₃ receptors, whereas several of these derivatives (11 and 13) showed roughly 1 order of magnitude increase in affinity for adenosine A_2A receptor. Affinity at A₁ receptors was highly variable, ranging from 5.9 nM (biphenyl derivative, 9) to 1.2 μM. For example, the o-chlorobenzoyl derivative (12) was 100-fold less potent at A₃ receptors and was 10-fold less potent at A₁ receptors than the corresponding urea derivative.³⁹ The p-methoxybenzoyl derivative (11) was 127-fold less potent at A₃ receptors than the corresponding p-methoxyphenyl urea derivative.³⁹ These results suggested that a small modification in the chain at the 6 position could modulate the affinity for A₁ or A₃ receptors. In the series of urea-derivatives (30–45), a substituted-phenyl (31, 32, and 37) or substituted-benzyl (30, 33, and 34) group led to higher affinity and selectivity at A₃ adenosine receptors. In fact, derivatives 31, 32, and 37 showed a high affinity (9.7–107 nM) at A₃ receptors with varying degrees of A₁/A₃ selectivity. In particular, compound 31 was less active than IB-MECA (9.7 nM vs 1.1 nM) but showed selectivity for A₃ vs either A₁ or A_2A receptors comparable to the reference compound. In addition, it seemed necessary to have a free amino group on the sulfonamido moiety, since its substitution with some heteroaryl groups (e.g., compounds 41–43) led to derivatives with modest affinity at rat A₃ adenosine receptors and low selectivity with respect to A₁ and A_2A receptors.

The data demonstrate the importance of a phenylurea moiety at the purine 6 position for an interaction with the A₃ adenosine receptor subtype.⁴⁶ When the phenyl ring was replaced with various heterocycles (35, 36, and 40) both affinity and selectivity for A₃ receptors decreased.

Moreover, the disubstitution at the 6 position led to derivatives (44–45) with diminished activity, in particular at A_2A and A₃ receptors. This could be explained by the absence of one hydrogen atom at N⁶, which could act as a hydrogen bond donor, already noted as a requirement for binding at the A₁ and A_2A receptor subtypes.⁴⁷

The benzylic series (30, 33, and 34) showed a SAR pattern comparable to the phenylic series. In fact, the unsubstituted compound 30 showed affinity and selectivity at A₃ receptors very similar to those of the previously reported phenyl derivative.³⁹ Interestingly, comparing the two diastereomers 33 and 34 in which substituents at the purine 6 position are of opposite configuration, the R-isomer (33) was more potent than the S-isomer (34) at the rat A_2A and A₃ adenosine receptor subtypes. The binding stereoselectivity of the N⁶ substituent confirms the importance of the stereochemistry at this subregion of the adenosine receptor binding site, similar to that already observed for the A₁ agonists R-PIA and S-PIA.⁴⁸ A similar observation has been made for the C² subregion of the A_2A subtype, both with geometric isomers of 2-alkenyl-NECA⁴⁹ and with diastereomers of 2-alkynyl derivatives of NECA [R- and S-PHPNECA].⁵⁰

The data also demonstrate the importance of chain length at the 6 position and suggest that phenyl or benzyl chains are the preferred substituents for the interaction with the A₃ adenosine receptor subtype. Longer chains, with respect to the phenylic or benzylic moieties, seemed not to improve the affinity at rat A₃ receptors. Thus, compounds, 38 and 39, bearing 2-phenylethyl and 3,4-dimethoxy-2-phenylethyl groups at the purine 6 position, respectively, showed submicromolar affinity but low selectivity at rat A₃ receptors.

These results and our previous work³⁹ seem to indicate that the lipophilicity of the para substituents of a phenyl ring plays a significant role in A₃ affinity. A quantitative correlation was not found, but compounds 31 (π SO₂NH₂ = −1.82), 32 (π COCH₃ = −0.55), and the previously reported 6-phenyl urea derivative substituted with a methoxy group at the para position (π OCH₃ = −0.02), bearing the least lipophilic substituents, show some of the most favorable K_i values within the series.⁵¹

Species differences, which are pronounced for antagonists but not for agonists, in binding of ligands to A₃ receptors have been noted. The most interesting derivative in the present study (31) showed affinity at human A₃ adenosine receptors (56.1 ± 9.1 nM) comparable to that at rat A₃ receptors, but there was 6-fold more potency at rat receptors. This finding confirms, as already observed, significant differences between rat and human A₃ adenosine receptor subtypes.²⁸

A functional assay indicated that compound 31 acted as a full agonist at rat A₃ receptors. The assay consisted of agonist-induced stimulation of binding of a guanine nucleotide to membranes of rat RBL-2H3 mast cells, which contain a high density of A₃ receptors.³⁰ Figure 1 shows that the potent A₃ agonists 1-[6-[[(4-amino-3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-β-d-ribofuranuronamide (I-AB-MECA) and 2-chloro-1-[6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-β-d-ribofuranuronamide (Cl-IB-MECA) increased binding of [³⁵S]GTP-γ-S in a dose-dependent manner and with 9-fold greater potency than NECA, as expected for A₃ receptors.

Figure 1. — Effect of the adenosine agonists NECA, compound 31, Cl-IB-MECA, and I-AB-MECA on the binding of [³⁵S]GTP-γ-S to membranes of rat RBL-2H3 mast cells. EC₅₀ values were NECA (10.6 μM), compound 31 (1.15 μM), Cl-IB-MECA (174 nM), and I-AB-MECA (86.8 nM).

A correlation between binding affinity and potency in this functional assay has been demonstrated.⁵²

Compound 31 was as efficacious as these two potent agonists but approximately 1 order of magnitude less potent, with an EC₅₀ value of 1.15 μM.

The introduction of halogens at the 2 position of selected urea derivatives at the purine 6 position led to compounds 52–55, which show an A₃ affinity ranging from 17 nM (53) to 315 nM (54). For example, the presence of a chlorine atom in the 2 position reduced the affinity and selectivity with respect to the corresponding 2-unsubstituted urea derivatives³⁹ (52, K_i at A₃ = 77.6 nM vs 4.4 nM; 53, K_i at A₃ = 17 nM vs 6.6 nM), unlike data for various other 6-substituted A₁⁵³ and A₃²⁹ receptor agonists. Similarly, the presence of an iodine atom further decreased affinity and selectivity (54, K_i at A₃ = 315 nM; 55, K_i at A₃ = 251 nM).

The introduction of an alkynyl chains at the 2 position of these 5′-carboxamide adenosine derivatives was expected to enhance the potency at A₃ receptors, consistent with the high affinity demonstrated by HENECA (K_i at A₃ = 25.6 nM).⁴¹ Surprisingly, the 2-alkynyl-substituted N⁶-urea 5′-carboxamide derivatives (56–62), in fact, showed a dramatic reduction of A_2A affinity, indicating the interdependence of substitution at the purine 2 and 6 positions. The triple substitution resulted in compounds with extremely high A₃ vs A_2A receptor selectivity, 60 > 61 > 58 > 57 (in order of selectivity). Thus, the most potent compounds in this group, showing high selectivity versus the A_2A subtype and which also showed a comparable affinity for A₁ and A₃ receptors, the 2-(3-hydroxy-3-phenyl-1-propyn-1-yl) derivative 61, could be an interesting candidate for the simultaneous stimulation of both A₁ and A₃ subtypes. This could be interesting since an interaction between these two subtypes has been demonstrated in the rat hippocampus, in the presynaptic control of release of glutamic acid.⁵⁴ Since both the A₁ and A₃ receptors are coupled to the inhibition of adenylate cyclase, it may prove useful to have such pharmacological probes of mixed A₁ and A₃ selectivity such that they would exert a dual, simultaneous action on the same second messenger system. Conversely, compound 62 showed very high selectivity for the A₃ versus the A₁ subtype.

Conclusions

The present study provides useful information concerning the structural requirements necessary for recognition by the A₃ adenosine receptor. It appears that a phenyl- or benzylcarbamoyl moiety at the purine 6 position seems favorable for A₃ adenosine receptor affinity and selectivity, while an arylcarboxamido group enhances A₁ affinity and selectivity. In particular, in the urea series, compound 31 was shown to be less potent but with approximately the same degree of A₃ selectivity vs that of either A₁ or A_2A with respect to the reference compound IB-MECA.

Furthermore, a simultaneous substitution at the 6 and 2 positions does not improve affinity and selectivity for A₃. However, the 2-(3-hydroxy-3-phenyl-1-propyn-1-yl) derivative 61 showed the capability to bind simultaneously to A₁ and A₃ receptor subtypes, excluding the A_2A receptor.

Experimental Section

Chemistry.

Reactions were routinely monitored by thin-layer chromatography (TLC) on silica gel (precoated F₂₅₄ Merck plates), and products were visualized with iodine or aqueous potassium permanganate. Infrared spectra (IR) were measured on a Perkin-Elmer 257 instrument. ¹H NMR spectra were determined in CDCl₃ or DMSO-d₆ solutions with a Bruker AC 200 spectrometer, peak positions are given in parts per million (δ) downfield from tetramethylsilane as internal standard, and J values are given in Hz. Light petroleum ether refers to the fractions boiling at 40–60 °C. Melting points were determined on a Buchi-Tottoli instrument and are uncorrected. Chromatography was performed with Merck 60–200 mesh silica gel. All products reported showed IR and ¹H NMR spectra in agreement with the assigned structures. Organic solutions were dried over anhydrous magnesium sulfate. Elemental analyses were performed by the microanalytical laboratory of Dipartimento di Chimica, University of Ferrara, and were within ±0.4% of the theoretical values for C, H, and N.

General Procedure for the Preparation of 1-Deoxy-1-[6-[[(substituted)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (4–8).

2′,3′-O-Isopropylidene-NECA 3 (0.43 mmol) was dissolved in freshly distilled dioxane (4 mL), and the appropriate acyl chloride (1.3 equiv) and a catalytic amount of triethylamine (2–3 drops) were added. The mixture was refluxed under argon for 15 h. The solvent was then removed under reduced pressure, and the residue was purified by flash chromatography (CH₂Cl₂–EtOAc 20%) to afford the desired compound 4–8.

1-Deoxy-1-[6-[[(4-biphenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (4):

yield 80%; pale yellow foam; IR (neat) cm⁻¹ 3445, 1720, 1640, 1575; ¹H NMR (CDCl₃) δ 0.84 (t, 3H, J = 7), 1.39 (s, 3H), 1.62 (s, 3H), 3.02–3.11 (m, 2H), 4.71 (d, 1H, J = 2), 5.39–5.46 (m, 2H), 6.19 (d, 1H, J = 2), 6.66 (t, 1H, J = 2), 7.40–7.74 (m, 7H), 8.09–8.16 (m, 3H), 8.74 (s, 1H), 9.44 (bs, 1H). Anal. (C₂₈H₂₈N₆O₅) C, H, N.

1-Deoxy-1-[6-[[(2,4-dichlorobenzyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (5):

yield 67%; white foam; IR (neat) cm⁻¹ 3440, 1730, 1625, 1565, 1210; ¹H NMR (CDCl₃) δ 0.82 (t, 3H, J = 7), 1.17 (s, 3H), 1.26 (s, 3H), 2.99–3.13 (m, 2H), 4.32 (s, 2H), 4.72 (d, 1H, J = 1.8), 5.38–5.48 (m, 2H), 6.16 (d, 1H, J = 1.8), 6.56 (t, 1H, J = 2), 7.20–7.32 (m, 3H), 7.41 (s, 1H), 8.20 (s, 1H), 8.67 (s, 1H). Anal. (C₂₃H₂₄Cl₂N₇O₅) C, H, N.

1-Deoxy-1-[6-[[(4-methoxyphenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (6):

yield 85%; white solid, mp 105–107 °C (CH₂Cl₂–Et₂O); IR (KBr) cm⁻¹ 3445, 1715, 1640, 1555, 1210; ¹H NMR (CDCl₃) δ 0.85 (t, 3H, J = 7), 1.40 (s, 3H), 1.63 (s, 3H), 3.03–3.12 (m, 2H), 3.90 (s, 3H), 4.72 (d, 1H, J = 2), 5.41–5.49 (m, 2H), 6.16 (d, 1H, J = 2), 6.61 (t, 1H, J = 2), 7.01 (d, 2H, J = 9), 8.02 (s, 2H, J = 9), 8.10 (s, 1H), 8.74 (s, 1H), 9.09 (bs, 1H). Anal. (C₂₃H₂₆N₆O₆) C, H, N.

1-Deoxy-1-[6-[[(2-chlorophenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (7):

yield 72%; white foam; IR (neat) cm⁻¹ 3435, 1720, 1620, 1550, 1230; ¹H NMR (CDCl₃) δ 0.83 (t, 3H, J = 7), 1.39 (s, 3H), 1.62 (s, 3H), 3.00–3.07 (m, 2H), 4.71 (d, 1H, J = 2), 5.41–5.44 (m, 2H), 6.18 (d, 1H, J = 2), 6.67 (t, 1H, J = 2), 7.30–7.44 (m, 3H), 7.71–7.76 (m, 1H), 8.16 (s, 1H), 8.69 (s, 1H), 10.24 (bs, 1H). Anal. (C₂₂H₂₃ Cl N₇O₅) C, H, N.

1-Deoxy-1-[6-[[(phenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (8):

yield 90%; pale yellow oil; IR (neat) cm⁻¹ 3425, 1730, 1640, 1555, 1240; ¹H N:MR (CDCl₃) δ 0.79 (t, 3H, J = 7), 1.39 (s, 3H), 1.61 (s, 3H), 2.91–3.03 (m, 2H), 4.71 (d, 1H, J = 2), 5.45–5.52 (m, 2H), 6.18 (d, 1H, J = 2), 6.40 (t, 1H, J = 2), 7.27–7.53 (m, 3H), 7.83–7.87 (m, 2H), 8.19 (s, 1H), 8.63 (s, 1H), 9.18 (bs, 1H). Anal. (C₂₂H₂₄N₆O₅) C, H, N.

General Procedure for the Preparation of 1-Deoxy-1-[6-[[[(substituted)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (14–29).

2′,3′-O-Isopropylidene-NECA 3 (0.43 mmol) was dissolved in freshly distilled THF (4 mL), and then the appropriate isocyanate (1.3 equiv) and a catalytic amount of triethylamine (two drops) were added. The mixture was refluxed under argon for 18 h. Next the solvent was removed under reduced pressure, and the residue was purified by flash chromatography (CH₂Cl₂–EtOAc 20%) to afford the desired compound 14–29.

1-Deoxy-1-[6-[[[(benzyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (14):

yield 82%; white solid, mp 109–111 °C (CH₂Cl₂–Et₂O); IR (KBr) cm⁻¹ 3440, 1740, 1610, 1550, 1210; ¹H NMR (CDCl₃) δ 0.76 (t, 3H, J = 7), 1.21 (s, 3H), 1.63 (s, 3H), 2.96–3.03 (m, 2H), 4.64 (d, 2H, J = 6), 4.71 (d, 1H, J = 1.8), 5.44–5.48 (m, 2H), 6.16 (d, 1H, J = 2), 6.52 (t, 1H, J = 2), 7.26–7.38 (m, 5H), 8.14 (s, 1H), 8.45 (s, 1H), 8.46 (s, 1H), 9.79 (bs, 1H). Anal. (C₂₆H₃₁N₇O₅) C, H, N.

1-Deoxy-1-[6-[[[(4-sulfonamidophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (15):

yield 60%; white solid, mp 207 °C (CH₂Cl₂–Et₂O); IR (KBr) cm⁻¹ 3450–3250, 1730, 1610, 1565, 1360, 1230; ¹H NMR (CDCl₃) δ 0.57 (t, 3H, J = 7), 1.35 (s, 3H), 1.54 (s, 3H), 2.96–3.03 (m, 2H), 4.60 (d, 1H, J = 2), 5.47–5.50 (m, 2H), 6.47 (d, 1H, J = 2), 6.60 (t, 1H, J = 2), 7.26 (d, 2H, J = 9), 7.74 (d, 2H, J = 9), 8.60 (s, 1H), 8.64 (s, 1H), 9.22 (s, 1H), 10.47 (bs, 1H), 12.05 (s, 1H). Anal. (C₂₂H₂₆N₈O₇S) C, H, N.

1-Deoxy-1-[6-[[[(4-acetylphenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (16):

yield 77%; white solid, mp 204 °C (CH₂Cl₂–Et₂O); IR (KBr) cm⁻¹ 3430, 1740, 1730, 1630, 1545, 1240; ¹H NMR (CDCl₃) δ 0.83 (t, 3H, J = 7), 1.42 (s, 3H), 1.64 (s, 3H), 2.61 (s, 3H), 2.99–3.08 (m, 2H), 4.74 (d, 1H, J = 2), 5.47–5.55 (m, 2H), 6.21 (d, 1H, J = 2), 6.47 (t, 1H, J = 2), 7.74 (d, 2H, J = 9), 7.98 (d, 2H, J = 9), 8.21 (s, 1H), 8.63 (s, 1H), 8.67 (bs, 1H), 12.01 (s, 1H). Anal. (C₂₁H₂₇N₇O₆) C, H, N.

1-Deoxy-1-[6-[[[((R)-1-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (17):

yield 65%; white solid, mp 110–111 °C (CH₂Cl₂–Et₂O); IR (KBr) cm⁻¹ 3430, 1730, 1620, 1555, 1230; ¹H NMR (CDCl₃) δ 0.68 (t, 3H, J = 7), 1.41 (s, 3H), 1.63 (s, 3H), 1.64 (d, 3H, J = 7), 2.87–2.96 (m, 2H), 4.71 (d, 1H, J = 1.8), 5.16 (m, 1H), 5.48–5.51 (m, 2H), 6.16 (d, 1H, J = 2), 6.52 (bs, 1H), 7.25–7.40 (m, 5H), 8.15 (s, 1H), 8.51 (s, 1H), 9.84 (bs, 1H), 10.94 (s, 1H). Anal. (C₂₃H₂₉N₇O₅) C, H, N.

1-Deoxy-1-[6-[[[((S)-1-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (18):

yield 82%; white solid, mp 107–108 °C (CH₂Cl₂–Et₂O); IR (KBr) cm⁻¹ 3445, 1725, 1615, 1560, 1230; ¹H NMR (CDCl₃) δ 0.85 (t, 3H, J = 7), 1.40 (s, 3H), 1.63 (s, 3H), 1.64 (d, 3H, J = 7), 3.00–3.09 (m, 2H), 4.72 (d, 1H, J = 1.8), 5.17 (m, 1H), 5.39–5.50 (m, 2H), 6.14 (d, 1H, J = 2), 6.57 (bs, 1H), 7.26–7.45 (m, 5H), 8.12 (s, 1H), 8.51 (s, 1H), 9.81 (bs, 1H), 10.87 (s, 1H). Anal. (C₂₃H₂₉N₇O₅) C, H, N.

1-Deoxy-1-[6-[[[(5-methyl-isoxazol-3-yl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (19):

yield 83%; pale yellow solid, mp 120 °C (CH₂Cl₂–Et₂O); IR (KBr) cm⁻¹ 3450, 1720, 1620, 1550, 1215; ¹H NMR (CDCl₃) δ 0.82 (t, 3H, J = 7), 1.20 (s 3H), 1.41 (s, 3H), 2.44 (s, 3H), 3.00–3.07 (m, 2H), 4.74 (d, 1H, J = 2), 5.46–5.50 (m, 2H), 6.21 (d, 1H, J = 2), 6.55 (t, 1H, J = 2), 6.70 (s, 1H), 8.37 (s, 1H), 8.63 (s, 1H), 9.39 (bs, 1H), 12.34 (bs, 1H). Anal. (C₂₀H₂₄N₈O₆) C, H, N.

1-Deoxy-1-[6-[[[(1,3,4-thiadiazol-2-yl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (20):

yield 70%; yellow solid, mp 148 °C (CH₂Cl₂–Et₂O); IR (KBr) cm⁻¹ 3430, 1730, 1615, 1560, 1240; ¹H NMR (CDCl₃) δ 0.86 (t, 3H, J = 7), 1.44 (s 3H), 1.67 (s, 3H), 3.07–3.14 (m, 2H), 4.78 (d, 1H, J = 1.8), 5.47–5.51 (m, 2H), 6.24 (d, 1H, J = 1.8), 6.52 (t, 1H, J = 2), 8.36 (s, 1H), 8.75 (s, 1H), 8.91 (bs, 1H), 9.40 (bs, 1H), 11.72 (bs, 1H). Anal. (C₁₈H₂₁N₉O₅S) C, H, N.

1-Deoxy-1-[6-[[[(4-n-propyloxy-phenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (21):

yield 70%; pale yellow foam; IR (neat) cm⁻¹ 3440, 1725, 1620, 1555, 1220; ¹H NMR (CDCl₃) δ 0.71 (t, 3H, J = 7), 0.90 (t, 3H, J = 7), 1.40 (s 3H), 1.62 (s, 3H), 1.81–2.03 (m, 2H), 2.95–3.01 (m, 2H), 3.90 (t, 2H, J = 7), 4.71 (d, 1H, J = 1.8), 5.46–5.51 (m, 2H), 6.19 (d, 1H, J = 1.8), 6.51 (t, 1H, J = 2), 6.89 (d, 2H, J = 9), 7.48 (d, 2H, J = 9), 8.25 (s, 1H), 8.56 (s, 1H), 8.80 (bs, 1H), 11.48 (s, 1H). Anal. (C₂₂H₂₅N₇O₅) C, H, N.

1-Deoxy-1-[6-[[[(2-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (22):

yield 60%; pale yellow solid, mp 88–90 °C (EtOAc–light petroleum); IR (KBr) cm⁻¹ 3440, 1730, 1615, 1550, 1225; ¹H NMR (CDCl₃) δ 0.70 (t, 3H, J = 7), 1.40 (s 3H), 1.62 (s, 3H), 2.94–3.23 (m, 4H), 3.67 (t, 2H, J = 6.8), 4.71 (d, 1H, J = 1.8), 5.46–5.51 (m, 2H), 6.18 (d, 1H, J = 1.8), 6.49 (t, 1H, J = 6), 7.24–7.32 (m, 5H), 8.25 (s, 1H), 8.30 (s, 1H), 9.48 (bs, 1H), 11.48 (s, 1H). Anal. (C₂₄H₂₉N₇O₅) C, H, N.

1-Deoxy-1-[6-[[[(3,4-dimethoxy-2-phenylethyl)amino]-carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (23):

yield 55%; yellow solid, 85–87 °C mp (EtOAc–light petroleum); IR (KBr) cm⁻¹ 3440, 1724, 1623, 1554, 1225; ¹H NMR (CDCl₃) δ 0.73 (t, 3H, J = 7), 1.23 (t, 2H, J = 8), 1.41 (s 3H), 1.62 (s, 3H), 2.95–3.07 (m, 2H), 3.85 (s, 3H), 3.87 (s, 3H), 4.71 (d, 1H, J = 1.8), 5.47–5.51 (m, 2H), 6.17 (d, 1H, J = 1.8), 6.51 (t, 1H, J = 2), 6.60 (s, 1H), 6.62 (s, 2H), 8.23 (s, 1H), 8.36 (s, 1H), 8.71 (s, 1H), 9.46 (bs, 1H). Anal. (C₂₆H₃₃N₇O₇) C, H, N.

1-Deoxy-1-[6-[[[(fur-2-yl-methyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (24):

yield 58%; yellow solid mp 90–93 °C (EtOAc–light petroleum); IR (KBr) cm⁻¹ 3445, 1727, 1620, 1550, 1222; ¹H NMR (CDCl₃) δ 0.72 (t, 3H, J = 7), 1.40 (s, 3H), 1.62 (s, 3H), 2.84–2.99 (m, 2H), 4.6 (d, 2H, J = 6), 4.71 (d, 1H, J = 1.8), 5.45–5.55 (m, 2H), 6.19 (d, 1H, J = 1.8), 6.30–6.36 (m, 2H), 6.60 (s, 1H), 6.5 (t, 1H, J = 6), 7.38 (d, 1H, J = 2), 8.32 (s, 1H), 8.49 (s, 1H), 9.00 (s, 1H), 9.84 (bs, 1H). Anal. (C₂₁H₂₅N₇O₆) C, H, N.

1-Deoxy-1-[6-[[[(4-(pyridin-2-yl-aminosulfonyl)phenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (25):

yield 48%; yellow solid mp 168–170 °C (EtOAc–light petroleum); IR (KBr) cm⁻¹ 3450, 1730, 1625, 1550, 1350, 1220; ¹H NMR (CDCl₃) δ 0.71 (t, 3H, J = 7), 1.41 (s, 3H), 1.60 (s, 3H), 2.80–2.95 (m, 2H), 4.73 (d, 1H, J = 1.8), 5.40–5.60 (m, 2H), 6.19 (d, 1H, J = 1.8), 6.65 (t, 1H, J = 6), 7.15–7.25 (m, 1H), 7.45 (d, 2H, J = 9), 7.55–7.75 (m, 3H), 7.92 (d, 2H, J = 9), 8.27 (s, 1H), 8.50 (s, 1H), 8.99 (s, 1H), 9.80 (bs, 1H), 11.02 (bs, 1H). Anal. (C₂₇H₂₉N₉O₇S) C, H, N.

1-Deoxy-1-[6-[[[(4-(5-methyl-isoxazol-3-yl-aminosulfonyl)phenyl)amino] carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuron-amide (26):

yield 56%; yellow solid mp 157–159 °C (EtOAc–light petroleum); IR (KBr) cm⁻¹ 3455, 1725, 1630, 1557, 1355, 1224; ¹H NMR (DMSO-d₆) δ 0.79 (t, 3H, J = 7), 1.40 (s, 3H), 1.62 (s, 3H), 2.30 (s, 3H), 2.91–3.04 (m, 2H), 4.68 (d, 1H, J = 1.8), 5.40–5.48 (m, 2H), 6.11 (d, 1H, J = 1.8), 6.39 (s, 1H), 7.06 (t, 1H, J = 6), 7.63 (d, 2H, J = 9), 7.85 (d, 2H, J = 9), 8.44 (s, 1H), 8.59 (s, 1H), 9.07 (s, 1H), 9.62 (bs, 1H), 12.11 (bs, 1H). Anal. (C₂₆H₂₉N₉O₈S) C, H, N.

1-Deoxy-1-[6-[[[(4-(pyrimidin-2-yl-aminosulfonyl)-phenyl)amino] carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (27):

yield 53%; yellow solid mp 157–159 °C (EtOAc–light petroleum); IR (KBr) cm⁻¹ 3450, 1722, 1633, 1552, 1350, 1220; ¹H NMR (DMSO-d₆) δ 0.57 (t, 3H, J = 7), 1.35 (s, 3H), 1.54 (s, 3H), 2.71–2.80 (m, 2H), 4.59 (d, 1H, J = 1.8), 5.40–5.46 (m, 2H), 6.46 (d, 1H, J = 1.8), 7.03 (t, 1H, J = 6), 7.56 (m, 1H), 7.60 (d, 2H, J = 9), 7.85 (d, 2H, J = 9), 8.48 (s, 1H), 8.51 (s, 1H), 8.60 (d, 2H, J = 7), 10.45 (bs, 1H), 11.71 (bs, 1H), 12.04 (s, 1H). Anal. (C₂₆H₂₈N₁₀O₇S) C, H, N.

1-Deoxy-1-[6,6-[[[bis-(4-nitrophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (28):

yield 65%; yellow solid, mp 261 °C (CH₂Cl₂–Et₂O); IR (KBr) cm⁻¹ 3430, 1730, 1620, 1555, 1350, 1210; ¹H NMR (DMSO-d₆) δ 0.56 (t, 3H, J = 7), 1.34 (s, 3H), 1.54 (s, 3H), 2.67–2.83 (m, 2H), 4.61 (d, 1H, J = 1.8), 5.47–5.50 (m, 2H), 6.47 (d, 1H, J = 1.8), 7.61 (t, 1H, J = 2), 7.71 (d, 2H, J = 9), 7.91 (d, 2H, J = 9), 8.22 (d, 2H, J = 9), 8.24 (s, 1H), 8.64 (d, 2H, J = 9), 9.72 (s, 1H), 10.63 (bs, 1H), 12.24 (s, 1H). Anal. (C₂₉H₂₈N₁₀O₁₀) C, H, N.

1-Deoxy-1-[6,6-[[[bis-(5-chloro-pyridin-2-yl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (29):

yield 60%; yellow foam; IR (neat) cm^–1 3440, 1710, 1630, 1540, 1220; ¹H NMR (DMSO-d₆) δ 0.58 (t, 3H, J = 7), 1.35 (s 3H), 1.54 (s, 3H), 2.71–2.85 (m, 2H), 4.60 (d, 1H, J = 2), 5.47–5.51 (m, 2H), 6.47 (d, 1H, J = 2), 7.52 (t, 1H, J = 2), 7.79–7.92 (m, 4H), 8.34–8.38 (m, 2H), 8.60 (s, 1H), 8.65 (s, 1H), 10.66 (bs, 1H), 12.22 (bs, 1H). Anal. (C₂₇H₂₆Cl₂N₈O₆) C, H, N.

General Procedure for the Preparation of 1-Deoxy-1-[6-[[(substituted)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (9–13) and 1-Deoxy-1-[6-[[[(substituted)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (30–45).

A solution of the isopropylidene derivative, 4–8 or 14–29, (0.084 mmol) in aqueous 1 N HCl (5 mL) and dioxane (5 mL) was stirred at 65 °C for 1 h. The solvent was then removed under reduced pressure, and the residue was crystallized from ethanol to afford the desired compound, 9–13 or 30–45.

1-Deoxy-1-[6-[[(4-biphenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (9):

yield 65%; white solid, mp 221 °C (EtOH); IR (KBr) cm⁻¹ 3500–3100, 1720, 1615, 1550; ¹H NMR (DMSO-d₆) δ 1.07 (t, 3H, J = 7), 3.17–3.24 (m, 2H), 4.23–4.25 (m, 1H), 4.36 (d, 1H, J = 2), 4.71–4.73 (m, 1H), 5.70 (d, 1H, J = 8), 5.79 (d, 1H, J = 4), 6.13 (d, 1H, J = 8), 7.43–7.53 (m, 3H), 7.78–7.89 (m, 4H), 8.16 (d, 2H, J = 11), 8.46 (t, 1H, J = 4), 8.80 (s, 1H), 8.82 (s, 1H), 11.34 (bs, 1H). Anal. (C₂₅H₂₄N₆O₅) C, H, N.

1-Deoxy-1-[6-[[(2,4-dichlorobenzyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (10):

yield 55%; off white solid, mp 132–134 °C (EtOH); IR (KBr) cm⁻¹ 3450–3050, 1730, 1635, 1545, 1230; ¹H NMR (DMSO-d₆) δ 1.08 (t, 3H, J = 7), 3.18–3.25 (m, 2H), 3.34 (s, 2H), 4.12–4.15 (m, 1H), 4.30 (s, 1H), 4.49–4.62 (m, 2H), 5.57 (d, 1H, J = 8), 5.77 (d, 1H, J = 4), 5.95 (d, 1H, J = 8), 7.44–7.51 (m, 3H), 8.19 (s, 1H), 8.39 (s, 1H), 8.95 (bs, 1H). Anal. (C₂₀H₂₀Cl₂N₆O₅) C, H, N.

1-Deoxy-1-[6-[[(4-methoxyphenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (11):

yield 80%; white solid, mp 167–169 °C (EtOH); IR (KBr) cm⁻¹ 3550–3150, 1710, 1640, 1530, 1270; ¹H NMR (DMSO-d₆) δ 1.07 (t, 3H, J = 7), 3.15–3.23 (m, 2H), 3.86 (s, 3H), 4.20–4.24 (m, 1H), 4.35 (d, 1H, J = 2), 4.70–4.75 (m, 1H), 5.69 (d, 1H, J = 7), 5.77 (d, 1H, J = 4), 6.11 (d, 1H, J = 7), 7.08 (d, 2H, J = 9), 8.04 (d, 2H, J = 9), 8.44 (bs, 1H), 8.77 (s, 1H), 8.78 (s, 1H), 11.12 (s, 1H). Anal. (C₂₀H₂₂N₆O₆) C, H, N.

1-Deoxy-1-[6-[[(2-chlorophenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (12):

yield 78%; white solid, mp 179–180 °C (EtOH); IR (KBr) cm⁻¹ 3510–3050, 1720, 1660, 1510, 1250; ¹H NMR (DMSO-d₆) δ 1.06 (t, 3H, J = 7), 3.15–3.22 (m, 2H), 4.22 (bs, 1H), 4.34 (s, 1H), 4.68–4.70 (m, 1H), 5.66 (d, 1H, J = 7), 5.76 (d, 1H, J = 4), 6.10 (d, 1H, J = 7), 7.43–7.61 (m, 4H), 8.44 (bs, 1H), 8.67 (s, 1H), 8.82 (bs, 1H), 11.53 (bs, 1H). Anal. (C₁₉H₁₉N₆O₅) C, H, N.

1-Deoxy-1-[6-[[(phenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (13):

yield 67%; white solid, mp 157–159 °C (EtOH); IR (KBr) cm⁻¹ 3500–3000, 1715, 1630, 1535, 1260; ¹H NMR (DMSO-d₆) δ 1.03 (t, 3H, J = 7), 3.16–3.19 (m, 2H), 3.74 (bs, 2H), 4.22 (bs, 1H), 4.36 (s, 1H), 4.51–4.63 (m, 1H), 6.13 (d, 1H, J = 6.8), 7.48–7.59 (m, 3H), 8.04–8.11 (m, 2H), 8.52 (bs, 1H), 8.79 (s, 1H), 8.83 (s, 1H), 11.22 (s, 1H). Anal. (C₁₉H₂₀N₆O₅) C, H, N.

1-Deoxy-1-[6-[[[(benzyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (30):

yield 70%;white solid, mp 184–186 °C (EtOH); IR (KBr) cm⁻¹ 3500–3100, 1675, 1620, 1565, 1520, 1310; ¹H NMR (DMSO-d₆) δ 1.06 (t, 3H, J = 7), 3.10–3.20 (m, 2H), 4.05 (bs, 2H), 4.17–4.20 (m, 1H), 4.36 (s, 1H), 4.47–4.50 (m, 2H), 4.55–4.65 (m, 1H), 6.05–6.10 (m, 1H), 7.17–7.40 (m, 5H), 8.30–8.40 (m, 1H), 8.62 (s, 1H), 8.86 (s, 1H), 9.50–9.60 (m, 1H), 10.30 (bs, 1H). Anal. (C₂₀H₂₄N₇O₅) C, H, N.

1-Deoxy-1-[6-[[[(4-sulfonamidophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (31):

yield 63%; white solid, mp 183–185 °C (EtOH); IR (KBr) cm⁻¹ 3550–3050, 1715, 1630, 1545, 1370, 1250; ¹H NMR (DMSO-d₆) δ 1.07 (t, 3H, J = 7), 3.16–3.26 (m, 2H), 4.21 (bs, 1H), 4.36 (s, 1H), 4.63–4.69 (m, 1H), 5.62 (d, 1H, J = 6.8), 5.78 (d, 1H, J = 4), 6.11 (d, 1H, J = 6.8), 7.31 (bs, 2H), 7.81 (s, 4H), 8.49 (bs, 1H), 8.75 (s, 1H), 8.89 (s, 1H), 9.69 (bs, 1H), 11.92 (bs, 1H). Anal. (C₁₉H₂₂N₈O₇S) C, H, N.

1-Deoxy-1-[6-[[[(4-acetylphenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (32):

yield 83%; white solid, mp 187 °C (EtOH); IR (KBr) cm⁻¹ 3550–3100, 1740, 1720, 1630, 1525, 1215; ¹H NMR (DMSO-d₆) δ 1.08 (t, 3H, J = 7), 2.55 (s, 3H), 3.16–3.23 (m, 2H), 4.18–4.20 (m, 1H), 4.21 (d, 1H, J = 2), 4.45 (bs, 2H), 4.64–4.69 (m, 1H), 6.11 (d, 1H, J = 7), 7.77 (d, 2H, J = 9), 7.98 (d, 2H, J = 9), 8.49 (bs, 1H), 8.76 (s, 1H), 8.88 (s, 1H), 10.62 (bs, 1H), 11.94 (bs, 1H). Anal. (C₂₁H₂₃N₇O₆) C, H, N.

1-Deoxy-1-[6-[[[((R)-1-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (33):

yield 65%; white solid, mp 153–155 °C (EtOH); IR (KBr) cm⁻¹ 3550–3100, 1720, 1635, 1535, 1240; ¹H NMR (DMSO-d₆) δ 1.05 (t, 3H, J = 7), 1.51 (d, 3H, J = 8), 3.14–3.21 (m, 2H), 3.35–3.41 (m, 1H), 4.20 (s, 1H), 4.36 (s, 1H), 4.93–5.00 (m, 3H), 6.09 (d, 1H, J = 6.8), 7.25–7.39 (m, 5H), 8.48 (bs, 1H), 8.65 (s, 1H), 8.89 (s, 1H), 9.51 (bs, 1H), 10.38 (bs, 1H). Anal. (C₂₁H₂₅N₇O₅) C, H, N.

1-Deoxy-1-[6-[[[((S)-1-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (34):

yield 72%; white solid, mp 151 °C (EtOH); IR (KBr) cm⁻¹ 3550–3100, 1725, 1630, 1525, 1240; ¹H NMR (DMSO-d₆) δ 1.06 (t, 3H, J = 7), 1.49 (d, 3H, J = 8), 3.15–3.21 (m, 2H), 3.28–3.39 (m, 1H), 4.00 (bs, 2H), 4.35 (s, 1H), 4.61–4.66 (m, 1H), 4.94–5.01 (m, 1H), 6.07 (d, 1H, J = 6.8), 7.24–7.39 (m, 5H), 8.48 (bs, 1H), 8.64 (s, 1H), 8.83 (s, 1H), 9.57 (bs, 1H), 10.12 (bs, 1H). Anal. (C₂₁H₂₅N₇O₅) C, H, N.

1-Deoxy-1-[6-[[[(5-methyl-isoxazol-3-yl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (35):

yield 58%; white solid, mp 197 °C (EtOH); IR (KBr) cm⁻¹ 3550–3050, 1715, 1620, 1535, 1240; ¹H NMR (DMSO-d₆) δ 1.06 (t, 3H, J = 7), 2.40 (s 3H), 3.12–3.25 (m, 2H), 4.21–4.23 (m, 1H), 4.36 (d, 1H, J = 2), 4.64–4.70 (m, 1H), 5.68 (m, 2H), 6.09 (d, 1H, J = 6.8), 6.67 (s, 1H), 8.46 (t, 1H, J = 6), 8.74 (s, 1H), 8.84 (s, 1H), 10.75 (bs, 1H), 12.19 (bs, 1H). Anal. (C₁₇H₂₀N₈O₆) C, H, N.

1-Deoxy-1-[6-[[[(1,3,4-thiadiazol-2-yl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (36):

yield 65%; white solid, mp 197 °C (EtOH); IR (KBr) cm⁻¹ 3450–300, 1720, 1635, 1515, 1240; ¹H NMR (DMSO-d₆) δ 1.06 (t, 3H, J = 7), 3.15–3.22 (m, 2H), 3.72 (bs, 2H), 4.21–4.24 (m, 1H), 4.36 (d, 1H, J = 2), 4.65–4.70 (m, 1H), 6.12 (d, 1H, J = 6.8), 8.45 (t, 1H, J = 7), 8.80 (s, 1H), 8.89 (s, 1H), 9.19 (s, 1H), 10.35 (bs, 1H), 11.38 (bs, 1H). Anal. (C₁₅H₁₇N₄O₅S) C, H, N.

1-Deoxy-1-[6-[[[(4-n-propyloxy-phenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (37):

yield 66%; white solid, mp 270 °C (EtOH); IR (KBr) cm⁻¹ 3450–3050, 1720, 1630, 1535, 1240; ¹H NMR (DMSO-d₆) δ 0.95 (t, 3H, J = 7), 1.09 (t, 3H, J = 7), 1.65–1.75 (m, 2H), 3.06–3.23 (m, 2H), 3.67 (bs, 2H), 3.89 (t, 2H, J = 7), 4.18–4.20 (m, 1H), 4.33 (s, 1H), 4.51–4.61 (m, 1H), 6.05 (d, 1H, J = 6.8), 6.91 (d, 2H, J = 9), 7.52 (d, 2H, J = 9), 7.92 (t, 1H, J = 7), 8.22 (s, 1H), 8.52 (s, 1H), 9.65 (bs, 1H), 11.52 (bs, 1H). Anal. (C₂₂H₂₇N₇O₆) C, H, N.

1-Deoxy-1-[6-[[[(2-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (38):

yield 82%; light brown solid, mp 166 °C (EtOH); IR (KBr) cm⁻¹ 3450–3000, 1725, 1610, 1560, 1210; ¹H NMR (DMSO-d₆) δ 1.04 (t, 3H, J = 7), 2.84–2.95 (m, 2H), 3.01–3.23 (m, 2H), 4.15–4.20 (m, 1H), 4.47 (s, 1H), 4.54–4.79 (m, 3H), 5.39–5.48 (m, 2H), 6.21 (d, 1H, J = 1.8), 6.53 (t, 1H, J = 6), 7.31–7.44 (m, 5H), 8.32 (s, 1H), 8.41 (s, 1H), 9.55 (bs, 1H), 11.18 (s, 1H). Anal. (C₂₁H₂₅N₇O₅) C, H, N.

1-Deoxy-1-[6-[[[(3,4-dimethoxy-2-phenylethyl)amino]-carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (39):

yield 68%; light brown solid, mp 120–122 °C (EtOH); IR (KBr) cm⁻¹ 3540–3060, 1734, 1633, 1500, 1215; ¹H NMR (DMSO-d₆) δ 1.11 (t, 3H, J = 7), 1.23 (t, 2H, J = 8), 2.88–3.01 (m, 2H), 3.78 (s, 3H), 3.81 (s, 3H), 4.18–4.23 (m, 1H), 4.42 (s, 1H), 4.54–4.59 (m, 1H), 4.87–5.02 (bs, 2H), 5.47–5.51 (m, 2H), 6.15 (d, 1H, J = 1.8), 6.59 (t, 1H, J = 2), 6.65 (s, 1H), 6.71 (s, 2H), 8.31 (s, 1H), 8.47 (s, 1H), 9.05 (s, 1H), 10.46 (bs, 1H). Anal. (C₂₃H₂₉N₇O₇) C, H, N.

1-Deoxy-1-[6-[[[(fur-2-yl-methyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (40):

yield 79%; brown solid mp 189–191 °C (EtOH); IR (KBr) cm⁻¹ 3500–3087, 1731, 1630, 1550, 1335, 1221; ¹H NMR (DMSO-d₆) δ 1.06 (t, 3H, J = 7), 2.94–3.05 (m, 2H), 4.20–4.24 (m, 1H), 4.38 (s, 1H), 4.56 (s, 2H, J = 6), 4.61–4.67 (m, 1H), 4.96–5.04 (bs, 2H), 6.17 (d, 1H, J = 1.8), 6.35–6.46 (m, 1H), 6.62 (s, 1H), 6.56 (t, 1H, J = 6), 7.48 (d, 1H, J = 2), 8.38 (s, 1H), 8.54 (s, 1H), 9.08 (s, 1H), 9.94 (bs, 1H). Anal. (C₁₈H₂₁N₇O₆) C, H, N.

1-Deoxy-1-[6-[[[(4-(pyridin-2-yl-aminosulfonyl)phenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (41):

yield 88%; brown solid mp 163 °C (EtOH); IR (KBr) cm⁻¹ 3500–3050, 1734, 1633, 1552, 1345, 1213; ¹H NMR (DMSO-d₆) δ 1.05 (t, 3H, J = 7), 2.99–3.18 (m, 2H), 4.23–4.26 (m, 1H), 4.42 (s, 1H), 4.54–4.62 (m, 1H), 4.91–5.15 (bs, 2H), 6.16 (d, 1H, J = 1.8), 6.71 (t, 1H, J = 6), 7.18–7.31 (m, 1H), 7.57 (d, 2H, J = 9), 7.63–7.71 (m, 3H), 7.95 (d, 2H, J = 9), 8.37 (s, 1H), 8.78 (s, 1H), 9.05 (s, 1H), 10.15 (bs, 1H), 12.45 (bs, 1H). Anal. (C₂₄H₂₅N₉O₇S) C, H, N.

1-Deoxy-1-[6-[[[(4-(5-methyl-isoxazol-3-yl-aminosulfonyl)phenyl)amino] carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (42):

yield 77%; pale yellow solid mp 184 °C (EtOH); IR (KBr) cm⁻¹ 3550–3150, 1730, 1638, 1550, 1350, 1210; ¹H NMR (DMSO-d₆) δ 1.08 (t, 3H, J = 7), 2.30 (s, 3H), 3.17–3.20 (m, 2H), 4.22–4.23 (m, 1H), 4.36 (s, 1H), 4.40–4.95 (m, 3H), 6.11 (d, 1H, J = 1.8), 6.15 (s, 1H), 6.68 (t, 1H, J = 6), 7.37 (d, 2H, J = 9), 7.84 (d, 2H, J = 9), 8.42 (bs, 1H), 8.73 (s, 1H), 8.85 (s, 1H), 9.63 (bs, 1H), 11.37 (bs, 1H), 12.02 (s, 1H). Anal. (C₂₃H₂₅N₉O₈S) C, H, N.

Deoxy-1-[6-[[[(4-(pyrimidin-2-yl-aminosulfonyl)-phenyl)amino] carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (43):

yield 80%; brown solid mp 184 °C (EtOH); IR (KBr) cm⁻¹ 3550–3100, 1720, 1630, 1555, 1355, 1200; ¹H NMR (DMSO-d₆) δ 1.08 (t, 3H, J = 7), 3.17–3.23 (m, 2H), 4.22–4.25 (m, 1H), 4.37 (s, 1H), 4.64–4.70 (m, 1H), 4.90–5.12 (bs, 2H), 6.11 (d, 1H, J = 1.8), 6.60 (t, 1H, J = 6), 7.03–7.08 (m, 1H), 7.81 (d, 2H, J = 9), 7.98 (d, 2H, J = 9), 8.51 (d, 2H, J = 6), 8.74 (s, 1H), 8.89 (s, 1H), 9.55 (bs, 1H), 11.19 (bs, 1H), 11.90 (s, 1H). Anal. (C₂₃H₂₄N₁₀O₇S) C, H, N.

1-Deoxy-1-[6,6-[[[bis-(4-nitrophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (44):

yield 52%; white solid, mp 213 °C (EtOH); IR (KBr) cm⁻¹ 3500–3050, 1720, 1630, 1545, 1340, 1220; ¹H NMR (DMSO-d₆) δ 1.07 (t, 3H, J = 7), 3.12–3.23 (m, 2H), 4.21–4.23 (m, 1H), 4.35 (d, 1H, J = 2), 4.64–4.69 (m, 1H), 5.69 (bs, 2H), 6.09 (d, 1H, J = 6.8), 7.68 (d, 2H, J = 9), 7.91 (d, 2H, J = 9), 8.18–8.29 (m, 4H), 8.83 (s, 1H), 9.69 (s, 1H), 10.68 (bs, 1H), 12.26 (bs, 1H). Anal. (C₂₆H₂₉N₁₀O₁₀) C, H, N.

1-Deoxy-1-[6,6-[[[bis-(5-chloro-pyridin-2-yl)amino]-carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (45):

yield 68%; white solid, mp 220 °C (EtOH); IR (KBr) cm⁻¹ 3550–3100, 1725, 1630, 1525, 1250; ¹H NMR (DMSO-d₆) δ 1.06 (t, 3H, J = 7), 3.15–3.22 (m, 2H), 4.22–4.24 (m, 1H), 4.36 (d, 2H, J = 2), 4.64–4.69 (m, 1H), 5.95 (bs, 2H), 6.10 (d, 1H, J = 6.8), 7.80–8.06 (m, 6H), 8.39 (t, 1H, J = 7), 8.76 (s, 1H), 8.93 (s, 1H), 10.39 (bs, 1H), 12.11 (bs, 1H). Anal. (C₂₄H₂₂Cl₂N₈O₆) C, H, N.

General Procedure for the Synthesis of 2-Chloro- and 2-Iodo-1-deoxy-1-[6-[[[(substituted)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (48–51).

The appropriate commercially available isocyanate (2.1 mmol) was added to a solution of 2′,3′-O-isopropylidene-2-chloro-NECA (46)⁴³ or to 2′,3′-O-isopropylidene-2-iodo-NECA (47)⁴⁴ (0.42 mmol) and a catalytic amount of triethylamine in THF (10 mL), and the mixture was heated at 110 °C in a steel bomb for 24 h. After the mixture was concentrated in vacuo, the residue was flash chromatographed on a silica gel column eluting with CHCl₃–MeOH (99:1) to give the desired protected nucleosides 48–51.

2-Chloro-1-deoxy-1-[6-[[[(3-chlorophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (48):

yield 33%; white solid, mp 98–100 °C dec (CH₃CN–EtOH); IR (KBr) cm⁻¹ 3400–3050, 1685, 1650, 1520, 1235; ¹H NMR (DMSO-d₆) δ 0.66 (t, 3H, J = 7.2 Hz), 1.37 (s, 3H), 1.56 (s, 3H), 2.74–2.92 (br m, 2H), 4.61 (s, 1H), 5.42 (s, 2H), 6.42 (s, 1H), 7.17 (d, 1H, J = 7.0 Hz), 7.32–7.42 (m, 2H),7.60–7.70 (m, 1H), 7.79 (s, 1H), 8.59 (s, 1H), 10.64 (s, 1H), 10.79 (s, 1H). Anal. (C₂₂H₂₃Cl₂N₇O₅) C, H, N.

2-Chloro-1-deoxy-1-[6-[[[(4-methoxyphenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (49):

yield 35%; vitreous solid; IR (KBr) cm⁻¹ 3450–3050, 1680, 1650, 1510, 1230; ¹H NMR (DMSO-d₆) δ 0.67 (t, 3H, J = 7.0 Hz), 1.38 (s, 3H), 1.56 (s, 3H), 2.76–2.94 (m, 2H), 3.76 (s, 3H), 4.61 (s, 1H), 5.42 (s, 2H), 6.41 (s, 1H), 6.93–7.03 (m, 2H), 7.41–7.53 (m, 2H), 7.65–7.75 (m, 1H), 8.61 (s, 1H), 10.43 (s, 1H), 11.11 (s, 1H). Anal. (C₂₃H₂₆ClN₇O₆) C, H, N.

2-Iodo-1-deoxy-1-[6-[[[(3-chlorophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (50):

yield 62%; white solid, mp 127–130 °C (CH₃CN–EtOH); IR (KBr) cm⁻¹ 3450–3050, 1690, 1650, 1520, 1240; ¹H NMR (DMSO-d₆) δ 0.66 (t, 3H, J = 7.2 Hz), 1.37 (s, 3H), 1.56 (s, 3H), 2.74–2.92 (br m, 2H), 4.61(s, 1H), 5.42 (s, 2H), 6.42 (s, 1H), 7.17 (d, 1H, J = 7.0 Hz), 7.32–7.42 (m, 2H), 7.56–7.66 (m, 1H), 7.79 (s, 1H), 8.50 (s, 1H), 10.64 (s, 1H), 11.33 (s, 1H). Anal. (C₂₂H₂₃ClIN₇O₅) C, H, N.

2-Iodo-1-deoxy-1-[6-[[[(4-methoxyphenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (51):

yield 70%; white solid, mp 127–130 °C (CH₃CN–EtOH); IR (KBr) cm⁻¹ 3450–3025, 1680, 1660, 1510, 1230; ¹H NMR (DMSO-d₆) δ 0.67 (t, 3H, J = 7.0 Hz), 1.38 (s, 3H), 1.56 (s, 3H), 2.76–2.94 (m, 2H), 3.76 (s, 3H), 4.61 (s, 1H), 5.42 (s, 2H), 6.41 (s, 1H), 6.93–7.03 (m, 2H), 7.41–7.53 (m, 2H), 7.54–7.65 (m, 1H), 8.49 (s, 1H), 10.43 (s, 1H), 11.11 (s, 1H). Anal. (C₂₃H₂₆IN₇O₆) C, H, N.

General Procedure for the Synthesis of 2-Chloro- and 2-Iodo-1-deoxy-1-[6-[[[(substituted)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (52–55).

To a solution of 2′,3′-O-isopropylidene-2-chloro-6-ureas (48–49) or 2′,3′–O-isopropylidene-2-iodo-6-ureas (50–51) (0.96 mmol) in dioxane (50 mL) was added 1 N HCl (50 mL), and the mixture was heated at 65 °C for 1 h. After the mixture was concentrated in vacuo, the residue was flash chromatographed on a silica gel column eluting with CHCl₃–MeOH (95:5) to give the desired compounds 52–55.

2-Chloro-1-deoxy-1-[6-[[[(3-chlorophenyl)amino]-carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (52):

yield 66%; white solid, mp 150–153 °C dec (EtOH); IR (KBr) cm⁻¹ 3500–3000, 1690, 1640, 1530, 1240; ¹H NMR (DMSO-d₆) δ 1.07 (t, 3H, J = 7.2 Hz), 3.13–3.31 (m, 2H), 4.25 (s, 1H,), 4.38 (d,1H, J = 2.1 Hz), 4.67 (pq, 1H), 6.05 (d, 1H, J = 6.7 Hz), 7.17 (d, 1H, J = 5.2 Hz), 7.35–7.44 (m, 2H), 8.82 (s, 1H), 8.26–8.33 (m, 1H), 8.87 (s, 1H), 10.80 (s, 1H), 11.33 (s, 1H). Anal. (C₁₉H₁₉Cl₂N₇O₅) C, H, N.

2-Chloro-1-deoxy-1-[6-[[[(4-methoxyphenyl)amino]-carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuron-amide (53):

yield 67%; white solid, mp 158–160 °C dec (EtOH); IR (KBr) cm⁻¹ 3400–3050, 1680, 1660, 1520, 1240; ¹H NMR (DMSO-d₆) δ 1.02–1.14 (m, 3H), 3.12–3.30 (m, 2H), 3.75 (s, 3H), 4.20–4.30 (m, 1H), 4.37 (s,1H), 4.61–4.72 (m, 1H), 6.04 (d, 1H, J = 6.8 Hz), 6.97 (d, 2H, J = 8.9 Hz), 7.47 (d, 2H, J = 8.9 Hz), 8.22–8.35 (m, 1H), 8.85 (s, 1H), 10.51 (s, 1H), 10.62 (s, 1H). Anal. (C₂₀H₂₂ClN₇O₆) C, H, N.

2-Iodo-1-deoxy-1-[6-[[[(3-chlorophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (54):

yield 85%; white solid, mp 202–204 °C dec (EtOH); IR (KBr) cm⁻¹ 3500–3000, 1700, 1640, 1530, 1240; ¹H NMR (DMSO-d₆) δ 1.07 (t, 3H, J = 7.2 Hz), 3.13–3.31 (m, 2H), 4.25 (s, 1H), 4.38 (d, 1H, J = 2.1 Hz), 4.67 (pq, 1H), 6.05 (d, 1H, J = 6.7 Hz), 7.17 (d, 1H, J = 5.2 Hz), 7.35–7.44 (m, 2H), 8.82 (s, 1H), 8.13–8.23 (m, 1H), 8.79 (s, 1H), 10.66 (s, 1H), 11.33 (s, 1H). Anal. (C₁₉H₁₉ClIN₇O₅) C, H, N.

2-Iodo-1-deoxy-1-[6-[[[(4-methoxyphenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuron-amide (55):

yield 89%; white solid, mp 214–218 °C dec (EtOH); IR (KBr) cm⁻¹ 3375–3050, 1680, 1650, 1510, 1240; ¹H NMR (DMSO-d₆) δ 1.01–1.12 (m, 3H), 3.12–3.32 (m, 2H), 3.76 (s, 3H), 4.20–4.28 (m, 1H), 4.36 (s,1H), 4.60–4.73 (m, 1H), 6.04 (d, 1H, J = 6.6 Hz), 6.93–7.04 (m, 2H), 7.44–7.56 (m, 2H), 8.11–8.21 (m, 1H), 8.76 (s, 1H), 10.45 (s, 1H), 11.09 (s, 1H). Anal. (C₂₀H₂₂IN₇O₆) C, H, N.

General Procedure for the Synthesis of 2-(Ar)alkynyl-1-deoxy-1-[6-[[[(substituted)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (56–62).

Cuprous iodide (0.12 mg), bis(triphenylphosphine)palladium dichloride (1.7 mg) and the appropriate terminal alkyne (0.6 mmol) were added to a solution of 2-iodo-6-ureas (54 or 55) (0.12 mmol) in a mixture of dry acetonitrile (4 mL), DMF (4 mL), and triethylamine (0.3 mL), and the mixture was stirred at room temperature for 16 h in an atmosphere of N₂. The solvent was removed under vacuum, and the residue was chromatographed on silica gel column, eluting with a suitable mixture of solvents (see below) to give the desired alkynyl derivatives 56–62.

2-(1-Hexyn-1-yl)-1-deoxy-1-[6-[[[(3-chlorophenyl)amino]-carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (56):

chromatography solvent, CHCl₃–MeOH (92:8); yield 75%; white solid, mp 196–198 °C dec (EtOH); IR (KBr) cm⁻¹ 3500–3150, 2235, 1680, 1660, 1545, 1260; ¹H NMR (DMSO-d₆) δ 0.91–1.0 (m, 3H), 1.02–1.13 (m, 3H), 1.40–1.71 (m, 4H), 2.52–2.60 (m, 2H), 3.19–3.34 (m, 2H), 4.18–4.26 (m, 1H), 4.37 (d, 1H, J = 2.2 Hz), 4.60–4.70 (m, 1H), 6.07 (d, 1H, J = 7.1 Hz), 7.11–7.22 (m, 1H), 7.41 (d, 2H, J = 5.1 Hz), 7.74 (s, 1H), 8.40–8.50 (m, 1H), 8.84 (s, 1H), 10.56 (s, 1H), 11.80 (s, 1H). Anal. (C₂₅H₂₈ClN₇O₅) C, H, N.

2-(Phenylethynyl)-1-deoxy-1-[6-[[[(3-chlorophenyl)-amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (57):

chromatography solvent, CHCl₃–MeOH (92:8); yield 59%; white solid, mp 210–214 °C dec (EtOH); IR (KBr) cm⁻¹ 3450–3050, 2210, 1680, 1670, 1550, 1270; ¹H NMR (DMSO-d₆) δ 1.02–1.13 (m, 3H), 3.17–3.34 (m, 2H), 4.20–4.29 (m, 1H), 4.39 (s, 1H), 4.63–4.74 (m, 1H), 6.13 (d, 1H, J = 7.0 Hz), 7.17 (d, 1H, J = 7.2 Hz), 7.36–7.59 (m, 5H), 7.69–7.79 (m, 3H), 8.35–8.44 (m, 1H), 8.92 (s, 1H), 10.61 (s, 1H), 11.65 (s, 1H). Anal. (C₂₇H₂₄ClN₇O₅) C, H, N.

2-(3-Hydroxy-3-phenyl-1-propyn-1-yl)-1-deoxy-1-[6-[[[(3-chlorophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (58):

chromatography solvent, CHCl₃-cC₆H₁₂–CH₃CN–MeOH (73:10:5:12); yield 43%; yellow solid, mp 214–216 °C dec (EtOH); IR (KBr) cm⁻¹ 3450–3050, 2325, 1700, 1640, 1540, 1250; ¹H NMR (DMSO-d₆) δ 0.96–1.14 (m, 3H), 3.04–3.30 (m, 2H), 4.30–4.45 (m, 2H), 4.78–4.90 (m, 1H), 6.19 (d, 1H, J = 6.6 Hz), 7.12–8.34 (m, 11H), 8.94 (s, 1H), 10.51 (s, 1H), 11.47 (s, 1H). Anal. (C₂₈H₂₆ClN₇O₆) C, H, N.

2-(1-Hexyn-1-yl)-1-deoxy-1-[6-[[[(4-methoxyphenyl)-amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (59):

chromatography solvent, CHCl₃–C₆H₆–MeOH (81:10:9); yield 71%; white solid, mp 175–178 °C dec (EtOH); IR (KBr) cm⁻¹ 3500–3000, 2240, 1660, 1600, 1520, 1240; ¹H NMR (DMSO-d₆) δ 0.91–1.02 (m, 3H), 1.04–1.17 (m, 3H), 1.43–1.70 (m, 4H), 2.50–2.61 (m, 2H), 3.18–3.32 (m, 2H), 3.77 (s, 3H), 4.18–4.28 (m, 1H), 4.37 (d, 1H, J = 1.7 Hz), 4.66 (pq, 1H), 6.07 (d, 1H, J = 7.3 Hz), 6.96 (d, 2H, J = 8.9 Hz), 7.48 (d, 2H, J = 9.1 Hz), 8.40–8.51 (m, 1H), 8.82 (s, 1H), 10.31 (s, 1H), 11.49 (s, 1H). Anal. (C₂₆H₃₁N₇O₆) C, H, N.

2-(Phenylethynyl)-1-deoxy-1-[6-[[[(4-methoxyphenyl)-amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (60):

chromatography solvent, CHCl₃–C₆H₆–MeOH (80:10:10); yield 60%; white solid, mp 204–206 °C dec (EtOH); IR (KBr) cm⁻¹ 3400–3050, 2205, 1675, 1660, 1510, 1240; ¹H NMR (DMSO-d₆) δ 0.98–1.14 (m, 3H), 3.15–3.34 (m, 2H), 3.77 (s, 3H), 4.20–4.30 (m, 1H), 4.39 (s, 1H), 4.65–4.75 (m, 1H), 6.12 (d, 1H, J = 6.9 Hz), 6.99 (d, 2H, J = 8.8 Hz), 7.44–7.62 (m, 5H), 7.64–7.78 (m, 2H), 8.34–8.47 (m, 1H), 8.90 (s, 1H), 10.36 (s, 1H), 11.29 (s, 1H). Anal. (C₂₈H₂₇N₇O₆) C, H, N.

2-(3-Hydroxy-3-phenyl-1-propyn-1-yl)-1-deoxy-1-[6-[[[(4-methoxyphenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (61):

chromatography solvent, CHCl₃–C₆H₆–MeOH (80:10:10); yield 40%; yellow solid, mp 218–220 °C dec (EtOH); IR (NaCl) cm⁻¹ 3450–3050, 2300, 1680, 1650, 1510, 1240; ¹H NMR (DMSO-d₆) δ 0.90–1.10 (m, 3H), 3.13–3.23 (m, 2H), 3.77 (s, 3H), 4.30–4.46 (m, 2H), 4.76–4.89 (m, 1H), 6.19 (d, 1H, J = 6.7 Hz), 6.88–7.02 (m, 2H), 7.43–7.83 (m, 5H), 8.03–8.30 (m, 4H), 8.92 (s, 1H), 10.30 (s, 1H), 11.32 (s, 1H). Anal. (C₂₉H₂₉N₇O₇) C, H, N.

2-(5-Phenyl-1-pentyn-1-yl)-1-deoxy-1-[6-[[[(4-methoxyphenyl)amino] carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (62):

chromatography solvent, CHCl₃–MeOH (94:6); yield 77%; white solid, mp 210–212 °C dec (EtOH); IR (NaCl) cm⁻¹ 3450–3050, 2240, 1680, 1660, 1510, 1240; ¹H NMR (DMSO-d₆) δ 1.02–1.16 (m, 3H), 1.86–2.04 (m, 2H), 2.49–2.64 (m, 2H), 2.73–2.88 (m, 2H), 3.16–3.32 (m, 2H), 3.71 (s, 3H), 4.18–4.29 (m, 1H), 4.37 (s, 1H), 4.66 (pq, 1H), 6.07 (d, 1H, J = 7.0 Hz), 6.84 (d, 2H, J = 8.1 Hz), 7.17–7.38 (m, 5H), 7.41–7.54 (m, 2H), 8.39–8.52 (m, 1H), 8.83 (s, 1H), 10.32 (s, 1H), 11.50 (s, 1H). Anal. (C₃₁H₃₃N₇O₆) C, H, N.

Methods for Receptor Binding Assays.

Procedures for preparation of rat brain membranes and CHO cell membranes were reported previously.^30,38,55 For binding experiments, membrane homogenates were frozen and stored at −20 °C for ≤2 months. Adenosine deaminase (ADA) was from Boehringer Mannheim (Indianapolis, IN). [³H]R-PIA was from Amersham (Arlington Heights, IL), and [³H]CGS 21680 was from DuPont NEN (Boston, MA). [¹²⁵I]-AB-MECA was prepared as described by Olah et al.³⁰

Binding of [¹²⁵I]-AB-MECA to CHO cells stably transfected with the rat A₃ receptor clone or to HEK-293 cells stably expressing the human A₃ receptor was performed essentially as described.^28,30,38,55 Assays were performed in 50 mM Tris/10 mM MgCl₂/1 mM EDTA buffer (adjusted to pH 8.26 at 5 °C) in glass tubes containing 100 μL of the membrane suspension, 50 μL of [¹²⁵I]-AB-MECA (final concentration 0.3 nM), and 50 μL of inhibitor. Inhibitors were routinely dissolved in DMSO. Concentrations of DMSO in incubations never exceeded 1%, at which concentration [¹²⁵I]-AB-MECA binding was not affected. Incubations were carried out in duplicate for 1 h at 37 °C and were terminated by rapid filtration over Whatman GF/B filters, using a Brandell cell harvester (Brandell, Gaithersburg, MD). Tubes were washed three times with 3 mL of buffer. Radioactivity was determined in a Beckman gamma 5500B γ-counter. Nonspecific binding was determined in the presence of 200 μM NECA. K_i values were calculated according to Cheng–Prusoff⁵⁶ assuming a K_d for [¹²⁵I]-AB-MECA of 1.48 nM.

Binding of [³H]R-PIA to A₁ receptors from rat cortical membranes and of [³H]CGS 21680 to A_2A receptors from rat striatal membranes was performed as described previously.⁵⁷Adenosine deaminase (2 units/mL) was present during the preparation of brain membranes. Additional deaminase was not added during incubation with the radioligand.

Binding of [³⁵S]GTP-γ-S.

The binding of [³⁵S]GTP-γ-S (Amersham, Chicago, IL, specific activity 1275 Ci/mmol) was carried out using rat RBL-2H3 mast cell membranes by the general method of Lorenzen et al.⁵⁸ Membranes (approximately 5 μg) were suspended in a buffer containing 50 mM Tris (pH 7.4), 3 units/mL adenosine deaminase, 1 mM EDTA, 1 mM DTT, 10 μM GDP, 100 mM NaCl, and 10 mM MgCl₂. [³⁵S]GTP-γ-S was added to a final concentration of 0.1 nM in a total volume of 125 μL, and the mixture was incubated for 2 h at 30 °C. Nonspecific binding was determined in the presence of 10 μM GTP-γ-S (Sigma, St. Louis, MO). Incubation of the reaction mixture was terminated by filtration over GF/B glass filter using a Brandell cell harvester and washed twice with the same buffer.

Acknowledgment.

This work was partially supported by Consiglio Nazionale delle Ricerche (CNR), Ministero della Ricerca Scientifica e Tecnologica (MURST, grant 40 and 60%).

Abbreviations:

[125I]AB-MECA: [125I]1-[6-[[(4-amino-3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-β-d-ribofuranuronamide
HENECA: that 2-(1-hexyn-1-yl)-1-(6-amino-9H-purin-9-yl)-1-deoxy-N-ethyl-β-d-ribofuranuronamide
CHO: cells, Chinese hamster ovary cells
CGS 21680: 2-[4-[(2-carboxyethyl)phenyl]ethyl-amino]1-(6-amino-9H-purin-9-yl)-1-deoxy-N-ethyl-β-d-ribofuranuronamide
Cl-IB-MECA: 2-Chloro-1-[6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-β-d-ribofuranuronamide
R-PIA: 1-[6-[[(phenylisopropyl]amino]-9H-purin-9-yl]-1-deoxy-β-d-ribofuranose
DMSO: dimethysulfoxide
EDTA: ethylenediaminotetraacetate, I-AB-MECA, 1-[6-[[(4-amino-3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-β-d-ribofuranuronamide
IB-MECA: 1-[6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-β-d-ribofuranuronamide
Ki: equilibrium inhibition constant
NECA: 1-(6-amino-9H-purin-9-yl)-1-deoxy-N-ethyl-β-d-ribofuranuronamide
PHPNECA: 2-(3-hydroxy-3-phenyl-1-propyn-1-yl)-1-(6-amino-9H-purin-9-yl)-1-deoxy-N-ethyl-β-d-ribofuranuronamide
THF: tetrahydrofuran
Tris: tris(hydroxymethyl)aminomethane

References

(1).(a) Libert F; Schiffmann SN; Lefort A; Parmentier M; Gerard C; Dumont JE; Vanderhaeghen JJ; Vassart G The orphan receptor cDNA RDC7 encodes an A₁ adenosine receptor. EMBO J. 1991, 10, 1677–1682. [DOI] [PMC free article] [PubMed] [Google Scholar]; (b) Fredholm BB; Abbracchio MP; Burnstock G; Daly JW; Harden TK; Jacobson KA: Williams M Nomenclature and classification of purinoceptors. Pharmacol. Rev 1994, 46, 143–156. [PMC free article] [PubMed] [Google Scholar]
(2).Maenhaut C; Sande JV; Libert F; Abramowicz M; Parmentier M; Vanderhaeghen JJ; Dumont JE; Vassart G; Schiffmann S RDC8 codes for an adenosine A₂ receptor with physiological constitutive activity. Biochem. Biophys. Res. Commun 1990, 173, 1169–1178. [DOI] [PubMed] [Google Scholar]
(3).Stehle JH; Rivkees SA; Lee JJ; Weaver DR; Deeds JD; Reppert SM Molecular cloning and expression of the cDNA for a novel A₂-adenosine receptor subtype. Mol. Endocrinol 1992, 6, 384–393. [DOI] [PubMed] [Google Scholar]
(4).Zhou QY; Li CY; Olah ME; Johnson RA; Stiles GL; Civelli O Molecular cloning and characterization of an adenosine receptor-the A₃ adenosine receptor. Proc. Natl. Acad. Sci. U.S.A 1992, 89, 7432–7436. [DOI] [PMC free article] [PubMed] [Google Scholar]
(5).Londos C; Cooper DM; Wolff J Subclasses of external adenosine receptors. Proc. Natl. Acad. Sci. U.S.A 1980, 77, 2551–2554. [DOI] [PMC free article] [PubMed] [Google Scholar]
(6).Hamprecht B; Van Calker D Nomenclature of adenosine receptors. Trends Pharmacol. Sci 1985, 6, 153–154. [Google Scholar]
(7).Ukena D; Olsson RA; Daly JW Definition of subclasses of adenosine receptors associated with adenylate cyclase: interaction of adenosine analogs with inhibitory A₁ receptors and stimulatory A₂ receptors. Can. J. Physiol. Pharmacol 1987, 65, 365–376. [DOI] [PubMed] [Google Scholar]
(8).Ramkumar V; Stiles GL; Beaven MA; Ali H The A₃AR is the unique adenosine receptor which facilitates release of allergic mediators in mast cells. J. Biol. Chem 1993, 268, 6871–6890. [PubMed] [Google Scholar]
(9).Abbracchio MP; Brambilla R; Ceruti S; Kim HO; von Lubitz DKJE; Jacobson KA; Cattabeni F G-protein-dependent activation of phospholipase C by adenosine A₃ receptors in rat brain. Mol. Pharmacol 1995, 48, 1038–1045. [PubMed] [Google Scholar]
(10).(a) Bruns RF; Daly JW; Snyder SH Adenosine receptors in brain membranes: binding of N-cyclohexyl[³H] adenosine and 1,3-diethyl-8[³H] phenylxanthine. Proc. Natl. Acad. Sci. U.S.A 1980, 77, 5547–5551. [DOI] [PMC free article] [PubMed] [Google Scholar]; (b) Jacobson KA; van Rhee AM Development of selective purinoceptor agonists and antagonists. In Purinergic Approaches Experimental Therapy; Jacobson KA, Jarvis MF, Eds; Wiley-Liss Publisher: New York, 1997; pp 101–128. [Google Scholar]
(11).Olsson RA; Kusachi S; Thompson SD; Ukena D; Padgett W; Daly JW N⁶-substituted N-alkyladenosine-5′-uronamides: bifunctional ligands having recognition groups for A₁ and A₂ adenosine receptors. J. Med. Chem 1986, 29, 1683–1689. [DOI] [PubMed] [Google Scholar]
(12).Jacobson KA; Nikodijevic O; Ji XD; Berkich DA; Eveleth D; Dean RL; Hiramatsu K; Kassel NF; van Galen PJM; Lee KS; Bartus RT; Daly JW; Lanoue KF; Maillard M Synthesis and biological activity of N6-(p-sulfophenyl) alkyl and N⁶-sulfoalkyl derivatives of adenosine. Water soluble and peripherally selective adenosine agonists. J. Med. Chem 1992, 35, 4143–4149. [DOI] [PMC free article] [PubMed] [Google Scholar]
(13).Shamim MT; Ukena D; Padgett WL; Hong O; Daly JW 8-Aryl- and 8-cycloalkyl-1,3-dipropylxanthines: further potent and selective antagonists for A₁-adenosine receptors. J. Med. Chem 1988, 31, 613–617. [DOI] [PubMed] [Google Scholar]
(14).Linden J; Patel A; Earl CQ; Craig RH; Daluge SM [¹²⁵I]-labelled 8-phenylxanthine derivatives: antagonist radio-ligands for adenosine A₁ receptors. J. Med. Chem 1988, 31, 745–751. [DOI] [PubMed] [Google Scholar]
(15).Jacobson KA; de la Cruz R; Schulick R; Kiriasis KL; Padgett W; Pfleiderer W; Kirk KL; Neumeyer JL; Daly JW 8-substituted xanthines as antagonists at A₁ and A₂ adenosine receptors. Biochem. Pharmacol 1988, 37, 3653–3661. [DOI] [PMC free article] [PubMed] [Google Scholar]
(16).Cristalli G; Eleuteri A; Vittori S; Volpini R; Lohse MJ; Klotz KN 2-alkynyl derivatives of adenosine and adenosine-5′-N-ethyluronamide as selective agonists at A₂ adenosine receptors. J. Med. Chem 1992, 35, 2363–2368. [DOI] [PubMed] [Google Scholar]
(17).Jarvis MF; Schulz R; Hutchison AJ; Do UH; Sills MA; Williams M [³H]CGS 21680, a selective A₂ adenosine receptor agonist directly labels A₂ receptors in rat brain. J. Pharmacol. Exp. Ther 1989, 251, 888–893. [PubMed] [Google Scholar]
(18).Shimada J; Suzuki K; Nonaka H; Ishii A; Ichikawa S (E)-1,3-dialkyl-7-methyl-8-(3,4,5-trimethoxystyryl)xanthines: potent and selective adenosine A₂ antagonists. J. Med. Chem 1992, 35, 2342–2345. [DOI] [PubMed] [Google Scholar]
(19).(a) Jacobson KA; Gallo-Rodriguez C; Melman N; Fischer B; Maillard M; van Bergen A; van Galen PJM; Karton Y Structure–activity relationship of 8-styrylxanthines as A₂-selective adenosine antagonists. J. Med. Chem 1993, 36, 1333–1342. [DOI] [PMC free article] [PubMed] [Google Scholar]; (b) Baraldi PG; Manfredini S; Simoni D; Zappaterra L; Zocchi C; Dionisotti S; Ongini E Synthesis and activity of new pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c] pyrimidine and 1,2,3-triazolo[4,5-e]1,2,4-triazolo[1,5-c]pyrimidine displaying potent and selective activity as A_2A adenosine receptor antagonist. Bioorg. Med. Chem. Lett 1994, 4, 2539–2544. [Google Scholar]; (c) Baraldi PG; Cacciari B; Spalluto G; Borioni A; Viziano M; Dionisotti S; Ongini E Current developments of A_2A adenosine receptor antagonists. Cur. Med. Chem 1995, 2, 707–722. [Google Scholar]
(20).Richardson PJ; Kase H; Jenner PG Trends Pharmacol. Sci. 1997, 18, 338–344. [DOI] [PubMed] [Google Scholar]
(21).Schingnitz G; Küfner-Mühl U; Ensinger H; Lehr E; Kuhn FJ Selective A₁-antagonists for treatment of cognitive deficits. Nucleosides Nucleotides. 1991, 10, 1067–1076. [Google Scholar]
(22).(a) Bridges AJ; Moos WH; Szotek DL; Trivedi BK; Bristol JA; Heffner TG; Bruns RF; Downs DA N⁶-(2,2-diphenylethyl)adenosine, a novel adenosine receptor agonist with antipsychotic-like activity. J. Med. Chem 1987, 30, 1709–1711. [DOI] [PubMed] [Google Scholar]; (b) Ferré S; O’Connor WT; Snaprud P; Ungerstedt U; Fuxe K Antagonistic interactions between adenosine A_2A receptors and dopamine D₂ receptors in the ventral striopallidal system-implications for the treatment of schizophrenia. Neuroscience. 1994, 63, 765–773. [DOI] [PubMed] [Google Scholar]
(23).(a) Knutsen LJS; Murray TF Adenosine and ATP in epilepsy. In Purinergic Approaches Experimental Therapy Jacobson KA, Jarvis MF, Eds; Wiley-Liss Publisher: New York, 1997; pp 423–447. [Google Scholar]; (b) Jacobson KA; Trivedi BK; Churchill PC; Williams M Novel therapeutics acting via purine receptors. Biochem. Pharmacol 1991, 41, 1399–1410. [DOI] [PMC free article] [PubMed] [Google Scholar]
(24).Jacobson KA; Kim HO; Siddiqi SM; Olah ME; Stiles G; von Lubitz DKJ A3 adenosine receptors: design of selective ligands and therapeutic prospects. Drugs Future. 1995, 20, 689–699. [DOI] [PMC free article] [PubMed] [Google Scholar]
(25).Ji XD; Melman N; Jacobson KA Interactions of flavonoids and other phytochemicals with adenosine receptors. J. Med. Chem 1996, 39, 781–788. [DOI] [PMC free article] [PubMed] [Google Scholar]
(26).Karton Y; Jiang J-I; Ji XD; Melman N; Olah ME; Stiles GL; Jacobson KA Synthesis and biological activities of flavonoid derivatives as A₃ adenosine receptor antagonists. J. Med. Chem 1996, 39, 2293–2301. [DOI] [PMC free article] [PubMed] [Google Scholar]
(27).van Rhee AM; Jiang J-I; Melman N; Olah ME; Stiles GL; Jacobson KA Intercaction of 1,4-dihydropyridine and pyridine derivative with adenosine receptors: selectivity for A₃ receptors. J. Med. Chem 1996, 39, 2980–2989. [DOI] [PMC free article] [PubMed] [Google Scholar]
(28).Kim Y-C, Ji XD; Jacobson KA Derivatives of the triazoloquinazoline adenosine antagonist (CGS15943) are selective for the human A3 receptor subtype. J. Med. Chem 1996, 39, 4142–4148. [DOI] [PMC free article] [PubMed] [Google Scholar]
(29).Kim HO; Ji XD; Siddiqi SM; Olah ME; Stiles GL; Jacobson KA 2-substitution of N⁶-benzyladenosine-5′-uronamides enhances selectivity for A₃ adenosine receptors. J. Med. Chem 1994, 37, 3614–3621. [DOI] [PMC free article] [PubMed] [Google Scholar]
(30).Olah ME; Gallo-Rodriguez C; Jacobson KA; Stiles GL [¹²⁵I]AB-MECA, a high affinity radioligand for the rat A₃ adenosine receptor. Mol. Pharmacol 1994, 45, 978–982. [PMC free article] [PubMed] [Google Scholar]
(31).Linden J Cloned adenosine A₃ receptors-pharmacological properties, species, differences and receptor functions. Trends Pharmacol. Sci 1994, 15, 298–306. [DOI] [PubMed] [Google Scholar]
(32).Hannon JP; Pfannkuche HJ; Fozard JR A role for mast cells in adenosine A₃ receptor-mediated hypothension in the rat. Br. J. Pharmacol 1995, 115, 945–952. [DOI] [PMC free article] [PubMed] [Google Scholar]
(33).Fozard JR; Pfannkuche HJ; Schuurman HJ Mast cell degranulation following adenosine A₃ receptor activation in rats. Eur. J. Pharmacol 1996, 298, 293–297. [DOI] [PubMed] [Google Scholar]
(34).Jacobson KA; Nikodijevic O; Shi D; Gallo-Rodriguez C; Olah ME; Stiles GL; Daly JW A role for central A₃-adenosine receptors: Mediation of behavioral depressant effects. FEBS Lett. 1993, 336, 57–60. [DOI] [PMC free article] [PubMed] [Google Scholar]
(35).von Lubitz DKJE; Lin RCS; Popik P; Carter MF; Jacobson KA Adenosine A₃ receptor stimulation and cerebral ischemia. Eur. J. Pharmacol 1994, 263, 59–67. [DOI] [PMC free article] [PubMed] [Google Scholar]
(36).Kohno Y; Sei Y; Koshiba M; Kim HO; Jacobson KA Induction of apoptosis in HL-60 human promyelocytic leukemia cells by selective adenosine A₃ receptor agonists. Biochem. Biophys. Res. Commun 1996, 219, 904–910. [DOI] [PMC free article] [PubMed] [Google Scholar]
(37).Jacobson KA; van Galen PJM; Williams M Perspective Adenosine Receptors: pharmacology, structure–activity relationship and therapeutic potential. J. Med. Chem 1992, 35, 407–422. [DOI] [PMC free article] [PubMed] [Google Scholar]
(38).Gallo-Rodriguez C; Ji XD; Melman N; Siegman BD; Sanders LH; Orlina J; Pu QL; Olah ME; van Galen PJM; Stiles GL; Jacobson KA Structure–activity relationship of N⁶-benzyladenosine-5′-uronamides as A₃-selective adenosine agonists. J. Med. Chem 1994, 37, 636–646. [DOI] [PMC free article] [PubMed] [Google Scholar]
(39).Baraldi PG; Cacciari B; Spalluto G; Ji XD; Olah ME; Stiles G; Dionisotti S; Zocchi C; Ongini E; Jacobson KA Novel N6-(substituted-phenylcarbamoyl)adenosine-5′-uronamides as potent agonists for A₃ adenosine receptors. J. Med. Chem 1996, 39, 802–806. [DOI] [PubMed] [Google Scholar]
(40).Cristalli G; Volpini R; Vittori S; Camaioni E; Monopoli A; Conti A; Dionisotti S; Zocchi C; Ongini E 2-Alkynyl derivatives of adenosine-5′-N-ethyluronamide: selective A₂ adenosine receptor agonists with potent inhibitory activity on platelet aggregation. J. Med. Chem 1994, 37, 1720–1726. [DOI] [PubMed] [Google Scholar]
(41).Siddiqui SM; Jacobson KA; Esker JL; Olah ME; Ji X; Melman N; Tiwari KN; Secrist JA III; Schneller SW; Cristalli G; Stiles GL; Johnson CR; IJzerman AP Search for new purine- and ribose-modified adenosine analogues as selective agonists and antagonists at adenosine receptors. J. Med. Chem 1995, 38, 1174–1188. [DOI] [PMC free article] [PubMed] [Google Scholar]
(42).Kurita K; Iwakura Y Trichloromethyl-Chloroformate as a phosgene equivalent: 3-isocyanatopropanoylchloride. Organic Synthesis; Wiley & Sons: New York, 1988; Collect. Vol VI, pp 715–718. [Google Scholar]
(43).Hutchison AJ; Williams M; de Jesus R; Yokoyama R; Oei HH; Ghai GR; Webb RL; Zoganas HC; Stone GA; Jarvis MF 2-(Arylalkylamino)adenosin-5′-uronamides: a new class of highly selective adenosine A₂ receptor ligands. J. Med. Chem 1990, 33, 1919–1924. [DOI] [PubMed] [Google Scholar]
(44).Cristalli G; Volpini R; Vittori S; Camaioni E; Monopoli A; Conti A; Dionisotti S; Zocchi C; Ongini E 2-Aralkynyl and 2-heteroalkynyl derivatives of adenosine-5′-N-ethyluronamide as selective A_2a adenosine receptor agonists. J. Med. Chem 1995, 38, 1462–1472. [DOI] [PubMed] [Google Scholar]
(45).Bruns RF; Lu GH; Pugsley TA Characterization of the A₂ adenosine receptor labeled by [³H]NECA in rat striatal membranes. Mol. Pharmacol 1986, 29, 331–346. [PubMed] [Google Scholar]
(46).Siddiqi SM; Pearlstein RA; Sanders LH; Jacobson KA Comparative molecular field analysis of selective A₃ adenosine receptor agonists. Bioorg. Med. Chem 1995, 3, 1331–1343. [DOI] [PMC free article] [PubMed] [Google Scholar]
(47).van Galen PJM; Stiles GL; Michaels G; Jacobson KA Adenosine A1 and A2 receptors: structure-function relationships. Med. Res. Rev 1992, 12, 423–471. [DOI] [PMC free article] [PubMed] [Google Scholar]
(48).Müller CE; Scior T Adenosine receptors and their modulators. Pharmaceutica Acta Helvitiae. 1993, 68, 77–111. [DOI] [PubMed] [Google Scholar]
(49).Vittori S; Camaioni E; Di Francesco E; Volpini R; Monopoli A; Dionisotti S; Ongini E; Cristalli G 2-Alkenyl and 2-alkyl derivatives of adenosine and adenosine-5′-N-ethyluronamide: different affinity and selectivity of E- and Z-diastereomers at A_2A adenosine receptors. J. Med. Chem 1996, 39, 4211–4217. [DOI] [PubMed] [Google Scholar]
(50).Camaioni E; Di Francesco E; Vittori S; Volpini R; Cristalli G Adenosine receptor agonists: synthesis and biological evaluation of the diastereoisomers of 2-(3-hydroxy-3-phenyl-1-propyn-1-yl)NECA. J. Med. Chem 1997, 5, 2267–2275. [DOI] [PubMed] [Google Scholar]
(51).Bodwen K Electronic effects in drugs. In Comprehensive Medicinal Chemistry, 1st ed.; Hansch C, Sammes PG, Taylor JB, Eds.; Pergamon Press: Elmsford, NY, 1990; Vol. 4, pp 205–239. [Google Scholar]; (b) Leo AJ Methods of calculating partition coefficients. In Comprehensive Medicinal Chemistry, 1st ed.; Hansch C, Sammes PG, Taylor JB, Eds.; Pergamon Press: Elmsford, NY, 1990; Vol. 4, pp 295–319. [Google Scholar]
(52).Park KS; Hoffmann C; Kim HO; Padgett WL; Daly JW; Brambilla R; Motta C; Abbracchio MP; Jacobson KA Activation and desensitization of rat A₃-adenosine receptors by selective adenosine derivatives and xanthine-7-ribosides. Drug Dev. Res, submitted. [DOI] [PMC free article] [PubMed] [Google Scholar]
(53).Lohse MJ; Klotz K-N; Schwabe U; Cristalli G; Vittori S; Grifantini M 2-Chloro- N⁶-cyclopentyladenosine: a highly selective agonist at A₁ adenosine receptors. Naunyn–Schmiedeberg’s Arch. Pharmacol 1988, 337, 687–689. [DOI] [PubMed] [Google Scholar]
(54).Macek TA; Conn PJ PKC Inhibition of Group II mGlu Autoreceptor Function at Medial and Lateral Performant. Path. Soc. Neurosci 1997, Abstr. 685.17, 23, pp 1754. [Google Scholar]
(55).van Galen PJM; van Bergen AH; Gallo-Rodriguez C; Melman N; Olah ME; IJzerman AP; Stiles GL; Jacobson KA A binding site model and structure-activity relationship for the rat A₃ adenosine receptor. Mol. Pharmacol 1994, 45, 1101–1111. [PMC free article] [PubMed] [Google Scholar]
(56).Cheng YC; Prusoff HR Relationships between the inhibition constant (K_i) and the concentration of inhibitor which causes 50% inhibition (IC₅₀) of an enzymatic reaction. Biochem. Pharmacol 1973, 22, 3099–3108. [DOI] [PubMed] [Google Scholar]
(57).van Bergen A; van Galen PJM; Stiles GL; Jacobson KA A₃ receptors: Structure activity relationship and molecular modeling. ACS 206th National Meeting, Chicago, IL, August, 1993, Abstract MEDI217. [Google Scholar]
(58).Lorenzen A; Guerra L; Vogt H; Schwabe U Interaction of full and partial agonists of the A₁ adenosine receptor with receptor/G protein complexes in rat brain membranes. Mol. Pharmacol 1996, 49, 915–926. [PubMed] [Google Scholar]

[R1] (1).(a) Libert F; Schiffmann SN; Lefort A; Parmentier M; Gerard C; Dumont JE; Vanderhaeghen JJ; Vassart G The orphan receptor cDNA RDC7 encodes an A₁ adenosine receptor. EMBO J. 1991, 10, 1677–1682. [DOI] [PMC free article] [PubMed] [Google Scholar]; (b) Fredholm BB; Abbracchio MP; Burnstock G; Daly JW; Harden TK; Jacobson KA: Williams M Nomenclature and classification of purinoceptors. Pharmacol. Rev 1994, 46, 143–156. [PMC free article] [PubMed] [Google Scholar]

[R2] (2).Maenhaut C; Sande JV; Libert F; Abramowicz M; Parmentier M; Vanderhaeghen JJ; Dumont JE; Vassart G; Schiffmann S RDC8 codes for an adenosine A₂ receptor with physiological constitutive activity. Biochem. Biophys. Res. Commun 1990, 173, 1169–1178. [DOI] [PubMed] [Google Scholar]

[R3] (3).Stehle JH; Rivkees SA; Lee JJ; Weaver DR; Deeds JD; Reppert SM Molecular cloning and expression of the cDNA for a novel A₂-adenosine receptor subtype. Mol. Endocrinol 1992, 6, 384–393. [DOI] [PubMed] [Google Scholar]

[R4] (4).Zhou QY; Li CY; Olah ME; Johnson RA; Stiles GL; Civelli O Molecular cloning and characterization of an adenosine receptor-the A₃ adenosine receptor. Proc. Natl. Acad. Sci. U.S.A 1992, 89, 7432–7436. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R5] (5).Londos C; Cooper DM; Wolff J Subclasses of external adenosine receptors. Proc. Natl. Acad. Sci. U.S.A 1980, 77, 2551–2554. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R6] (6).Hamprecht B; Van Calker D Nomenclature of adenosine receptors. Trends Pharmacol. Sci 1985, 6, 153–154. [Google Scholar]

[R7] (7).Ukena D; Olsson RA; Daly JW Definition of subclasses of adenosine receptors associated with adenylate cyclase: interaction of adenosine analogs with inhibitory A₁ receptors and stimulatory A₂ receptors. Can. J. Physiol. Pharmacol 1987, 65, 365–376. [DOI] [PubMed] [Google Scholar]

[R8] (8).Ramkumar V; Stiles GL; Beaven MA; Ali H The A₃AR is the unique adenosine receptor which facilitates release of allergic mediators in mast cells. J. Biol. Chem 1993, 268, 6871–6890. [PubMed] [Google Scholar]

[R9] (9).Abbracchio MP; Brambilla R; Ceruti S; Kim HO; von Lubitz DKJE; Jacobson KA; Cattabeni F G-protein-dependent activation of phospholipase C by adenosine A₃ receptors in rat brain. Mol. Pharmacol 1995, 48, 1038–1045. [PubMed] [Google Scholar]

[R10] (10).(a) Bruns RF; Daly JW; Snyder SH Adenosine receptors in brain membranes: binding of N-cyclohexyl[³H] adenosine and 1,3-diethyl-8[³H] phenylxanthine. Proc. Natl. Acad. Sci. U.S.A 1980, 77, 5547–5551. [DOI] [PMC free article] [PubMed] [Google Scholar]; (b) Jacobson KA; van Rhee AM Development of selective purinoceptor agonists and antagonists. In Purinergic Approaches Experimental Therapy; Jacobson KA, Jarvis MF, Eds; Wiley-Liss Publisher: New York, 1997; pp 101–128. [Google Scholar]

[R11] (11).Olsson RA; Kusachi S; Thompson SD; Ukena D; Padgett W; Daly JW N⁶-substituted N-alkyladenosine-5′-uronamides: bifunctional ligands having recognition groups for A₁ and A₂ adenosine receptors. J. Med. Chem 1986, 29, 1683–1689. [DOI] [PubMed] [Google Scholar]

[R12] (12).Jacobson KA; Nikodijevic O; Ji XD; Berkich DA; Eveleth D; Dean RL; Hiramatsu K; Kassel NF; van Galen PJM; Lee KS; Bartus RT; Daly JW; Lanoue KF; Maillard M Synthesis and biological activity of N6-(p-sulfophenyl) alkyl and N⁶-sulfoalkyl derivatives of adenosine. Water soluble and peripherally selective adenosine agonists. J. Med. Chem 1992, 35, 4143–4149. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R13] (13).Shamim MT; Ukena D; Padgett WL; Hong O; Daly JW 8-Aryl- and 8-cycloalkyl-1,3-dipropylxanthines: further potent and selective antagonists for A₁-adenosine receptors. J. Med. Chem 1988, 31, 613–617. [DOI] [PubMed] [Google Scholar]

[R14] (14).Linden J; Patel A; Earl CQ; Craig RH; Daluge SM [¹²⁵I]-labelled 8-phenylxanthine derivatives: antagonist radio-ligands for adenosine A₁ receptors. J. Med. Chem 1988, 31, 745–751. [DOI] [PubMed] [Google Scholar]

[R15] (15).Jacobson KA; de la Cruz R; Schulick R; Kiriasis KL; Padgett W; Pfleiderer W; Kirk KL; Neumeyer JL; Daly JW 8-substituted xanthines as antagonists at A₁ and A₂ adenosine receptors. Biochem. Pharmacol 1988, 37, 3653–3661. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R16] (16).Cristalli G; Eleuteri A; Vittori S; Volpini R; Lohse MJ; Klotz KN 2-alkynyl derivatives of adenosine and adenosine-5′-N-ethyluronamide as selective agonists at A₂ adenosine receptors. J. Med. Chem 1992, 35, 2363–2368. [DOI] [PubMed] [Google Scholar]

[R17] (17).Jarvis MF; Schulz R; Hutchison AJ; Do UH; Sills MA; Williams M [³H]CGS 21680, a selective A₂ adenosine receptor agonist directly labels A₂ receptors in rat brain. J. Pharmacol. Exp. Ther 1989, 251, 888–893. [PubMed] [Google Scholar]

[R18] (18).Shimada J; Suzuki K; Nonaka H; Ishii A; Ichikawa S (E)-1,3-dialkyl-7-methyl-8-(3,4,5-trimethoxystyryl)xanthines: potent and selective adenosine A₂ antagonists. J. Med. Chem 1992, 35, 2342–2345. [DOI] [PubMed] [Google Scholar]

[R19] (19).(a) Jacobson KA; Gallo-Rodriguez C; Melman N; Fischer B; Maillard M; van Bergen A; van Galen PJM; Karton Y Structure–activity relationship of 8-styrylxanthines as A₂-selective adenosine antagonists. J. Med. Chem 1993, 36, 1333–1342. [DOI] [PMC free article] [PubMed] [Google Scholar]; (b) Baraldi PG; Manfredini S; Simoni D; Zappaterra L; Zocchi C; Dionisotti S; Ongini E Synthesis and activity of new pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c] pyrimidine and 1,2,3-triazolo[4,5-e]1,2,4-triazolo[1,5-c]pyrimidine displaying potent and selective activity as A_2A adenosine receptor antagonist. Bioorg. Med. Chem. Lett 1994, 4, 2539–2544. [Google Scholar]; (c) Baraldi PG; Cacciari B; Spalluto G; Borioni A; Viziano M; Dionisotti S; Ongini E Current developments of A_2A adenosine receptor antagonists. Cur. Med. Chem 1995, 2, 707–722. [Google Scholar]

[R20] (20).Richardson PJ; Kase H; Jenner PG Trends Pharmacol. Sci. 1997, 18, 338–344. [DOI] [PubMed] [Google Scholar]

[R21] (21).Schingnitz G; Küfner-Mühl U; Ensinger H; Lehr E; Kuhn FJ Selective A₁-antagonists for treatment of cognitive deficits. Nucleosides Nucleotides. 1991, 10, 1067–1076. [Google Scholar]

[R22] (22).(a) Bridges AJ; Moos WH; Szotek DL; Trivedi BK; Bristol JA; Heffner TG; Bruns RF; Downs DA N⁶-(2,2-diphenylethyl)adenosine, a novel adenosine receptor agonist with antipsychotic-like activity. J. Med. Chem 1987, 30, 1709–1711. [DOI] [PubMed] [Google Scholar]; (b) Ferré S; O’Connor WT; Snaprud P; Ungerstedt U; Fuxe K Antagonistic interactions between adenosine A_2A receptors and dopamine D₂ receptors in the ventral striopallidal system-implications for the treatment of schizophrenia. Neuroscience. 1994, 63, 765–773. [DOI] [PubMed] [Google Scholar]

[R23] (23).(a) Knutsen LJS; Murray TF Adenosine and ATP in epilepsy. In Purinergic Approaches Experimental Therapy Jacobson KA, Jarvis MF, Eds; Wiley-Liss Publisher: New York, 1997; pp 423–447. [Google Scholar]; (b) Jacobson KA; Trivedi BK; Churchill PC; Williams M Novel therapeutics acting via purine receptors. Biochem. Pharmacol 1991, 41, 1399–1410. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R24] (24).Jacobson KA; Kim HO; Siddiqi SM; Olah ME; Stiles G; von Lubitz DKJ A3 adenosine receptors: design of selective ligands and therapeutic prospects. Drugs Future. 1995, 20, 689–699. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R25] (25).Ji XD; Melman N; Jacobson KA Interactions of flavonoids and other phytochemicals with adenosine receptors. J. Med. Chem 1996, 39, 781–788. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R26] (26).Karton Y; Jiang J-I; Ji XD; Melman N; Olah ME; Stiles GL; Jacobson KA Synthesis and biological activities of flavonoid derivatives as A₃ adenosine receptor antagonists. J. Med. Chem 1996, 39, 2293–2301. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R27] (27).van Rhee AM; Jiang J-I; Melman N; Olah ME; Stiles GL; Jacobson KA Intercaction of 1,4-dihydropyridine and pyridine derivative with adenosine receptors: selectivity for A₃ receptors. J. Med. Chem 1996, 39, 2980–2989. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R28] (28).Kim Y-C, Ji XD; Jacobson KA Derivatives of the triazoloquinazoline adenosine antagonist (CGS15943) are selective for the human A3 receptor subtype. J. Med. Chem 1996, 39, 4142–4148. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R29] (29).Kim HO; Ji XD; Siddiqi SM; Olah ME; Stiles GL; Jacobson KA 2-substitution of N⁶-benzyladenosine-5′-uronamides enhances selectivity for A₃ adenosine receptors. J. Med. Chem 1994, 37, 3614–3621. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R30] (30).Olah ME; Gallo-Rodriguez C; Jacobson KA; Stiles GL [¹²⁵I]AB-MECA, a high affinity radioligand for the rat A₃ adenosine receptor. Mol. Pharmacol 1994, 45, 978–982. [PMC free article] [PubMed] [Google Scholar]

[R31] (31).Linden J Cloned adenosine A₃ receptors-pharmacological properties, species, differences and receptor functions. Trends Pharmacol. Sci 1994, 15, 298–306. [DOI] [PubMed] [Google Scholar]

[R32] (32).Hannon JP; Pfannkuche HJ; Fozard JR A role for mast cells in adenosine A₃ receptor-mediated hypothension in the rat. Br. J. Pharmacol 1995, 115, 945–952. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R33] (33).Fozard JR; Pfannkuche HJ; Schuurman HJ Mast cell degranulation following adenosine A₃ receptor activation in rats. Eur. J. Pharmacol 1996, 298, 293–297. [DOI] [PubMed] [Google Scholar]

[R34] (34).Jacobson KA; Nikodijevic O; Shi D; Gallo-Rodriguez C; Olah ME; Stiles GL; Daly JW A role for central A₃-adenosine receptors: Mediation of behavioral depressant effects. FEBS Lett. 1993, 336, 57–60. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R35] (35).von Lubitz DKJE; Lin RCS; Popik P; Carter MF; Jacobson KA Adenosine A₃ receptor stimulation and cerebral ischemia. Eur. J. Pharmacol 1994, 263, 59–67. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R36] (36).Kohno Y; Sei Y; Koshiba M; Kim HO; Jacobson KA Induction of apoptosis in HL-60 human promyelocytic leukemia cells by selective adenosine A₃ receptor agonists. Biochem. Biophys. Res. Commun 1996, 219, 904–910. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R37] (37).Jacobson KA; van Galen PJM; Williams M Perspective Adenosine Receptors: pharmacology, structure–activity relationship and therapeutic potential. J. Med. Chem 1992, 35, 407–422. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R38] (38).Gallo-Rodriguez C; Ji XD; Melman N; Siegman BD; Sanders LH; Orlina J; Pu QL; Olah ME; van Galen PJM; Stiles GL; Jacobson KA Structure–activity relationship of N⁶-benzyladenosine-5′-uronamides as A₃-selective adenosine agonists. J. Med. Chem 1994, 37, 636–646. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R39] (39).Baraldi PG; Cacciari B; Spalluto G; Ji XD; Olah ME; Stiles G; Dionisotti S; Zocchi C; Ongini E; Jacobson KA Novel N6-(substituted-phenylcarbamoyl)adenosine-5′-uronamides as potent agonists for A₃ adenosine receptors. J. Med. Chem 1996, 39, 802–806. [DOI] [PubMed] [Google Scholar]

[R40] (40).Cristalli G; Volpini R; Vittori S; Camaioni E; Monopoli A; Conti A; Dionisotti S; Zocchi C; Ongini E 2-Alkynyl derivatives of adenosine-5′-N-ethyluronamide: selective A₂ adenosine receptor agonists with potent inhibitory activity on platelet aggregation. J. Med. Chem 1994, 37, 1720–1726. [DOI] [PubMed] [Google Scholar]

[R41] (41).Siddiqui SM; Jacobson KA; Esker JL; Olah ME; Ji X; Melman N; Tiwari KN; Secrist JA III; Schneller SW; Cristalli G; Stiles GL; Johnson CR; IJzerman AP Search for new purine- and ribose-modified adenosine analogues as selective agonists and antagonists at adenosine receptors. J. Med. Chem 1995, 38, 1174–1188. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R42] (42).Kurita K; Iwakura Y Trichloromethyl-Chloroformate as a phosgene equivalent: 3-isocyanatopropanoylchloride. Organic Synthesis; Wiley & Sons: New York, 1988; Collect. Vol VI, pp 715–718. [Google Scholar]

[R43] (43).Hutchison AJ; Williams M; de Jesus R; Yokoyama R; Oei HH; Ghai GR; Webb RL; Zoganas HC; Stone GA; Jarvis MF 2-(Arylalkylamino)adenosin-5′-uronamides: a new class of highly selective adenosine A₂ receptor ligands. J. Med. Chem 1990, 33, 1919–1924. [DOI] [PubMed] [Google Scholar]

[R44] (44).Cristalli G; Volpini R; Vittori S; Camaioni E; Monopoli A; Conti A; Dionisotti S; Zocchi C; Ongini E 2-Aralkynyl and 2-heteroalkynyl derivatives of adenosine-5′-N-ethyluronamide as selective A_2a adenosine receptor agonists. J. Med. Chem 1995, 38, 1462–1472. [DOI] [PubMed] [Google Scholar]

[R45] (45).Bruns RF; Lu GH; Pugsley TA Characterization of the A₂ adenosine receptor labeled by [³H]NECA in rat striatal membranes. Mol. Pharmacol 1986, 29, 331–346. [PubMed] [Google Scholar]

[R46] (46).Siddiqi SM; Pearlstein RA; Sanders LH; Jacobson KA Comparative molecular field analysis of selective A₃ adenosine receptor agonists. Bioorg. Med. Chem 1995, 3, 1331–1343. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R47] (47).van Galen PJM; Stiles GL; Michaels G; Jacobson KA Adenosine A1 and A2 receptors: structure-function relationships. Med. Res. Rev 1992, 12, 423–471. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R48] (48).Müller CE; Scior T Adenosine receptors and their modulators. Pharmaceutica Acta Helvitiae. 1993, 68, 77–111. [DOI] [PubMed] [Google Scholar]

[R49] (49).Vittori S; Camaioni E; Di Francesco E; Volpini R; Monopoli A; Dionisotti S; Ongini E; Cristalli G 2-Alkenyl and 2-alkyl derivatives of adenosine and adenosine-5′-N-ethyluronamide: different affinity and selectivity of E- and Z-diastereomers at A_2A adenosine receptors. J. Med. Chem 1996, 39, 4211–4217. [DOI] [PubMed] [Google Scholar]

[R50] (50).Camaioni E; Di Francesco E; Vittori S; Volpini R; Cristalli G Adenosine receptor agonists: synthesis and biological evaluation of the diastereoisomers of 2-(3-hydroxy-3-phenyl-1-propyn-1-yl)NECA. J. Med. Chem 1997, 5, 2267–2275. [DOI] [PubMed] [Google Scholar]

[R51] (51).Bodwen K Electronic effects in drugs. In Comprehensive Medicinal Chemistry, 1st ed.; Hansch C, Sammes PG, Taylor JB, Eds.; Pergamon Press: Elmsford, NY, 1990; Vol. 4, pp 205–239. [Google Scholar]; (b) Leo AJ Methods of calculating partition coefficients. In Comprehensive Medicinal Chemistry, 1st ed.; Hansch C, Sammes PG, Taylor JB, Eds.; Pergamon Press: Elmsford, NY, 1990; Vol. 4, pp 295–319. [Google Scholar]

[R52] (52).Park KS; Hoffmann C; Kim HO; Padgett WL; Daly JW; Brambilla R; Motta C; Abbracchio MP; Jacobson KA Activation and desensitization of rat A₃-adenosine receptors by selective adenosine derivatives and xanthine-7-ribosides. Drug Dev. Res, submitted. [DOI] [PMC free article] [PubMed] [Google Scholar]

[R53] (53).Lohse MJ; Klotz K-N; Schwabe U; Cristalli G; Vittori S; Grifantini M 2-Chloro- N⁶-cyclopentyladenosine: a highly selective agonist at A₁ adenosine receptors. Naunyn–Schmiedeberg’s Arch. Pharmacol 1988, 337, 687–689. [DOI] [PubMed] [Google Scholar]

[R54] (54).Macek TA; Conn PJ PKC Inhibition of Group II mGlu Autoreceptor Function at Medial and Lateral Performant. Path. Soc. Neurosci 1997, Abstr. 685.17, 23, pp 1754. [Google Scholar]

[R55] (55).van Galen PJM; van Bergen AH; Gallo-Rodriguez C; Melman N; Olah ME; IJzerman AP; Stiles GL; Jacobson KA A binding site model and structure-activity relationship for the rat A₃ adenosine receptor. Mol. Pharmacol 1994, 45, 1101–1111. [PMC free article] [PubMed] [Google Scholar]

[R56] (56).Cheng YC; Prusoff HR Relationships between the inhibition constant (K_i) and the concentration of inhibitor which causes 50% inhibition (IC₅₀) of an enzymatic reaction. Biochem. Pharmacol 1973, 22, 3099–3108. [DOI] [PubMed] [Google Scholar]

[R57] (57).van Bergen A; van Galen PJM; Stiles GL; Jacobson KA A₃ receptors: Structure activity relationship and molecular modeling. ACS 206th National Meeting, Chicago, IL, August, 1993, Abstract MEDI217. [Google Scholar]

[R58] (58).Lorenzen A; Guerra L; Vogt H; Schwabe U Interaction of full and partial agonists of the A₁ adenosine receptor with receptor/G protein complexes in rat brain membranes. Mol. Pharmacol 1996, 49, 915–926. [PubMed] [Google Scholar]

PERMALINK

Synthesis and Biological Activity of a New Series of N6-Arylcarbamoyl, 2-(Ar)alkynyl-N6-arylcarbamoyl, and N6-Carboxamido Derivatives of Adenosine-5′-N-ethyluronamide as A1 and A3 Adenosine Receptor Agonists

Pier Giovanni Baraldi

Barbara Cacciari

Maria José Pineda de Las Infantas

Romeo Romagnoli

Giampiero Spalluto

Rosaria Volpini

Stefano Costanzi

Sauro Vittori

Gloria Cristalli

Neli Melman

Kyung-Sun Park

Xiao-duo Ji

Kenneth A Jacobson

Abstract

Introduction

Results and Discussion

Chemistry.

Scheme 1.

Scheme 2.

Scheme 3.

Scheme 4.

Biological Activity.

Table 1.

Figure 1.

Conclusions

Experimental Section

Chemistry.

General Procedure for the Preparation of 1-Deoxy-1-[6-[[(substituted)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (4–8).

1-Deoxy-1-[6-[[(4-biphenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (4):

1-Deoxy-1-[6-[[(2,4-dichlorobenzyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (5):

1-Deoxy-1-[6-[[(4-methoxyphenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (6):

1-Deoxy-1-[6-[[(2-chlorophenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (7):

1-Deoxy-1-[6-[[(phenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (8):

General Procedure for the Preparation of 1-Deoxy-1-[6-[[[(substituted)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (14–29).

1-Deoxy-1-[6-[[[(benzyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (14):

1-Deoxy-1-[6-[[[(4-sulfonamidophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (15):

1-Deoxy-1-[6-[[[(4-acetylphenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (16):

1-Deoxy-1-[6-[[[((R)-1-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (17):

1-Deoxy-1-[6-[[[((S)-1-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (18):

1-Deoxy-1-[6-[[[(5-methyl-isoxazol-3-yl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (19):

1-Deoxy-1-[6-[[[(1,3,4-thiadiazol-2-yl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (20):

1-Deoxy-1-[6-[[[(4-n-propyloxy-phenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (21):

1-Deoxy-1-[6-[[[(2-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (22):

1-Deoxy-1-[6-[[[(3,4-dimethoxy-2-phenylethyl)amino]-carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (23):

1-Deoxy-1-[6-[[[(fur-2-yl-methyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (24):

1-Deoxy-1-[6-[[[(4-(pyridin-2-yl-aminosulfonyl)phenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (25):

1-Deoxy-1-[6-[[[(4-(5-methyl-isoxazol-3-yl-aminosulfonyl)phenyl)amino] carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuron-amide (26):

1-Deoxy-1-[6-[[[(4-(pyrimidin-2-yl-aminosulfonyl)-phenyl)amino] carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (27):

1-Deoxy-1-[6,6-[[[bis-(4-nitrophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (28):

1-Deoxy-1-[6,6-[[[bis-(5-chloro-pyridin-2-yl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (29):

General Procedure for the Preparation of 1-Deoxy-1-[6-[[(substituted)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (9–13) and 1-Deoxy-1-[6-[[[(substituted)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (30–45).

1-Deoxy-1-[6-[[(4-biphenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (9):

1-Deoxy-1-[6-[[(2,4-dichlorobenzyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (10):

1-Deoxy-1-[6-[[(4-methoxyphenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (11):

1-Deoxy-1-[6-[[(2-chlorophenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (12):

1-Deoxy-1-[6-[[(phenyl)carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (13):

1-Deoxy-1-[6-[[[(benzyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (30):

1-Deoxy-1-[6-[[[(4-sulfonamidophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (31):

1-Deoxy-1-[6-[[[(4-acetylphenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (32):

1-Deoxy-1-[6-[[[((R)-1-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (33):

1-Deoxy-1-[6-[[[((S)-1-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (34):

1-Deoxy-1-[6-[[[(5-methyl-isoxazol-3-yl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (35):

1-Deoxy-1-[6-[[[(1,3,4-thiadiazol-2-yl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (36):

1-Deoxy-1-[6-[[[(4-n-propyloxy-phenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (37):

1-Deoxy-1-[6-[[[(2-phenylethyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (38):

1-Deoxy-1-[6-[[[(3,4-dimethoxy-2-phenylethyl)amino]-carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (39):

1-Deoxy-1-[6-[[[(fur-2-yl-methyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (40):

1-Deoxy-1-[6-[[[(4-(pyridin-2-yl-aminosulfonyl)phenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (41):

1-Deoxy-1-[6-[[[(4-(5-methyl-isoxazol-3-yl-aminosulfonyl)phenyl)amino] carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (42):

Deoxy-1-[6-[[[(4-(pyrimidin-2-yl-aminosulfonyl)-phenyl)amino] carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (43):

1-Deoxy-1-[6,6-[[[bis-(4-nitrophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (44):

1-Deoxy-1-[6,6-[[[bis-(5-chloro-pyridin-2-yl)amino]-carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (45):

General Procedure for the Synthesis of 2-Chloro- and 2-Iodo-1-deoxy-1-[6-[[[(substituted)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (48–51).

2-Chloro-1-deoxy-1-[6-[[[(3-chlorophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (48):

2-Chloro-1-deoxy-1-[6-[[[(4-methoxyphenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (49):

2-Iodo-1-deoxy-1-[6-[[[(3-chlorophenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (50):

2-Iodo-1-deoxy-1-[6-[[[(4-methoxyphenyl)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-2,3-O-(1-methylethylidene)-β-d-ribofuranuronamide (51):

General Procedure for the Synthesis of 2-Chloro- and 2-Iodo-1-deoxy-1-[6-[[[(substituted)amino]carbonyl]amino]-9H-purin-9-yl]-N-ethyl-β-d-ribofuranuronamide (52–55).

Synthesis and Biological Activity of a New Series of N⁶-Arylcarbamoyl, 2-(Ar)alkynyl-N⁶-arylcarbamoyl, and N⁶-Carboxamido Derivatives of Adenosine-5′-N-ethyluronamide as A₁ and A₃ Adenosine Receptor Agonists

Binding of [³⁵S]GTP-γ-S.