Synthesis and antitumor activity of fluorine-substituted 4-amino-2(1H)-pyridinones and their nucleosides. 3-Deazacytosines

Novel fluorine-substituted deaza analogues of 5-azacytidine (AZC) and 5-aza-2'-deoxycytidine (dAZC) (3-deazacytosines) have been synthesized and tested for antitumor activity. Thus, 4-amino-3,5-difluoro-1-beta-D-ribofuranosyl-2(1H)-pyridinone (16), 4-amino-3-fluoro-1-beta-D-ribofuranosyl-2(1H)-pyridinone (17), 4-amino-5-fluoro-1-beta-D-ribofuranosyl-2(1H)-pyridinone (18), 4-amino-1-(2-deoxy-beta-D-erythro-pentofuranosyl)-3,5-difluoro-2 (1H)-pyridinone (25), 4-amino-1-(2-deoxy-beta-D-erythro-pentofuranosyl)-3-fluoro-2(1H)-pyridin one (26) 4-amino-1-(2-deoxy-alpha-D-erythro-pentofuranosyl)-3,5-difluoro-2(1H)-++ +pyridinon e (27), and 4-amino-1-(2-deoxy-alpha-D-erythro-pentofuranosyl)-3-fluoro-2 (1H)-pyridinone (28) were prepared by standard glycosylation procedures. Requisite heterocycle 4-amino-3,5-difluoro-2(1H)-pyridinone (6) was prepared in five steps from pentafluoropyridine (1). Other requisite fluoro heterocycles, 4-amino-3-fluoro-2(1H)-pyridinone (7) and 4-amino-5-fluoro-2(1H)-pyridinone (8), were obtained from a bis-defluorination of 4-amino-3,5,6-trifluoro-2(1H)-pyridinone (3) with hydrazine. Acetylation of 17 provided 4-amino-3-fluoro-1-(2,3,5-tri-O-acetyl-beta-D-ribofuranosyl)-2(1H)-pyrid inone (29). Structure proof of target nucleosides and heterocyclic compounds was provided by X-ray diffraction, 19F and 1H NMR, and UV. The ID50 values of fluorine-substituted 3-deazacytosines and 3-deazacytidines were greater than 1 X 10(-5) M in L1210 lymphoid leukemia cells in culture. Nucleoside 17 and its tri- and tetraacetates were the most active compounds with ID50 values of 1.07 X 10(-5), 1.23 X 10(-5), and 1.25 X 10(-5) M, respectively. The target nucleosides and intermediate heterocycles were inactive against P388 and L1210 lymphocytic leukemia in mice, except nucleoside 17 (NSC-378066) and its triacetate 29 (NSC-382021). Nucleoside 17 exhibited confirmed DN2 activity (% T/C 169-230) at five dose levels (25-300 mg/kg). Prodrug 29 exhibited similarly confirmed L1210 in vivo activity.     

Direct C-glycosylation of guanine analogues: the synthesis and antiviral activity of certain 7- and 9-deazaguanine C-nucleosides

C-Glycosylation of two guanine analogues, 9-deaza- and 7-deazaguanine, has been achieved under Friedel-Crafts conditions, providing a direct synthetic route to 9-deazaguanosine (4; 2-amino-7-beta-D-ribofuranosyl-5H-pyrrolo[3,2-d]pyrimidin-4(3H)-one) and 8-beta-D-ribofuranosyl-7-deazaguanine (16), respectively. This electrophilic C-glycosylation was applied successfully to six guanine and substituted-guanine analogues resulting in yields of approximately 50%. This represents the first reported C-ribosylation of preformed nitrogen heterocycles isosteric with guanine. These C-nucleosides were evaluated for their ability to provide protection against a lethal Semliki Forest virus infection in mice, relative to 7-thia-8-oxoguanosine which was used as a positive control. Two of the C-nucleosides, 2-amino-6-chloro-5-methyl-7-beta-D-ribofuranosyl-5H-pyrrolo [3,2-d]pyrimidin-4(3H)-one (12) and the corresponding 6-bromo derivative (13), showed good prophylactic activity in this virus model system.     

Microbial growth-promotion activity of 3-hydroxymonoazine- and N-hydroxydiazine-type heterocycles

Three 3-hydroxymonoazine- and three N-hydroxydiazine-type heterocycles were tested whether they act as artificial siderophores toward Aureobacterium flavescens JG-9 (ATCC No. 25091). Among them, 1-hydroxy-3,5,6-trimethyl-2(1H)-pyrazinone (3) showed the highest growth-promotion activity comparable to desferrioxamine B (DFB), a natural trihydroxamate siderophore, at 48.5 microM or above, followed by 1-hydroxy-5,6-dimethyl-2(1H)-pyrazinone (2), 1-hydroxy-4,6-dimethyl-2(1H)-pyrimidinone (1), and 3-hydroxy-2-methyl-1-phenyl-4(1H)-pyridinone (6), while 3-hydroxy-1,2-dimethyl-4(1H)-pyridinone (5) did not show the bioactivity. These results are the first examples of N-hydroxydiazine-type heterocycles acting as artificial siderophores for A. flavescens JG-9.     

Escherichia coli mediated biosynthesis and in vitro anti-HIV activity of lipophilic 6-halo-2',3'-dideoxypurine nucleosides

A series of 6-substituted 2',3'-dideoxypurine ribofuranosides (ddP) was enzymatically synthesized with live E. coli in an effort to enhance the lipophilicity of this class of anti-human immunodeficiency virus (HIV) compounds and thereby facilitate drug delivery into the central nervous system. All 6-halo-substituted ddPs were substantially more lipophilic, as defined by their octanol-water partition coefficient (P), than their nonhalogenated congeners 2',3'-dideoxyinosine (ddI) or 2',3'-dideoxyguanosine (ddG). For this class of compounds, log P's ranged from +0.5 to -1.2 in the following order: 6-iodo, 2-amino-6-iodo greater than 6-bromo, 2-amino-6-bromo greater than 6-chloro, 2-amino-6-chloro greater than 6-fluoro, 2-amino-6-fluoro much greater than ddG greater than ddI. These compounds were evaluated in vitro for ability to suppress the infectivity, replication, and cytopathic effect of HIV. 2-Amino-6-fluoro-, 2-amino-6-chloro-, and 6-fluoro-ddP exhibited a potent activity against HIV comparable to that of ddI or ddG and completely blocked the infectivity of HIV without affecting the growth of target cells. The comparative order of in vitro anti-HIV activity was 2-amino-6-fluoro, 2-amino-6-chloro, 6-fluoro greater than 2-amino-6-bromo greater than 2-amino-6-iodo, 6-chloro greater than 6-bromo greater than 6-iodo. These compounds also exhibited potent in vitro activity against HIV-2 and 3'-azido-3'-deoxythymidine-resistant HIV-1 variants. All 2-amino-6-halo-ddPs and 6-halo-ddPs were substrates for adenosine deaminase (ADA) and were converted to ddG or ddI, respectively. In the presence of the potent ADA inhibitor 2'-deoxycoformycin, 6-halo-substituted ddPs failed to exert an in vitro antiretroviral effect. These dideoxypurine nucleoside analogues represent a new class of lipophilic prodrugs of ddG and ddI that possess the potential for more effective therapy of HIV-induced neurologic disorders.     

Synthesis and antitumor activity of certain 3-beta-D-ribofuranosyl-1,2,4-triazolo[3,4-f]-1,2,4-triazines related to formycin prepared via ring closure of a 1,2,4-triazine precursor

Several 3-beta-D-ribofuranosyl-1,2,4-triazolo[3,4-f]-1,2,4-triazines related to formycin were prepared and tested for their antitumor activity in cell culture. Dehydrative coupling of 3-amino-6-hydrazino-1,2,4-triazin-5(4H)-one (5) with 3,4,6-tri-O-benzoyl-2,5-anhydro-D-allonic acid (6a) and further ring closure of the reaction product (7) provided 6-amino-3-(2,3,5-tri-O-benzoyl-beta-D-ribofuranosyl)-1,2,4-triazolo[3,4- f]-1,2,4-triazin-8(7H)-one (8). Condensation of 5 with 3,4,6-tri-O-benzoyl-2,5-anhydro-D-allonic acid chloride (6b), followed by ring annulation, also gave 8 in good yield. Debenzoylation of 8 furnished the guanosine analogue 6-amino-3-beta-D-ribofuranosyl-1,2,4-triazolo[3,4-f]-1,2,4-triazin -8(7H)-one (4b). Thiation of 8 with P2S5, followed by debenzoylation of the thiated product (11a), afforded 6-amino-3-beta-D-ribofuranosyl-1,2,4-triazolo[3,4-f]-1,2,4-triazin -8(7H)- thione (11b). Methylation of the sodium salt of 11a gave the 8-methylthio derivative (10), which on ammonolysis furnished 6,8-diamino-3-beta-D-ribofuranosyl-1,2,4-triazolo[3,4-f]-1,2,4-triazine (9). Diazotization of 10 with tert-butyl nitrite (TBN) and SbCl3 in 1,2-dichloroethane gave the corresponding 6-chloro derivative (12a). Reaction of 10 with TBN in THF in the absence of a halogen source gave 8-(methylthio)-3-(2,3,5-tri-O-benzoyl-beta-D-ribofuranosyl)-1,2,4- triazolo[3,4-f]-1,2,4-triazine (12b). Ammonolysis of 12b gave the azaformycin A analogue 8-amino-3-beta-D-ribofuranosyl-1,2,4- triazolo[3,4-f]-1,2,4-triazine (3), which on deamination afforded 3-beta-D-ribofuranosyl-1,2,4-triazolo[3,4- f]-1,2,4-triazin-8(7H)-one (4a). The azaformycin A analogue (3) showed pronounced inhibitory effects against L1210, WIL2, and CCRF-CEM cell lines with ID50 values ranging from 5.0 to 7.3 microM.     

Synthesis and antirhinovirus activity of 8-substituted analogues of 6-(dimethylamino)-9-(4-methylbenzyl)-2-(trifluoromethyl)-9H-purine.

Several 8-substituted analogues of 6-(dimethylamino) -9-(4-methylbenzyl)-2-(trifluoromethyl)-9H-purine (1) were synthesized and tested for activity against rhinovirus type 1B. Among 16 8-substituted analogues, the 8-amino (3) and 8-bromo (2) analogues were most active with IC50s of 0.36 and 1.4 microM, respectively, under conditions where 1 had an IC50 of 0.03 microM.     

Synthesis and biological activity of 6-azacadeguomycin and certain 3,4,6-trisubstituted pyrazolo[3,4-d]pyrimidine ribonucleosides

Several 3,4,6-trisubstituted pyrazolo[3,4-d]pyrimidine ribonucleosides were prepared and tested for their biological activity. High-temperature glycosylation of 3,6-dibromoallopurinol with 1-O-acetyl-2,3,5-tri-O-benzoyl-D-ribofuranose in the presence of BF3 X OEt2, followed by ammonolysis, provided 6-amino-3-bromo-1-beta-D-ribofuranosylpyrazolo-[3,4-d]pyrimidin-4(5H)-on e. Similar glycosylation of either 3-bromo-4(5H)-oxopyrazolo [3,4-d]pyrimidin-6-yl methyl sulfoxide or 6-amino-3-bromopyrazolo [3,4-d]pyrimidin-4(5H)-one, and subsequent ammonolysis, also gave 7a. The structural assignment of 7a was on the basis of spectral studies, as well as its conversion to the reported guanosine analogue 1d. Application of this glycosylation procedure to 6-(methylthio)-4(5H)-oxopyrazolo[3,4-d]pyrimidine-3-carboxamide gave the corresponding N-1 glycosyl derivative. Dethiation and debenzoylation of 16a provided an alternate route to the recently reported 3-carbamoylallopurinol ribonucleoside thus confirming the structural assignment of 16a and the nucleosides derived therefrom. Oxidation of 16a and subsequent ammonolysis afforded 6-amino-1-beta-D-ribofuranosyl-4(5H)-oxopyrazolo[3, 4-d]pyrimidine-3-carboxamide. Alkaline treatment of 15a gave 6-azacadeguomycin. Acetylation of 15a, followed by dehydration with phosgene, provided the versatile intermediate 6-amino-1-(2,3,5-tri-O-acetyl-beta-D-ribofuranosyl)-4(5H)-oxopyrazolo [3, 4-d]pyrimidine-3-carbonitrile. Deacetylation of 19 gave 6-amino-1-beta-D-ribofuranosyl-4(5H)-oxopyrazolo[3, 4-d]pyrimidine-3-carbonitrile. Reaction of 19 with H2S gave 6-amino-1-beta-D-ribofuranosyl-4(5H)-oxopyrazolo[3, 4-d]pyrimidine-3-thiocarboxamide. All of these compounds were tested in vitro against certain viruses and tumor cells. Among these compounds, the guanosine analogues 7a and 20a showed significant activity against measles in vitro and were found to exhibit moderate antitumor activity in vitro against L1210 and P388 leukemia. 6-Azacadeguomycin and all other compounds were inactive against the viruses and tumor cells tested in vitro.     

Synthesis, antibacterial, antifungal and anti-HIV evaluation of Schiff and Mannich bases of isatin derivatives with 3-amino-2-methylmercapto quinazolin-4(3H)-one

Isatin (Indole 2,3-dione), its 5-chloro and 5-bromo derivatives were added to 3-amino-2-methylmercapto quinazolin-4(3H)-one to form Schiff bases and the N-Mannich bases of these compounds were synthesized by reacting with formaldehyde and several secondary amines. Their chemical structures have been confirmed by means of their IR, 1H-NMR data and by elemental analysis. Investigation of antimicrobial activity of compounds was done by an agar dilution method against 26 pathogenic bacteria, 8 pathogenic fungi and anti-HIV activity against replication of HIV-1 (III B) in MT-4 cells. Among the compounds tested 5-chloro-3-(3',4'-dihydro-2'-methylmercapto-4'-oxoquinazolin-3'-yl )- l-morpholino methyl imino isatin was the most active antimicrobial agent.     

Guanosine analogues. Synthesis of nucleosides of certain 3-substituted 6-aminopyrazolo[3,4-d]pyrimidin-4(5H)-ones as potential immunotherapeutic agents

Several guanosine analogues were synthesized in the pyrazolo[3,4-d]pyrimidine ring system with various substituents at the 3-position. The new analogues prepared here include the CH3 (2-amino-3-methyl-1-beta-D-ribofuranosylpyrazolo[3,4-d]pyrimidin-4 (5H)-one, 13a), the phenyl (2-amino-3-phenyl-1-beta-D-ribofuranosylpyrazolo[3,4-d]pyrimidin-4 (5H)-one, 13b), and the NH2 (3,6-diamino-1-beta-D-ribofuranosylpyrazolo[3,4-d]pyrimidin-4(5H)- one, 17) substituted derivatives. These new agents, as well as several other 3-substituted derivatives including H, Br, OCH3, COOH, and oxo, were evaluated for their ability to potentiate certain murine immune functions relative to the known active agent 5-amino-3-beta-D-ribofuranosylthiazolo[4,5-d]pyrimidine-2,7(3H,6H) -dione (4, 7-thia-8-oxoguanosine). The biological evaluation included the (1) ex vivo determination of increased natural killer cell function and (2) in vivo antiviral protection against a lethal challenge of Semliki Forest virus. The 3-unsubstituted (5a) and the 3-bromo (5c) derivatives were found to be the most active immunopotentiators in this series.     

Synthesis, antibacterial, antifungal and anti-HIV evaluation of Schiff and Mannich bases of isatin and its derivatives with triazole

Isatin (indole 2,3-dione) and its 5-chloro and 5-bromo derivatives have been reacted with 3-(4'-pyridyl)-4-amino-5-mercapto-4-(H)-1,2,4-triazole to form Schiff bases and the N-Mannich bases of these compounds were synthesised by reacting them with formaldehyde and several secondary amines. Their chemical structures have been confirmed by means of their IR, 1H-NMR data and by elemental analysis. Investigation of antimicrobial activity of compounds was done by agar dilution method against 27 pathogenic bacteria, 8 pathogenic fungi and anti-HIV activity against replication of HIV-1 (III B) in MT-4 cells. Among the compounds tested 1-(piperidinomethyl) 5-bromo 3-[3'-(4"-pyridyl)-5'-mercapto-4'-(H)-1',2',4'-triazol 4'-yl]imino isatin showed the most favourable antimicrobial activity.     

Synthesis and antiviral activity of 9-[4-hydroxy-3-(hydroxymethyl)but-1-yl]purines

Alkylation of 2-amino-6-chloropurine with 5-(2-bromoethyl)-2,2-dimethyl-1,3-dioxane (5) provided 2-amino-6-chloro-9-[2,(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine (6) in high yield. This aminochloropurine 6 was readily converted to the antiviral acyclonucleoside 9-[4-hydroxy-3-(hydroxymethyl)but-1-yl]guanine (1) and to its 6-chloro (10), 6-thio (11), 6-alkoxy (12-17), 6-amino (20), and 6-deoxy (21) purine analogues. The guanine derivative 1 was converted to its xanthine analogue 9. Similarly, alkylation of 6-chloropurine with 5 provided a route to 8, the hypoxanthine analogue of 1. Of these 9-substituted purines, the guanine derivative 1 showed the highest activity against herpes simplex virus types 1 and 2 in cell cultures, and in some tests it was more active than acyclovir, with no evidence of toxicity for the cells. A series of monoesters (30-33) and diesters (24-27, 29) of 1 were prepared, and some of these also showed antiherpes virus activity in cell cultures, the most active ester being the dihexanoate 27.     

Synthesis and biological evaluation of certain 3-beta-D-ribofuranosyl-1,2,4-triazolo[4,3-b)pyridazines related to formycin prepared via ring closure of pyridazine precursors

All three amino-substituted 3-beta-D-ribofuranosyl-1,2,4-triazolo[4,3-b]pyridazines (5, 19, and 20) structurally related to formycin A were prepared and tested for their antitumor and antiviral activity in cell culture. Dehydrative coupling of 4-amino-5-chloro-3-hydrazinopyridazine (7) with 3,4,6-tri-O-benzoyl-2,5-anhydro-D-allonic acid (6) in the presence of DCC and subsequent thermal ring closure of the reaction product (8) provided 8-amino-7-chloro-3-(2,3,5-tri-O-benzoyl-beta-D-ribofuranosyl)- triazolo[4,3-b]pyridazine (9). Dehalogenation of 9, followed by debenzoylation, gave the formycin congener 8-amino-3-beta-D-ribofuranosyl-1,2,4- triazolo[4,3-b]pyridazine (5). Similar condensation of 5-amino-4-chloro-3-hydrazinopyridazine (13) with 6 and dehalogenation of the cyclized product (16), followed by debenzoylation, gave the isomeric 7-amino-3-beta-D-ribofuranosyl-1,2,4- triazolo[4,3-b]pyridazine (19). DCC-mediated coupling of 6 with 6-chloro-3-hydrazinopyridazine (12), followed by ammonolysis of the cyclized product (21) with liquid NH3, provided a convenient route to 6-amino-3-beta-D-ribofuranosyl-1,2,4-triazolo[4,3-b]pyridazine (20). The structural assignment of 5 was made by single-crystal X-ray diffraction analysis. Compounds 5, 19, 20, and certain deprotected nucleoside intermediates were evaluated against L1210, WI-L2, and CCRF-CEM tumor cell lines, as well as against DNA and RNA viruses in culture. These compounds did not exhibit any significant antitumor or antiviral activity in vitro.     

Adenosine receptor agonists: synthesis and biological evaluation of 1-deaza analogues of adenosine derivatives

In a search for more selective A1 adenosine receptor agonists, N6-[(R)-(-)-1-methyl-2-phenethyl]-1-deazaadenosine (1-deaza-R-PIA, 3a), N6-cyclopentyl-1-deazaadenosine (1-deazaCPA, 3b), N6-cyclohexyl-1-deazaadenosine (1-deazaCHA, 3c), and the corresponding 2-chloro derivatives 2a-c were synthesized from 5,7-dichloro-3-beta-D-ribofuranosyl-3H-imidazo[4,5-b]pyridine. On the other hand, N-ethyl-1'-deoxy-1'-(1-deaza-6-amino-9H-purin-9-yl)-beta-D-ribofuranu ronamide (1-deazaNECA, 10) was prepared from 7-nitro-3-beta-D-ribofuranosyl-3H-imidazo[4,5-b]pyridine, in an attempt to find a more selective A2 agonist. The activity of all deaza analogues at adenosine receptors has been determined in adenylate cyclase and in radioligand binding studies. 1-DeazaNECA proved to be a nonselective agonist at both subtypes of the adenosine receptor. It is about 10-fold less active than NECA but clearly more active than the parent compound 1-deazaadenosine as an inhibitor of platelet aggregation and as a stimulator of cyclic AMP accumulation. The N6-substituted 1-deazaadenosines largely retain the A1 agonist activity of their parent compounds, but lose some of their A2 agonist activity. This results in A1-selective compounds, of which N6-cyclopentyl-2-chloro-1-deazaadenosine (1-deaza-2-Cl-CPA, 2b) was identified as the most selective agonist at A1 adenosine receptors so far known. The activity of all 1-deaza analogues confirms that the presence of the nitrogen atom at position 1 of the purine ring is not critical for A1 receptor mediated adenosine actions.     

Paullones, a series of cyclin-dependent kinase inhibitors: synthesis, evaluation of CDK1/cyclin B inhibition, and in vitro antitumor activity.

The paullones represent a novel class of small molecule cyclin-dependent kinase (CDK) inhibitors. To investigate structure-activity relationships and to develop paullones with antitumor activity, derivatives of the lead structure kenpaullone (9-bromo-7,12-dihydroindolo[3,2-d][1]benzazepin-6(5H)-one, 4a) were synthesized. Paullones with different substituents in the 2-, 3-, 4-, 9-, and 11-positions were prepared by a Fischer indole reaction starting from 1H-[1]benzazepine-2,5(3H,4H)-diones 5. Selective substitutions at either the lactam or the indole nitrogen atom were accomplished by treating kenpaullone with alkyl halides in the presence of sodium hydride/THF or potassium hydroxide/acetone, respectively. S-Methylation of the kenpaullone-derived thiolactam 18 yielded the methylthioimidate 19, which gave the hydroxyamidine 20 upon reaction with hydroxylamine. The new paullones were tested both in a CDK1/cyclin B inhibition assay and in the in vitro antitumor cell line-screening program of the National Cancer Institute (NCI). With respect to the CDK1/cyclin B inhibition, electron-withdrawing substituents in the 9-position as well as a 2,3-dimethoxy substitution on the paullone basic scaffold turned out to be favorable. A 9-trifluoromethyl substituent was found to be equivalent to the 9-bromo substituent of kenpaullone. Replacement of the 9-bromo substituent of kenpaullone by a 9-cyano or 9-nitro group produced a substantial increase in enzyme-inhibiting potency. Substitutions in other positions or the replacement of the lactam moiety led to decreased CDK1 inhibition. Noteworthy in vitro antitumor activities (GI(50) values between 1 and 10 microM) were found with the 9-bromo-2,3-dimethoxy-7,12-dihydroindolo[3, 2-d][1]benzazepin-6(5H)-one (4t), its 9-trifluoromethyl analogue 4u, the 12-Boc-substituted paullone15, and the methylthioimidate 19, respectively. The 9-nitro-7,12-dihydroindolo[3, 2-d][1]benzazepin-6(5H)-one (4j, named alsterpaullone) showed a high CDK1/cyclin B inhibitory activity (IC(50) = 0.035 microM) and exceeded the in vitro antitumor potency of the other paullones by 1 order of magnitude (log GI(50) mean graph midpoint = -6.4 M).     

Synthesis, antiproliferative, and antiviral activity of certain 4-aminopyrrolo[2,3-d]pyridazine nucleosides: an entry into a novel series of adenosine analogues.

The preparation of novel heterocyclic base modified adenosine analogues, the 4-aminopyrrolo[2,3-d]pyridazine nucleosides, is described. Crucial to their successful preparation was the use of the pyrrole glycoside intermediates 3-cyano-2-formyl-1-(2,3,5-tri-O-benzyl-beta-D-ribofuranosyl)pyrrole (11) and 3-cyano-2-formyl-1-(2,3,5-tri-O-benzyl-beta-D-arabinofuranosyl)pyrrole (17). Treatment of 11 and 17 with hydrazine dihydrochloride followed by treatment with boron trichloride provided 4-amino-1-beta-D-ribofuranosylpyrrolo[2,3-d]pyridazine (2) and 4-amino-1-beta-D-arabinofuranosylpyrrolo[2,3-d]pyridazine (3), respectively. 4-Amino-3-bromo-1-beta-D-ribofuranosylpyrrolo[2,3-d]-pyridazine (4) was prepared by a bromination of 4-amino-1-(2,3,5-tri-O-benzyl-beta-D-ribofuranosyl)pyrrolo[2,3-d]pyri daz ine (12) and subsequent removal of the benzyl groups with boron trichloride. Compounds 2-4 were evaluated for antiproliferative and antiviral activity. The tubercidin analogue (2) and its arabinosyl derivative (3) were virtually inactive in all assays. In contrast, the 3-bromo analogue 4 inhibited growth of L1210 and H. Ep. 2 cells. Compound 4 was also active against human cytomegalovirus and herpes simplex virus type 1, but the antiviral activity was not completely separated from cytotoxicity.     

Structural determinants for AMPA agonist activity of aryl or heteroaryl substituted AMPA analogues. Synthesis and pharmacology

We have previously reported the synthesis and pharmacological characterization of analogues of 2-amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic acid (AMPA, 1a), in which the methyl group was replaced by a phenyl group (APPA, 1b) or heteroaryl groups. While 2b and its 3-pyridyl analogue 2-amino-3-[3-hydroxy-5-(3-pyridyl)-4-isoxazolyl]propionic acid (3-Py-AMPA, 3) show very low affinity for AMPA receptors, introduction of heteroaryl substituents containing heteroatom in the 2-position provides potent AMPA receptor agonists. We here report the synthesis and pharmacology of 2-amino-3-(3-hydroxy-5-pyrazinyl-4-isoxazolyl)propionic acid (7) (IC50 = 1.2 microM), which is weaker as an AMPA agonist than AMPA (IC50 = 0.040 microM; EC50 = 3.5 microM) but comparable in potency with 2-Py-AMPA (4) (IC50 = 0.57 microM; EC50 = 7.4 microM), as determined in radioligand binding and electrophysiological experiments, respectively. The AMPA analogues 8a-c, containing 2-, 3-, or 4-methoxyphenyl substituents, respectively, and the corresponding hydroxyphenyl analogues, 9a-c, were also synthesized and evaluated pharmacologically. With the exception of 2-amino-3-[3-hydroxy-5-(2-hydroxyphenyl)-4-isoxazolyl]propionic acid (9a), which is a very weak AMPA agonist (IC50 = 45 microM; EC50 = 324 microM), none of these compounds showed detectable effect at AMPA receptors.     

Synthesis and antiviral activity of certain 4- and 4,5-disubstituted 7-[(2-hydroxyethoxy)methyl]pyrrolo[2,3-d]pyrimidines

In vitro evaluation of a series of previously prepared tubercidin analogues revealed that certain 5-halogen-substituted analogues were active against human cytomegalovirus (HCMV) at concentrations lower than those that produced comparable cytotoxicity in uninfected cells. In contrast, tubercidin was cytotoxic at all antiviral concentrations. Even though the antiviral selectivity of the 5-substituted compounds was slight, this observation led us to prepare a series of acyclic analogues. Treatment of the sodium salt of 4-chloropyrrolo[2,3-d]pyrimidine (2) with (2-acetoxyethoxy)methyl bromide (2a) provided the acyclic nucleoside 4-chloro-7-[(2-acetoxyethoxy)methyl]pyrrolo[2,3-d]pyrimidine (3). A nucleophilic displacement of the 4-chloro group with methoxide, methylamine, and dimethylamine yielded the corresponding 4-substituted compounds 4, 5, and 6, respectively, in good yield. Electrophilic substitution (chlorination, bromination, and iodination) was effected at the C-5 position of compound 3 with N-chlorosuccinimide, N-bromosuccinimide, and iodine monochloride, respectively, in methylene chloride. Removal of the acetyl group from these intermediates (7a-9a) with methanolic ammonia at room temperature afforded the 5-chloro (7b), 5-bromo (8b), and 5-iodo (9b) derivatives of 4-chloro-7-[(2-hydroxyethoxy)methyl]pyrrolo[2,3-d]pyrimidine. Treatment of compounds 7b-9b with methanolic ammonia at an elevated temperature produced the corresponding 5-halotubercidin analogues 10, 11, and 12, respectively. An alternate procedure for the preparation of these 4,5-disubstituted 7-[(2-hydroxyethoxy)methyl]pyrrolo[2,3-d]pyrimidines involved an electrophilic substitution prior to the condensation of the heterocycle with 2a. Treatment of 2 with N-chlorosuccinimide and N-bromosuccinimide gave compounds 13a and 13b, respectively. The condensation of 13a and 13b with 2a and subsequent treatment with methylamine and ethylamine furnished the corresponding 5-halo-4-substituted-pyrrolo[2,3-d]pyrimidines 14a, 14b, 14c, and 14d, respectively. Evaluation of the target compounds (4-6, 7b-9b, 10-12, and 14a-14d) for cytotoxicity and activity against HCMV and herpes simplex virus type 1 (HSV-1) revealed that all compounds except the 5-halogen-substituted compounds 10, 11, and 12 were inactive. Compounds 10, 11, and 12 were active against both viruses at noncytotoxic concentrations. The activity of compound 11 was particularly noteworthy, being at least 10-fold more potent than acyclovir.     

Studies on beta-lactam antibiotics. I. Synthesis and in vitro anti-pseudomonal activity of 3-isothiazole-cephalosporin derivatives

The synthesis and in vitro activity of 7 beta-[(Z)-2-(2-amino-4-thiazolyl)-2-(2-carboxy-2- alkoxyimino)acetamido]cephalosporins with a (4-carboxy-3-hydroxy-5-isothiazolyl)thiomethyl group at the 3-position are described. These cephalosporins (9a approximately 9i) showed excellent activity against Gram-negative bacteria including beta-lactamase producing strains. The most interesting compound of the series was 7 beta-[(Z)-2-(2-amino-4-thiazolyl)-2- (2-carboxy-2-propoxyimino)acetamido]-3-cephem-4-carboxylic acid (9g, YM-13115) because of its outstanding inhibitory potency against Pseudomonas aeruginosa and highly prolonged plasma half-life in rats.     

9-Benzyl-6-(dimethylamino)-9H-purines with antirhinovirus activity

A series of 9-benzyl-6-(dimethylamino)-9H-purines and 9-benzyl-2-chloro-6-(dimethylamino)-9H-purines were synthesized and tested in cell culture for activity against rhinovirus type 1B. The 9-benzylpurines that were unsubstituted in the 2-position had weak activity. However, introduction of a 2-chloro substituent resulted in a substantial increase in antiviral activity. One of the most active compounds, 2-chloro-6-(dimethylamino)-9-(4-methylbenzyl)-9H-purine (29), had an IC50 value of 0.08 microM against serotype 1B. Four compounds were tested against 18 other rhinovirus serotypes, but the majority tested were less sensitive than type 1B. The range of serotype sensitivity for 29 varied from 0.08 to 14 microM. These 9-benzyl-2-chloro-9H-purines represent a new class of antiviral agents with in vitro activity against rhinoviruses.     

Selective antagonists at group I metabotropic glutamate receptors: synthesis and molecular pharmacology of 4-aryl-3-isoxazolol amino acids

Homologation of (S)-glutamic acid (Glu, 1) and Glu analogues has previously provided ligands with activity at metabotropic Glu receptors (mGluRs). The homologue of ibotenic acid (7), 2-amino-3-(3-hydroxy-5-isoxazolyl)propionic acid (HIBO, 8), and the 4-phenyl derivative of 8, compound 9a, are both antagonists at group I mGluRs. Here we report the synthesis and molecular pharmacology of HIBO analogues 9b-h containing different 4-aryl substituents. All of these compounds possess antagonist activity at group I mGluRs but are inactive at group II and III mGluRs.