Dose-dependent pharmacokinetics of 1-(2-deoxy-beta-D- ribofuranosyl)-2,4-difluoro-5-iodobenzene: a potential mimic of 5-iodo-2'-deoxyuridine

The dose-range pharmacokinetics of l-(2-deoxy-beta-D-ribofuranosyl)-2,4-difluoro-5-iodobenzene (5-IDFPdR), a C-aryl nucleoside mimic of IUdR, were studied in male Sprague-Dawley rats following single intravenous (i.v.) and oral doses. After i.v. administration, the blood clearance decreased from approximately 32 ml/min/kg at a dose of 15 mg/kg, to approximately 19 ml/min/kg when dosed at 54 mg/kg, and the elimination half-life increased from 8.4 min to 21.5 min, for the respective doses. While the dose-normalized area under the concentration-time curve (AUCnorm) remained practically unchanged (0.132 kg min ml(-1)) upon increasing the i.v. dose from 5 to 15 mg/kg, it increased by about 44% ( approximately 0.19 kg min ml(-1)) when the i.v. dose was increased from 15 to 54 mg/kg. Similarly, there was a dose-dependent increase in AUCnorm with increasing oral doses: AUCnorm increased by 49% as the oral dose increased from 20 to 40 mg/kg, and further by 55% as the oral dose was increased from 40 mg/kg to 54 mg/kg. For the respective oral doses, the elimination half-life increased from 24.5 min to 176 min, while blood clearance was reduced from approximately 37 ml/min/kg to approximately 17 ml/min/kg. The urinary recoveries of unchanged 5-IDFPdR and its glucuronides (as percent of the dose) were somewhat increased at higher doses. This increase was more pronounced following the highest oral dose. The total biliary recovery of 5-IDFPdR (as percent of the dose) was, however, decreased with increasing doses. The overall kinetic profile of 5-IDFPdR based on these data is suggestive of dose-dependent pharmacokinetics. Decreased elimination of 5-IDFPdR with increasing dose, as supported by longer elimination half-lives at higher doses, is one likely mechanism contributing to the dose-dependent behaviour of this compound. Saturable non-renal metabolism might explain the reduced total body clearance of 5-IDFPdR at higher doses, despite the unchanged or increased urinary clearance.For drugs exhibiting nonlinear kinetics, the dosage regimens may need to be carefully designed to avoid potential unpredictable toxicity and/or lack of pharmacological response associated with the disproportional changes in steady state drug concentrations on changing dose. Manifestation in the rat of nonlinear kinetics at doses of 5-IDFPdR, which may be of therapeutic relevance, warrants extended dose-range evaluations of this compound in future preclinical and clinical studies, to establish safe and efficacious dosage regimens.     

Metabolism of anti-herpes agent 5-(2-chloroethyl)-2'-deoxyuridine in mice and rats

The enzymatic splitting and metabolic elimination of anti-viral agent 5-(2-chloroethyl)-2'-deoxyuridine [CEDU] have been studied. For elucidation of structures of metabolites, several different kinds of extraction, purification and spectroscopic methods were used (Extrelut LC, TLC, HPLC, MS, NMR, IR, UV and CD). For mass spectral analysis, various ionization techniques (EI, CI and FAB-MS) were performed as complementary methods. After oral administration of [14C]-CEDU to mice and rats, the parent compound, 5-(2chloroethyl) uracil [CEU] and hydroxylated CEU metabolites were isolated and identified from urine and faeces by the above mentioned methods. The CEDU showed rapid phosphorolysis in vitro with thymidine phosphorylase Km 41.0 +/- 5.0; and uridine phosphorylase Km 10.0 +/- 1.5. The cleavage of the N-glycosidic bond of the nucleoside analogue and a new metabolic pathway of CEDU [stereoselective oxidation of 5-(2-chloroethyl) uracil] was observed in both species.     

Inhibition of nucleoside phosphorylase cleavage of 5-fluoro-2'-deoxyuridine by 2,4- pyrimidinedione derivatives

Several 2,4-pyrimidinedione (uracil) derivatives were evaluated as inhibitors of the pyrimidine nucleoside phosphorylases that cleave 5-fluorn-2'-deoxyuridine (FUdR) to 5-fluorouracil. Pyrimidinediones substituted at either N-1 or C-5, or both, markedly inhibited the phosphorolysis of FUdR by the uridine-deoxyuridine phosphorylases of Ehrlich ascites and Novikoff hepatoma cells. The most potent inhibitors were 5-benzyluracil derivatives substituted with alkoxy groups on the meta-position of the benzyl moiety; the most active of these was 5-{[3-(phenylmethoxy)phenyl]methyl}uracil. The same derivatives, however, did not inhibit the phosphorolysis of FUdR by the thymidine phosphorylases of murine liver, human leukocytes and HeLa (S3) cells. 6-Anilino and 6-(1-naphthylmethylamino) derivatives of uracil, which have been shown by others to inhibit the cleavage of FUdR by the thymidine phosphorylase activity of Escherichia coli, did not inhibit any of the mammalian thymidine or uridinedeoxyuridine phosphorylase activities. By contrast, pyrimidinediones substituted with smaller, non-hydrophobic groups at either C-5 or C-6, or both, inhibited the cleavage of FUdR by both the mammalian thymidine and uridine-deoxyuridine phosphorylases. The most active of these, 6-aminothymine, was also the best inhibitor of thymidine phosphorylase. Our results demonstrate differences in the active sites of the various pyrimidine nucleoside phosphorylases, and should provide a basis for the design of more potent and specific inhibitors of the nucleoside phosphorylase(s) responsible for the cleavage of FUdR in man.     

Phosphorolysis of (E)-5-(2-bromovinyl)-2'-deoxyuridine (BVDU) and other 5-substituted-2'-deoxyuridines by purified human thymidine phosphorylase and intact blood platelets

Various 5-substituted-2'-deoxyuriclines (dUrd), inclucling 5-ethyl-,5-propyl-, 5-trifluoromethyl-, 5-hydroxymethyl-, 5-formyl-, 5-vinyl-, (E)-5-(2-chlorovinyl)-, (E)-5-(2-bromovinyl)-, 5-fluoro-. 5-chloro-. 5-bromo-. 5-iodo-, 5-cyano-, 5-thiocyano-, 5-nitro- and 5-amino-dUrd. were shown to be effective substrates for the thymidine (dThd) phosphorylase isolated from human blood platelets. Some of dUrd analogs, i.e. the highly potent and selective antiherpes agent (E)-5-(2-bromovinyl)-dUrd. were degraded more rapidly than the natural substrates, dUrd and dThd. All dUrd analogs were also readily catabolised by intact human blood platelets. The potent inhibitors of thymicline phosphorylase, 6-amino-thymine and 6-amino-5-bromo-uracil, strongly inhibited the phosphorolysis of (E)-5-(2-bromovinyl)-dUrd by both purified enzyme and intact platelets.     

Structure-activity relationships of 1-alkyl-5-(3,4-dichlorophenyl)- 5-[2-[(3-substituted)-1-azetidinyl]ethyl]-2-piperidones. 1. Selective antagonists of the neurokinin-2 receptor

The design, synthesis, and pharmacological evaluation of a novel class of neurokinin-2 (NK2) antagonists 1-alkyl-5-(3,4-dichlorophenyl)-5-[2-[(3-substituted)-1-azetidinyl]ethyl]-2-piperidones (5-44) are described. These compounds are formally derived from 2 by incorporating the metabolically vulnerable N-methylamide function into a more stable six-membered ring lactam 4, resulting in increased stability in human liver microsome (HLM) preparations relative to 2 (T1/2(HLM) of 30 min vs <10 min for 2). This series was further optimized by replacing the 4,4-disubstituted piperidine functionality found in 4 with simple 3-substituted azetidines. This series, exemplified by 1-benzyl-5-(3,4-dichlorophenyl)-5-[2-[3-(4-morpholinyl)-1-azetidinyl]ethyl]-2-piperidone 5, was found to possess excellent functional potency for the NK2 receptor in the Rabbit pulmonary artery (RPA) assay (pA2 = 9.3) and increased in vitro metabolic stability (T1/2(HLM) = 70 min) relative to 4. Metabolic route identification studies revealed that N-benzyl oxidation was a major route in this relatively lipophilic lead (log D = 3.2). Further exploration of the N-lactam substituent SAR targeting reduced lipophilicity to attenuate P-450 metabolism revealed that incorporation of a cyclopropylmethyl group in this region of the molecule gave a balance of good potency and high metabolic stability. For example, the significantly less lipophilic analogue 29 (log D = 2.3) possessed both good functional potency (RPA, pA2 = 8.1) and high in vitro metabolic stability (T1/2(HLM) = 120 min). Optimization in this N-cyclopropylmethyllactam series by modification of the nature of the azetidine 3-substituent as a strategy to further increase potency and moderate log D led to the identification of sulfamide analogue 33, which possessed both excellent metabolic stability in vitro (T1/2(HLM) >120 min) and high potency in both RPA (pA2 = 8.9) and human bladder smooth muscle (pK(b) = 8.9) functional assays. In addition, NK2 antagonist 33 (IC50 = 4 nM) showed excellent selectivity over both the related human neurokinin receptors h-NK1 (IC50 = 7.9 microM) and h-NK3 (IC50 = 1.8 microM) in radioligand binding studies.     

Biochemical and pharmacokinetic evaluation of a novel pyrimidine nucleoside nitric oxide donor as a potential anticancer/antiviral agent.

The objective of this study was to determine the physiochemical, biochemical and pharmacokinetic properties of 5-iodo-3'-O-nitro-2'-deoxyuridine (INUdR), a novel prodrug releasing both nitric oxide (NO) and 5-iodo-2'-deoxyuridine. The INUdR partition coefficient (log P=1.12) was determined by both the shake-flask method and by calculation using Interactive Analysis Log P Program. In vitro binding of INUdR to bovine serum albumin (BSA) was estimated using an ultrafiltration method to be 65 to 77%, depending on the INUdR concentration. INUdR was stable in phosphate buffer (pH 7.4) and in water, at both ambient temperature and at 37 degrees C. INUdR was resistant to phosphorolysis when incubated with thymidine phosphorylase. Plasma, L-cysteine and glutathione catalyzed release of NO from INUdR, as determined using the Griess reaction. In all three systems, the release of NO by INUdR was equal to or greater than that of the reference drug isosorbide dinitrate. The pharmacokinetics of INUdR following single intravenous bolus and oral doses of INUdR (40 mg/kg) to male Sprague-Dawley rats were characterized by a short elimination half-life (T(1/2) 0.27 h), a large steady-state volume of distribution (V(ss) 0.89 l/kg) and high oral bioavailability (F=0.95). In conclusion, INUdR lipophilicity, shelf-stability, and resistance towards catabolic breakdown by thymidine phosphorylase, together with its non-spontaneous, yet considerable NO release, constitute favorable characteristics of a potential anticancer/antiviral agent.     

The metabolism of the anti-tumour agent 5-(1-aziridinyl)-2,4-dinitrobenzamide (CB 1954)

3he metabolism in the rat of 5-(1-aziridinyl)-2,4-dinitrobenzamide (CB 1954), an antitumour agent having a highly specific action on the Walker 256 carcinoma in rats, has been studied. Unchanged CB 1954 is the most abundant radioactive constituent in the urine of rats treated with the tritium-labelled drug. The principal urinary metabolite, 4-amino-5-(1-aziridinyl)-2-nitrobenzamide, was also tumour inhibitory, though less so than CB 1954. The isomeric 2-amino-4-nitro derivative, which was not a urinary metabolite was much less active. Other urinary metabolites identified were 5-amino-2,4-dinitrobenzamide and 5-(2-hydroxyethyl)amino-2,4-dinitrobenzamide.The results of this metabolic study are considered in relation to possible explanations for the specificity of CB 1954 towards the Walker tumour, and are compared with the earlier findings for the parent member of the series, 1-(1-aziridinyl)-2,4-dinitrobenzene (CB 1837).     

Phosphorolysis of 5′-deoxy-5-fluorouridine in human plasma,serum and blood platelets

During the course of the study of 5′-deoxy-5-fluorouridine (5′dFUrd) plasma protein binding using ¹⁹F NMR spectroscopy, phosphorolytic cleavage of 5′dFUrd into 5-fluorouracil (5FU) was observed. This transformation was due to the enzymatic content of residual blood cells present in plasma, since the percentage conversion of 5′dFUrd into 5FU was lower as the number of residual blood cells fell, and an ‘acellular’ plasma or a serum effected a negligible phosphorolyis of 5′dFUrd. As platelets were the contaminants of the plasma samples studied, and a concentrate of human platelets demonstrated a high phosphorolytic activity towards 5′dFUrd, it was concluded that these blood cells were responsible for the 5′dFUrd cleavage. Thymidine phosphorylase, being the only pyrimidine nucleoside phosphorylase in human platelets, is suggested by the present results to catalyse 5′dFUrd phosphorolysis and is therefore not as specific for 2′-deoxyribonucleosides as has been reported.     

Synthesis and biological activity of 5-(2,2-difluorocyclopropyl)-2'-deoxyuridine deoxyuridine diastereomers.

The synthesis of the two diastereomers (9 and 10) of 5-(2,2-difluorocyclopropyl)-2'-deoxyuridine are described. Their antiviral and cytotoxic activities were determined, in comparison with (E)-5-(2-bromovinyl)-2'-deoxyuridine (BVDU) and 5-fluoro-2'-deoxyuridine (FDU), respectively. 5-[(1R)-2,2-Difluorocyclopropyl]-2'-deoxyuridine (10) was the most active antiviral agent against HSV-1 (IC50 = 5 micrograms/ml) relative to BVDU (IC50 = 0.082 micrograms/ml), and cytotoxic agent in the CCRF-CEM (IC50 = 230 microM) screen relative to FDU (IC50 = 4.7 x 10(-3) microM). The 5-[(1S)-2,2-difluorocyclopropyl] diastereomer was inactive in both screens. Partition coefficients (P) and affinity for the mouse erythrocyte nucleoside transporter (Ki) were not determinants of antiviral or cytotoxic activities. However, the (1R)-diastereomer (10) was more resistant to glycosidic bond cleavage by thymidine phosphorylase than the (1S)-diastereomer (9).     

4-[5-Fluoro-3-[4-(2-methyl-1H-imidazol-1-yl)benzyloxy]phenyl]-3,4,5,6- tetrahydro-2H-pyran-4-carboxamide, an orally active inhibitor of 5-lipoxygenase with improved pharmacokinetic and toxicology characteristics

Described herein are structure-activity relationships (SARs) of 4-[5-fluoro-3-[4-(2-methyl-1H-imidazol-1-yl)benzyloxy]-phenyl]-4-methoxy-3,4,5,6-tetrahydro-2H-pyran (1, CJ-12,918), an imidazole 5-lipoxygenase (5-LO) inhibitor. When 1 was tested in preclinical studies, cataract formation was observed in rats; however, this compound was metabolized extensively in vivo and showed low systemic exposure. To eliminate this side effect and enhance bioavailability, structural modification was focused on replacing the methoxy group of 1 by modulating lipophilicity (i.e., predicted log D at pH 7.4). The SARs led to the discovery of 4-[5-fluoro-3-[4-(2-methyl-1H-imidazol-1-yl)benzyloxy]phenyl]-3,4,5,6-tetrahydro-2H-pyran-4-carboxamide (10, CJ-13,454), which was less lipophilic by 1.2 log D units and showed in vivo potency (ED(50) = 4-9 mg/kg) equipotent to 1. Enhanced metabolic stability resulted in fewer in vivo metabolites, as well as improved bioavailability and a better toxicological profile. Thus, 10 was found to be a more practical lead for an orally active 5-LO inhibitor.     

Intracellular metabolism of the new antiviral compound 1-(S)-[3-hydroxy-2-(phosphonomethoxy)propyl]-5-azacytosine

1-(S)-[3-hydroxy-2-(phosphonomethoxy)propyl]-5-azacytosine [HPMP-5-azaC], the 5-azacytosine analogue of cidofovir (HPMPC), represents a new acyclic nucleoside phosphonate with pronounced activity against DNA viruses, and a selectivity index superior to that of cidofovir. Here we investigated the intracellular metabolic pathway of [6-³H]-HPMP-5-azaC. By comparing the metabolism in mouse lymphosarcoma S49-wild type (S49-WT) and mutant cells deficient for dCMP deaminase, we identified the mono- and diphosphate metabolites generated from HPMP-5-azaC and its deaminated product HPMP-5-azaU. In human lung carcinoma A549 cells, the relative formation of the deaminated metabolites was only 6%, implying that deamination plays a minor role in the overall metabolism of HPMP-5-azaC. The diphosphorylated metabolite of HPMP-5-azaC accounted for 60% of the total radioactivity, and reached intracellular levels which were 60-fold higher in absolute value than the corresponding diphosphate levels obtained with cidofovir. Consequently to its increased activation, HPMP-5-azaC showed about 45-fold higher incorporation into cellular DNA than cidofovir. Herpes-, pox- or adenovirus infection had no marked influence on the metabolism of HPMP-5-azaC. The HPMP-5-azaC-diphosphate metabolite was shown to have long intracellular stability (half-life: 63 h), suggesting that infrequent administration of HPMP-5-azaC should be possible. HPMP-5-azaC represents a new acyclic nucleoside phosphonate compound with promising anti-DNA virus activity and a favorable metabolic profile that is characterized by low sensitivity to catabolic deamination and a high rate of phosphorylation and DNA incorporation.     

Metabolic and photochemical hydroxylation of 5-nitro-2-furancarboxaldehyde derivatives.

The potassium salt of 1-[[(5-aco-nitro-4,5-dihydro-4-oxo-2-furanyl)methylene]amino]-2,4-imidazolidinedione (4) was isolated from the urine of rats fed nitrofurantoin. An aldehyde absorbing at 400 nm was synthesized photochemically, in less than 1% yield, from 5-nitro-2-furancarboxaldehyde diacetate (1), and the hydroxylamine (2), 3-amino-2-oxazolidinone (3a-c), and 1-amino-2,4-imidazolidinedione (4) derivatives were prepared. On the basis of ir and NMR data 2, 3b,c, and 4 are considered derivatives of 4-hydroxy-5-nitro-2-furancarboxaldehyde which are mainly in the aci-nitro form. Methyl and ethyl nitronic esters of 3b were synthesized. The photochemical hydroxylation of 1 also yields 3,4-dihydroxy-5-nitro-2-furancarboxaldehyde, isolated as 3-[[(3,4-dimethoxy-5-nitro-2-furanyl)-methylene[amino[-2-oxazolidinone (7).     

Structure-activity studies of 5-[[4-(4,5-dihydro-2-oxazolyl) phenoxy]alkyl]-3-methylisoxazoles: inhibitors of picornavirus uncoating

A series of substituted phenyl analogues of 5-[[4-(4,5-dihydro-2-oxazolyl) phenoxy]alkyl]-3-methylisoxazoles has been synthesized and evaluated in vitro against several human rhinovirus (HRV) serotypes. Substituents in the 2-position greatly enhanced activity when compared to the unsubstituted compound. Many of these compounds exhibited mean MICs (MIC) against five serotypes as low as 0.40 microM. The mean MIC correlated well (r = 0.83) with the MIC80 (the concentration that inhibited 80% of the serotypes tested). A quantitative structure-activity relationship study indicated a strong dependency of MIC on lipophilicity (log P) in combination with inductive effects (sigma m) and bulk factors (MW).     

5-Substituted-2,4-diamino-6-[2-(phosphonomethoxy)ethoxy]pyrimidines-acyclic nucleoside phosphonate analogues with antiviral activity

2,4-Diamino-6-hydroxypyrimidines substituted in position 5 by an allyl, benzyl, cyanomethyl, ethoxycarbonylmethyl, phenyl, cyclopropyl, or methyl group were prepared either by C5-alkylation or by formation of the pyrimidine ring by cyclization. Their alkylation with 2-[(diisopropoxyphosphoryl)methoxy]ethyl tosylate afforded N1- and O6-regioisomers that were separated and converted to the free phosphonic acids by treatment with bromotrimethylsilane followed by hydrolysis. Reaction of 2,4-diamino-6-[[(diisopropoxyphosphoryl)methoxy]ethoxy]pyrimidine with elemental bromine, N-chloro-, or N-iodosuccinimide gave the corresponding phosphorus-protected 5-bromo-, 5-chloro-, and 5-iodo derivatives, respectively. Their deprotection afforded 2,4-diamino-5-bromo- and -5-chloro-6-[2-(phosphonomethoxy)ethoxy]pyrimidines. 2,4-Diamino-5-methyl-6-[2-(phosphonomethoxy)ethoxy]pyrimidine was synthesized also by cross-coupling of the 5-bromo compound with AlMe(3), followed by deprotection. The compounds showed poor, if any, inhibitory activity against DNA viruses such as herpes simplex virus type 1 and type 2, cytomegalovirus, varicella-zoster virus, and vaccinia virus. In contrast, several 5-substituted 2,4-diaminopyrimidine derivatives markedly inhibited retrovirus replication in cell culture. The 5-methyl derivative was exquisitely inhibitory to human immunodeficiency virus and Moloney murine sarcoma virus-induced cytopathicity in cell culture (EC(50) approximately 0.00018 mumol/mL) but also cytostatic to CEM cell cultures. In contrast, the 5-halogen-substituted derivatives showed pronounced antiretroviral activity (EC(50) = 0.0023-0.0110 mumol/mL), comparable to that of the reference drugs adefovir and tenofovir, but were devoid of measurable toxicity at 0.3 mumol/mL.     

1-((S)-2-aminopropyl)-1H-indazol-6-ol: a potent peripherally acting 5-HT2 receptor agonist with ocular hypotensive activity

Serotonin 5-HT(2) receptor agonists have been identified as a potential new class of agents for the treatment of ocular hypertension and glaucoma. The initially reported tryptamine analogues displayed either poor solution stability, potent central nervous system activity, or both of these undesirable characteristics and were unacceptable for clinical evaluation. A series of 1-(2-aminopropyl)-1H-indazole analogues was synthesized and evaluated for their suitability for consideration as clinical candidates. 1-((S)-2-Aminopropyl)-1H-indazol-6-ol (9) was identified as a peripherally acting potent 5-HT(2) receptor agonist (EC(50) = 42.7 nM, E(max) = 89%) with high selectivity for the 5-HT(2) receptors relative to other serotonergic receptor subtypes and other families of receptors and has significantly greater solution stability than alpha-methyl-5-hydroxytryptamine. Additionally, 9 potently lowers intraocular pressure in conscious ocular hypertensive monkeys (-13 mmHg, 33%); this reduction appears to be through a local rather than a centrally mediated effect. Compound 9 appears to be an excellent 5-HT(2) receptor agonist for conducting further studies directed toward a clinical proof-of-concept study for this class of ocular hypotensive agents.     

Differential selectivity of 5-fluorouracil and 5'-deoxy-5-fluorouridine in cultured human B lymphocytes and mouse L1210 leukemia

The role of differential metabolic activation of a 5-fluorouracil (FU) prodrug, 5'-deoxy-5-fluorouridine (dFUR), in achieving selective cytotoxicity was investigated in cultured human (dFUR), in achieving selective cytotoxocity was investigated in cultured human B lymphocytes and murine leukemia L1210 cells. B cells were cross-sensitive to FU and dFUR. On the other hand, leukemia L1210 cells were sensitive to FU but resistant to dFUR. The difference in the biological activities of FU and dFUR in B and L1210 cells correlated with (a) the metabolism of dFUR to FU by intact B (60% conversion) and L1210 (no conversion) cells, and (b) the phosphorylase activity of B (660 nmoles converted in 2 hr per mg protein) and L1210 (undetectable) cells. The intracellular metabolism of FU and dFUR was studied using a reversed-phase ion-pair high pressure liquid chromatographic assay. FU and dFUR shared similar metabolic pathways in B cells; their anabolites included FU ribose and deoxyribose nucleosides and nucleotides. In L1210 cells, FU was anabolized to 5-fluorouridine triphosphate and 5-fluorodeoxyuridine monophosphate, whereas dFUR was present mainly as the unchanged drug. Further metabolism studies using dFUR with tritium label in either the FU moiety or the altered sugar moiety established that the metabolic pathway of dFUR to cytotoxic FU anabolites in the B cells was via phosphorolysis to FU. These data indicate that, on a cellular level, an FU prodrug such as dFUR, which is activated by cytosolic enzyme, has a different selectivity from that of FU, and that the basis of differential selectivity is the initial phosphorolysis to FU.     

Discovery and optimization of 1-(4-(pyridin-2-yl)benzyl)imidazolidine-2,4-dione derivatives as a novel class of selective cannabinoid CB2 receptor agonists

Here, we report the identification and optimization of 1-(4-(pyridin-2-yl)benzyl)imidazolidine-2,4-dione derivatives as a novel chemotype with selective cannabinoid CB2 receptor agonist activity. 1 is a potent and selective cannabinoid CB2 receptor agonist (hCB2 pEC(50) = 8.6). The compound was found to be metabolically unstable, which resulted in low oral bioavailability in rat (F(po) = 4%) and possessed off-target activity at the hERG ion channel (pK(i) = 5.5). Systematic modification of physicochemical properties, such as lipophilicity and basicity, was used to optimize the pharmacokinetic profile and hERG affinity of this novel class of cannabinoid CB2 receptor agonists. This led to the identification of 44 as a potent, selective, and orally bioavailable cannabinoid CB2 receptor agonist (hCB2 pEC(50) = 8.0; hERG pK(i) < 4; F(po) = 100%), which was active in a rat spinal nerve ligation model of neuropathic pain.     

Synthesis and Structure-Opioid Activity Relationships of trans-(±)-3,4-Dichloro-N-methyl-N-[4- or 5-hydroxy-2-(1-pyrrolidiny)cyclohexyl]benzeneacetamides

To explore the effects of attaching a hydroxy function to the cyclohexane ring of κ-selective opioid N-[2-(1-pyrrolidinyl)cyclohexyl]benzeneacetamides, trans-(±)-3,4-dichloro-N-methyl-N-[4- or 5-hydroxy-2-(1-pyrrolidiny)cyclohexyl]benzeneacetamides (1–4) and their benzoates (5–8) have been synthesized in a divergent and stereoselective manner. When compared with the parent compound U-50488, hydroxy derivatives 1–4 maintained high selectivity towards the κ-opioid receptor (μ/κ ratio = 24 to >91); while displaying significant reduction in binding affinity (K_i,κ = 75–218 nm ). The lowest κ-affinity was observed with compound 4, where the hydroxy group is attached at the 5-axial or 5-β position. Further reduction in κ-affinity was observed when the hydroxy function was benzoylated. However, the 4β, 5α, and 5β isomers (6–8) maintained varying degrees of κ-selectivity; the 4α-isomer compound 5, with its benzoate moiety situated at the 4-axial position is now a moderately potent μ-selective opioid (K_i/μ= 168 nm , μ/κ = 0.076). The results suggest the importance of lipophilicity in binding to opioid receptors and the presence of a specific lipophilic binding site on the μ-opioid receptor.     

Synthesis and biological activity of 5-alkyl-6-(alkylsulfanyl)- or 5-alkyl-6-(arylsulfanyl)pyrazine-2-carboxamides and corresponding thioamides

Nucleophilic substitution of chlorine in 5-alkyl-6-chloropyrazine-2-carboxamides with various alkyl and arylthiolates afforded 20 5-alkyl-6-(alkylsulfanyl)- and 5-alkyl-6-(arylsulfanyl)pyrazine-2-carboxamides. The reaction of the amides with Lawesson's reagent yielded the corresponding thioamides. The assessment of in vitro antimycobacterial and antifungal activity of the compounds was carried out. In both series, the antimycobacterial activity increases with increasing molecular weight of the alkylsulfanyl group in position 6 of the pyrazine ring. Thioamides exhibited higher activity than the corresponding amides. 5-Butyl-6-(phenylsulfanyl)pyrazine-2-carbothioamide (2j) possessed the highest activity (91% inhibition) against Mycobacterium tuberculosis and also the highest lipophilicity (log P = 4.95). Only a poor in vitro antifungal effect was noted in 5-butyl-6-(butylsulfanyl)pyrazine-2-carboxamide (1i) and 6-(ethylsulfanyl)-5-isobutylpyrazine-2-carbothioamide (2q) against Trichophyton mentagrophytes and Absidia corymbifera.     

Synthesis and antiviral properties of 5-(2-substituted vinyl)-6-aza-2'-deoxyuridines

The following 5-(2-substituted vinyl)-6-aza-2'-deoxyuridines were synthesized: (E)-5-(2-bromovinyl) (2) (6-aza-BVDU), 5-(2-bromo-2-fluorovinyl) (a mixture of E and Z isomers) (3), (E)-5-(2-chlorovinyl) (4), (E)-5-[2-(methylthio)vinyl] (5), 5-(2,2-dibromovinyl) (6), and 5-(3-furyl) (7). The synthesis of 2-6 utilized Wittig-type reactions on 5-formyl-1-(2'-deoxy-3', 5'-di-O-p-toluoyl-beta-D-erythro-pentofuranosyl)-6-azauracil (16). 6-Aza-BVDU (and its alpha-anomer) was also synthesized from (E)-5-(2-bromovinyl)-6-azauracil (12) by using standard deoxyribosidation methodology. Compound 7 was prepared from 5-(3-furyl)-6-azauracil (33) via a ribosidation/deoxygenation sequence. An attempt to prepare the corresponding 5-(2,2-difluorovinyl) analogue afforded instead a mixture of the 5-[(2,2-difluoro-2-methoxy)ethyl] and 5-(2,2,2-trifluoroethyl) derivatives 29 and 30. Compounds 2-7, 29, and 30 were tested for in vitro activity against herpes simplex virus types 1 and 2 (HSV-1, HSV-2). 6-Aza-BVDU (2) exhibited ID50s of 8 micrograms/mL vs. HSV-1 and 190 micrograms/mL vs. HSV-2. BVDU (1) had ID50s of 0.015 and 1.6 micrograms/mL against HSV-1 and HSV-2, respectively. Compound 4 showed a similar profile of activity, but the other analogues were either weakly active or inactive.