NPP-669, a Novel Broad-Spectrum Antiviral Therapeutic with Excellent Cellular Uptake, Antiviral Potency, Oral Bioavailability, Preclinical Efficacy, and a Promising Safety Margin.

DNA viruses are responsible for many diseases in humans. Current treatments are often limited by toxicity, as in the case of cidofovir (CDV, Vistide), a compound used against cytomegalovirus (CMV) and adenovirus (AdV) infections. CDV is a polar molecule with poor bioavailability, and its overall clinical utility is limited by the high occurrence of acute nephrotoxicity. To circumvent these disadvantages, we designed nine CDV prodrug analogues. The prodrugs modulate the polarity of CDV with a long sulfonyl alkyl chain attached to one of the phosphono oxygens. We added capping groups to the end of the alkyl chain to minimize ß-oxidation and focus the metabolism on the phosphoester hydrolysis, thereby tuning the rate of this reaction by altering the alkyl chain length. With these modifications, the prodrugs have excellent aqueous solubility, optimized metabolic stability, increased cellular permeability, and rapid intracellular conversion to the pharmacologically active diphosphate form (CDV-PP). The prodrugs exhibited significantly enhanced antiviral potency against a wide range of DNA viruses in infected human foreskin fibroblasts. Single-dose intravenous and oral pharmacokinetic experiments showed that the compounds maintained plasma and target tissue levels of CDV well above the EC50 for 24 h. These experiments identified a novel lead candidate, NPP-669. NPP-669 demonstrated efficacy against CMV infections in mice and AdV infections in hamsters following oral (p.o.) dosing at a dose of 1 mg/kg BID and 0.1 mg/kg QD, respectively. We further showed that NPP-669 at 30 mg/kg QD did not exhibit histological signs of toxicity in mice or hamsters. These data suggest that NPP-669 is a promising lead candidate for a broad-spectrum antiviral compound.

The Intriguing Effects of Substituents in the N-Phenethyl Moiety of Norhydromorphone: A Bifunctional Opioid from a Set of "Tail Wags Dog" Experiments.

(-)-N-Phenethyl analogs of optically pure N-norhydromorphone were synthesized and pharmacologically evaluated in several in vitro assays (opioid receptor binding, stimulation of [35S]GTPγS binding, forskolin-induced cAMP accumulation assay, and MOR-mediated ß-arrestin recruitment assays). "Body" and "tail" interactions with opioid receptors (a subset of Portoghese's message-address theory) were used for molecular modeling and simulations, where the "address" can be considered the "body" of the hydromorphone molecule and the "message" delivered by the substituent (tail) on the aromatic ring of the N-phenethyl moiety. One compound, N-p-chloro-phenethynorhydromorphone ((7aR,12bS)-3-(4-chlorophenethyl)-9-hydroxy-2,3,4,4a,5,6-hexahydro-1H-4,12-methanobenzofuro[3,2-e]isoquinolin-7(7aH)-one, 2i), was found to have nanomolar binding affinity at MOR and DOR. It was a potent partial agonist at MOR and a full potent agonist at DOR with a δ/µ potency ratio of 1.2 in the ([35S]GTPγS) assay. Bifunctional opioids that interact with MOR and DOR, the latter as agonists or antagonists, have been reported to have fewer side-effects than MOR agonists. The p-chlorophenethyl compound 2i was evaluated for its effect on respiration in both mice and squirrel monkeys. Compound 2i did not depress respiration (using normal air) in mice or squirrel monkeys. However, under conditions of hypercapnia (using air mixed with 5% CO2), respiration was depressed in squirrel monkeys.

Modulation of opioid receptor affinity and efficacy via N-substitution of 9ß-hydroxy-5-(3-hydroxyphenyl)morphan: Synthesis and computer simulation study.

The enantiomers of a variety of N-alkyl-, N-aralkyl-, and N-cyclopropylalkyl-9ß-hydroxy-5-(3-hydroxyphenyl)morphans were synthesized employing cyanogen bromide and K2CO3 to improve the original N-demethylation procedure. Their binding affinity to the µ-, δ-, and κ-opioid receptors (ORs) was determined and functional (GTPγ35S) assays were carried out on those with reasonable affinity. The 1R,5R,9S-enantiomers (1R,5R,9S)-(-)-5-(3-hydroxyphenyl)-2-(4-nitrophenethyl)-2-azabicyclo[3.3.1]nonan-9-ol (1R,5R,9S-16), (1R,5R,9S)-(-) 2-cinnamyl-5-(3-hydroxyphenyl)-2-azabicyclo[3.3.1]nonan-9-ol (1R,5R,9S-20), and (1R,5R,9S)-(-)-5-(3-hydroxyphenyl)-2-(4-(trifluoromethyl)phenethyl)-2-azabicyclo[3.3.1]nonan-9-ol (1R,5R,9S-15), had high affinity for the µ-opioid receptor (e.g., 1R,5R,9S-16: Ki=0.073, 0.74, and 1.99nM, respectively). The 1R,5R,9S-16 and 1R,5R,9S-15 were full, high efficacy µ-agonists (EC50=0.74 and 18.5nM, respectively) and the former was found to be a partial agonist at δ-OR and an antagonist at κ-OR, while the latter was a partial agonist at δ-OR and κ-OR in the GTPγ35S assay. The enantiomer of 1R,5R,9S-16, (+)-1S,5S,9R-16 was unusual, it had good affinity for the µ-OR (Ki=26.5nM) and was an efficacious µ-antagonist (Ke=29.1nM). Molecular dynamics simulations of the µ-OR were carried out with the 1R,5R,9S-16 µ-agonist and the previously synthesized (1R,5R,9S)-(-)-5-(9-hydroxy-5-(3-hydroxyphenyl-2-phenylethyl)-2-azabicyclo[3.3.1]nonane (1R,5R,9S-(-)-NIH 11289) to provide a structural basis for the observed high affinities and efficacies. The critical roles of both the 9ß-OH and the p-nitro group are elucidated, with the latter forming direct, persistent hydrogen bonds with residues deep in the binding cavity, and the former interacting with specific residues via highly structured water bridges.

Cocaine and desipramine elicit distinct striatal noradrenergic and behavioral responses in selectively bred obesity-resistant and obesity-prone rats.

Previous studies have demonstrated a role for norepinephrine (NE) in energy regulation and feeding, and basal differences have been observed in hypothalamic NE systems in obesity-prone vs. obesity-resistant rats. Differences in the function of brain reward circuits, including in the nucleus accumbens (NAc), have been shown in obesity-prone vs. obesity-resistant populations, leading many researchers to explore the role of striatal dopamine in obesity. However, alterations in NE transmission also affect NAc mediated behaviors. Therefore, here we examined differences in striatal NE and the response to norepinephrine transporter blockers in obesity-prone and obesity-resistant rats. We found that striatal NE levels increase following systemic cocaine administration in obesity-prone, but not obesity-resistant rats. This could result from either blockade of striatal norepinephrine transporters (NET) by cocaine leading to reduced NE reuptake, or circuit-based responses following cocaine administration resulting in increased NE release. Retrodialysis of the NET inhibitor, desipramine, into the ventral striatum did not cause selective increases in striatal NE levels in obesity-prone rats, suggesting that circuit-based mechanisms underlie NE increases following systemic cocaine administration. Consistent with this, systemic desipramine treatment decreased locomotor activity in obesity-prone, but not obesity-resistant rats. Furthermore, obesity-prone rats were also more sensitive to desipramine-induced reductions in food intake compared to obesity-resistant rats. Taken together, these data expand our understanding of differences in NE systems of obesity-prone vs. resistant rats, and provide new insights into basal differences in striatal systems that may influence feeding behavior.

Pre-existing differences in motivation for food and sensitivity to cocaine-induced locomotion in obesity-prone rats.

Obesity is a significant problem in the United States, with roughly one third of adults having a body mass index (BMI) over thirty. Recent evidence from human studies suggests that pre-existing differences in the function of mesolimbic circuits that mediate motivational processes may promote obesity and hamper weight loss. However, few preclinical studies have examined pre-existing neurobehavioral differences related to the function of mesolimbic systems in models of individual susceptibility to obesity. Here, we used selectively bred obesity-prone and obesity-resistant rats to examine 1) the effect of a novel "junk-food" diet on the development of obesity and metabolic dysfunction, 2) over-consumption of "junk-food" in a free access procedure, 3) motivation for food using instrumental procedures, and 4) cocaine-induced locomotor activity as an index of general mesolimbic function. As expected, eating a sugary, fatty, "junk-food" diet exacerbated weight gain and increased fasted insulin levels only in obesity-prone rats. In addition, obesity-prone rats continued to over-consume junk-food during discrete access testing, even when this same food was freely available in the home cage. Furthermore, when asked to press a lever to obtain food in an instrumental task, rates of responding were enhanced in obesity-prone versus obesity-resistant rats. Finally, obesity-prone rats showed a stronger locomotor response to 15 mg/kg cocaine compared to obesity-resistant rats prior to any diet manipulation. This enhanced sensitivity to this dose of cocaine is indicative of basal differences in the function of mesolimbic circuits in obesity-prone rats. We speculate that pre-existing differences in motivational systems may contribute to over-consumption and enhanced motivation in susceptible individuals.