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Monophosphate prodrug analogs of 2’-deoxy-2’-fluoro-2’-C-methylguanosine have been reported as potent inhibitors of hepatitis C virus (HCV) RNA-dependent RNA polymerase. These prodrugs also display potent anti-dengue activities in cellular assays although their prodrug moieties were designed to produce high levels of triphosphate in the liver. Since peripheral blood mononuclear cells (PBMCs) are one of the major targets of dengue virus, different prodrug moieties were designed to effectively deliver 2’-deoxy-2’-fluoro-2’-C-methylguanosine monophosphate prodrugs and their corresponding triphosphates into PBMCs after oral administration. We identified a cyclic phosphoramidate prodrug 17 demonstrating a well-balanced anti-dengue cellular activity and in vitro stability profiles. In dogs, oral administration of 17 resulted in high PBMC triphosphate level, exceeding TP50 (the intracellular triphosphate concentration at which 50% of virus replication is inhibited) at 10 mg/kg. Compound 17 demonstrated 1.6- and 2.2 log viremia reduction in the dengue mouse model at 100 and 300 mg/kg twice daily, respectively. At 100 mg/kg twice daily, the terminal triphosphate concentration in PBMCs reached above TP50, defining for the first time the minimum efficacious dose for a nucleos(t)ide prodrug. In the two-week dog toxicity studies at 30 to 300 mg/kg/day, no observed adverse effect level (NOAEL) could not be achieved due to pulmonary inflammation and hemorrhage. The preclinical safety results suspended further development of 17. Nevertheless, present work has proven the concept that an efficacious monophosphate nucleoside prodrug could be developed for the potential treatment of dengue infection.

Objectives The neutropenic murine thigh infection model was used to assess the effectiveness of IID572, a novel β-lactamase inhibitor, in rescuing piperacillin activity against bacterial strains expressing various β-lactamase enzymes. Methods Mice (n = 4/group) were inoculated with Enterobacteriaceae or Staphylococcus aureus bacterial strains expressing a range of β-lactamases via intramuscular injection. Two hours after bacterial inoculation, subcutaneous treatment with piperacillin/IID572 or piperacillin/tazobactam every 3 h was initiated. Animals were euthanized via CO2 24 h after the start of therapy and bacterial cfu (log10 cfu) per thigh was determined, and the static dose was calculated. Results In a dose-dependent manner, piperacillin/IID572 reduced the thigh bacterial burden in models established with Enterobacteriaceae producing class A, C and D β-lactamases (e.g. ESBLs, KPC, CMY-2 and OXA-48). Piperacillin/IID572 was also efficacious against MSSA strains, including one producing β-lactamase. Static doses of piperacillin/IID572 were calculable from animals infected with all strains tested and the calculated static doses ranged from 195 to 4612 mg/kg/day piperacillin, the active component in the combination. Of the 13 strains investigated, a 1 log10 bacterial reduction was achieved for 9 isolates and a 2 log10 reduction was achieved for 3 isolates; piperacillin/tazobactam was not efficacious against 6 of the 13 isolates tested. Conclusions In contrast to tazobactam, IID572 was able to rescue piperacillin efficacy in murine thigh infection models established with β-lactamase-producing strains of Enterobacteriaceae and S. aureus, including those expressing ESBLs or serine carbapenemases.

Resistance in Gram-negative bacteria to β-lactam drugs is mediated primarily by the expression of β-lactamases, and co-dosing of β-lactams with a β-lactamase inhibitor (BLI) is a clinically proven strategy to address resistance. New β-lactamases that are not impacted by existing BLIs are spreading and creating the need for development of novel broader spectrum BLIs. IID572 is a novel broad spectrum BLI of the diazabicyclooctane (DBO) class that is able to restore the antibacterial activity of piperacillin against piperacillin/tazobactam-resistant clinical isolates. IID572 is differentiated from other DBOs by its broad inhibition of β-lactamases and the lack of intrinsic antibacterial activity.

Background We sought to characterise a refined rat model of respiratory infection with P. aeruginosa over an acute time course and test the antibiotic ciprofloxacin. Methods Agar beads were prepared±SPAN ® 80. Rats were inoculated with sterile agar beads or those containing 10 5 colony forming units (cfu) P. aeruginosa via intra-tracheal dosing. Bacterial load and inflammatory parameters were measured. Results Differing concentrations of SPAN ® 80 modified median agar bead diameter and reduced particle size distribution. Beads prepared with 0.01% v/v SPAN ® 80 were evaluated in vivo . A stable lung infection up to 7days post infection was achieved and induced BALF neutrophilia 2 and 5days post infection. Ciprofloxacin (50mg/kg) significantly attenuated infection without affecting the inflammatory parameters measured. Conclusion SPAN ® 80 can control the particle size and lung distribution of agar beads and P. aeruginosa -embedded beads prepared with 0.01%v/v SPAN ® 80 can induce infection and inflammation over 7days.

A workshop to discuss anti-inflammatory approaches in the treatment of CF was held at Novartis Institutes for Biomedical Research (NIBR, Horsham, UK) in March 2008. Key opinion leaders in the field (Hugo De Jonge, Stuart Elborn, Erich Gulbins, Mike Konstan, Rick Moss, Scott Randell and Adriano Rossi), and NIBR scientists were brought together to collectively address three main aims: (i) to identify anti-inflammatory targets in CF, (ii) to evaluate the pros and cons of targeting specific cell types and (iii) to discuss model systems to profile potential therapeutic agents. The highlights of the workshop are captured in this review.

Background Recent clinical data suggest statins have transient but significant effects in patients with pulmonary arterial hypertension. In this study we explored the molecular effects of statins on distal human pulmonary artery smooth muscle cells (PASMCs) and their relevance to proliferation and apoptosis in pulmonary arterial hypertension. Methods Primary distal human PASMCs from patients and controls were treated with lipophilic (simvastatin, atorvastatin, mevastatin and fluvastatin), lipophobic (pravastatin) and nitric-oxide releasing statins and studied in terms of their DNA synthesis, proliferation, apoptosis, matrix metalloproteinase-9 and endothelin-1 release. Results Treatment of human PASMCs with selected statins inhibited DNA synthesis, proliferation and matrix metalloproteinase-9 production in a concentration-dependent manner. Statins differed in their effectiveness, the rank order of anti-mitogenic potency being simvastatin > atorvastatin > > pravastatin. Nevertheless, a novel nitric oxide-releasing derivative of pravastatin (NCX 6550) was effective. Lipophilic statins, such as simvastatin, also enhanced the anti-proliferative effects of iloprost and sildenafil, promoted apoptosis and inhibited the release of the mitogen and survival factor endothelin-1. These effects were reversed by mevalonate and the isoprenoid intermediate geranylgeranylpyrophosphate and were mimicked by inhibitors of the Rho and Rho-kinase. Conclusions Lipophilic statins exert direct effects on distal human PASMCs and are likely to involve inhibition of Rho GTPase signalling. These findings compliment some of the recently documented effects in patients with pulmonary arterial hypertension.