Combinatorial gene expression using multiple episomal vectors.

Episomal vectors offer a powerful alternative to integrative recombination for transgene expression in mammalian cells. In this study, various combinations of G protein-coupled receptors (GPCRs) and the G protein subunit G(i2)alpha, were stably expressed from separate episomal vectors in 293-EBNA (293E) cells. Each episome did not adversely affect the others, as gauged by episomal copy number, steady-state mRNA levels and the presence of functional receptors and G protein. Cell lines expressing genes from multiple autonomously replicating vectors were stable just two weeks after transfection, and remained stable in continuous culture for at least 5months. Co-expression of supplementary G(i2)alpha with receptor amplifies the magnitude of signal transduction thereby permitting the development of more sensitive high throughput functional assays. Given these results, combinatorial transfection is the strategy of choice for generating stable cell lines expressing multiple genes for the study of signal-transduction pathways or the evaluation of receptor ligands.

Activity of temafloxacin against respiratory pathogens.

The activity of the quinolone temafloxacin against respiratory pathogens was compared with those of ciprofloxacin and ofloxacin. MICs for 90% of strains tested indicated that temafloxacin was at least two- to fourfold more potent than the other two quinolones against Staphylococcus aureus, Streptococcus pneumoniae, and Legionella pneumophila. Temafloxacin had potency equal to that of ciprofloxacin and was twofold more active than ofloxacin against Streptococcus pyogenes. Moraxella catarrhalis, and Bordetella pertussis. Against Haemophilus influenzae and Klebsiella pneumoniae, temafloxacin was four- and twofold less potent than ciprofloxacin, respectively. When administered orally in mouse protection tests against S. aureus, S. pneumoniae, and S. pyogenes, temafloxacin was at least eight times more potent than ciprofloxacin and was two to four times more active than ofloxacin. Against H. influenzae, temafloxacin was as active as ofloxacin and was two times less active than ciprofloxacin following oral administration in mice. In treating L. pneumophila in guinea pigs and H. influenzae otitis media in gerbils, temafloxacin and ofloxacin were more effective than ciprofloxacin. Against S. pneumoniae otitis media in gerbils, temafloxacin and ciprofloxacin were more active than ofloxacin. Following subcutaneous administration in mice, temafloxacin achieved higher lung levels than ciprofloxacin or ofloxacin did.

Antipneumococcal activity of ciprofloxacin, ofloxacin, and temafloxacin in an experimental mouse pneumonia model at various stages of the disease.

The efficacy of temafloxacin against Streptococcus pneumoniae in an experimental murine pneumonia model was compared with that of ofloxacin and ciprofloxacin. Erythromycin and amoxicillin were used as reference agents. Subcutaneous administration of antibiotics every 12 h for 3 days was initiated at various times after infection. The cumulative survival rates of mice treated with temafloxacin at 50 mg/kg were 100%, 92%, 81%, and 50% with treatment beginning 18, 48, 72, and 96 h after infection, respectively. The activity of temafloxacin at 50 mg/kg was not significantly different from that of erythromycin and amoxicillin but was superior to that of ofloxacin and ciprofloxacin. The maximum cumulative survival rates of mice treated with ofloxacin and ciprofloxacin at 100 mg/kg were 67% and 50%, respectively, with treatment beginning 18 h after infection. Treatment with ofloxacin and ciprofloxacin at 50 mg/kg 18 h after infection did not significantly increase survival rates compared with those of untreated controls.