FR264205 is a novel parenteral 3-aminopyrazolium cephalosporin. This study evaluated the in vitro and in vivo activities of FR264205 against Pseudomonas aeruginosa. The MIC of FR264205 at which 90% of 193 clinical isolates of P. aeruginosa were inhibited was 1 g/ml, 8-to 16-fold lower than those of ceftazidime (CAZ), imipenem (IPM), and ciprofloxacin (CIP). FR264205 also exhibited this level of activity against CAZ-, IPM-, and CIP-resistant P. aeruginosa. The reduction in the susceptibility of FR264205 by AmpC ␤-lactamase was lower than that of CAZ, indicating a relatively high stability of FR264205 against AmpC ␤-lactamase, the main resistance mechanism for cephalosporins. Neither expression of efflux pumps nor deficiency of OprD decreased the activity of FR264205. No spontaneous resistance mutants were selected in the presence of FR264205, and the reduction in susceptibility to FR264205 was lower than that to CAZ, IPM, and CIP after serial passage, suggesting that FR264205 has a low propensity for selecting resistance. In murine pulmonary, urinary tract, and burn wound models of infection caused by P. aeruginosa, the efficacy of FR264205 was superior or comparable to those of CAZ and IPM. These results indicate that FR264205 should have good potential as an antibacterial agent for P. aeruginosa.Nosocomial infections with gram-negative bacteria are a major problem for immunocompromised patients. Pseudomonas aeruginosa exhibits considerable inherent resistance, caused by low outer membrane permeability, multiple efflux pumps, and chromosomal AmpC ␤-lactamase (12). P. aeruginosa can also acquire additional resistance mechanisms, such as constitutive production of AmpC ␤-lactamase, OprD loss, and overproduction of efflux pumps. Although ceftazidime (CAZ) has been used as a first-line drug for P. aeruginosa infection, resistant mutants showing constitutive AmpC ␤-lactamase production can be selected in clinical settings, leading to therapeutic failure (2). P. aeruginosa has developed resistance not only to cephalosporins but also to carbapenems and quinolones. In 2003, the National Nosocomial Infections Surveillance System reported that resistance rates of P. aeruginosa to imipenem, quinolone, and broad-spectrum cephalosporins were 21.1, 29.5, and 31.9%, respectively. Compared to rates in the period between 1998 and 2002, these rates were increased by 15, 9, and 20%, respectively (15). Therefore, there is a critical need for new anti-P. aeruginosa agents that have no cross-resistance to currently marketed antibacterial agents and low propensities for inducing resistance.In order to generate promising anti-P. aeruginosa agents, research by our group has been directed toward the development of novel cephalosporins. As a result of exploration of structure-activity relationships of 3-(2,4-disubstituted 3-aminopyrazolio)methyl cephalosporins, FR264205 was discovered (Fig. 1). The antibacterial spectrum of FR264205 was similar to that of CAZ, and the MICs of FR264205 for Staphylococcus aureus ATCC 29213, Streptococcus pn...

The efficacy of intravenous injection of FK463, a novel water-soluble lipopeptide, was evaluated in mouse models of disseminated candidiasis and aspergillosis and was compared with those of fluconazole (FLCZ) and amphotericin B (AMPH-B). In the candidiasis model, FK463 significantly prolonged the survival of intravenously infected mice at doses of 0.125 mg/kg of body weight or higher. In disseminated candidiasis caused by Candida species, including FLCZ-resistant Candida albicans, FK463 exhibited an efficacy 1.4 to 18 times inferior to that of AMPH-B, with 50% effective doses (ED 50 s) ranging from 0.21 to 1.00 mg/kg and 0.06 to 0.26 mg/kg, respectively, and was much more active than FLCZ. The protective effect of FK463 was not obviously influenced by the fungal inoculum size, the starting time of the treatment, or the immunosuppressed status of the host. The reduction in efficacy was less than that observed with FLCZ or AMPH-B. The efficacy of FK463 was also evaluated in the disseminated candidiasis target organ assay and was compared with those of FLCZ and AMPH-B. Efficacies were evaluated on the basis of a comparison between the mean log 10 CFU in kidneys in the groups treated with antifungal agents and that in control group. A single dose of FK463 at 0.5 mg/kg or higher significantly reduced the viable counts in kidneys compared with the numbers of yeast cells before treatment, and its efficacy was comparable to that of AMPH-B, while FLCZ at 4 mg/kg showed only a suppressive effect on the growth of C. albicans in the kidneys. In the disseminated aspergillosis model, FK463 given at doses of 0.5 mg/kg or higher significantly prolonged the survival of mice infected intravenously with Aspergillus fumigatus conidia. The efficacy of FK463 was about 2 times inferior to that of AMPH-B, with ED 50 s ranging from 0.25 to 0.50 mg/kg and 0.11 to 0.29 mg/kg, respectively. These results indicate that FK463 may be a potent parenterally administered therapeutic agent for disseminated candidiasis and aspergillosis.People who have impaired immune systems are susceptible to fungal infections which can be life-threatening. Immune deficiencies resulting from AIDS, aggressive cancer treatment, the growing use of organ transplants, and other nosocomial situations have greatly increased the incidence of serious fungal infections (2,3,4,6) and have created a critical need for new, safe fungicidal agents that can be used to treat disseminated infections. Systemic mycoses are not easily diagnosed, and the patient usually has been infected for quite some time before symptoms appear. Thus, empiric therapy needs to begin immediately, but currently available treatments have problems with toxicity or resistance. Amphotericin B (AMPH-B) is the first-line therapy for systemic infections because of its broadspectrum and fungicidal activity. However, significant side effects limit its clinical utility to controlled intravenous administration (16). Lipid AMPH-B formulations have recently attracted much attention due to significantly lower toxic...

Sha (also known as Mrp/Mnh/Pha) is a Na؉ /H ؉ antiporter encoded by a cluster of six or seven genes that probably form a multisubunit transport complex. The Sha system is important for the homeostasis of H ؉ , Na ؉ , and other monovalent cations and plays a critical role in various functions, including alkaliphily, sporulation, and symbiosis. Here, we characterized the sha homologue genes from the opportunistic pathogen Pseudomonas aeruginosa, which exist as a cluster of six genes (PA1054 to PA1059). The gene cluster PA1054 to PA1059, but not the cluster with a deletion of PA1054, complemented a growth defect in the presence of 0.2 M NaCl and a defect in Na ؉ /H ؉ antiport activity of the Escherichia coli TO114 mutant lacking the three major Na ؉ /H ؉ antiporters, indicating that genes PA1054 to PA1059 are responsible for Na ؉ /H ؉ antiport activity. We disrupted PA1054 (a shaA homologue gene) and determined its effect on Na ؉ tolerance during growth, Na ؉ efflux, and pathogenicity in mice. Disruption of PA1054 resulted in severe Na ؉ sensitivity during growth and decreased Na ؉ efflux activity. In mice, the deletion mutant of PA1054 also exhibited an attenuated virulence in systemic, pulmonary, and urinary tract infections and also a decrease in colonization of the infected organs. From these results, we conclude that the genes PA1054 to PA1059 encode a Na ؉ /H ؉ antiporter that is largely responsible for Na ؉ extrusion in P. aeruginosa and has a role in the infection of the pathogen. We propose to designate PA1054 to PA1059 as the sha (sodium hydrogen antiporter) genes, shaABCDEFG.

The efficacy of FK463, a novel water-soluble lipopeptide, was evaluated in mouse models of pulmonary aspergillosis and was compared with that of amphotericin B (AMPH-B). In the pulmonary aspergillosis models induced by intranasal inoculation, FK463 exhibited good efficacy, with 50% effective doses in the range of 0.26 to 0.51 mg/kg of body weight; these values were comparable to those of AMPH-B. In an Aspergillus target organ assay with immunosuppressed mice, under conditions of constant plasma levels of FK463, using a subcutaneously implanted osmotic pressure pump, a significant reduction in viable fungal cells was observed at plasma FK463 levels of 0.55 to 0.80 g/ml or higher. We conclude that FK463 is highly effective in the treatment of pulmonary aspergillosis in this animal model. These results indicate that FK463 may be a potent parenterally administered antifungal agent for pulmonary aspergillosis.Deep-seated mycoses in immunocompromised hosts are becoming an increasingly important medicinal problem (10). Candidiasis is the most common and important fungal infection in humans, and aspergillosis is the next most common (2,8,9). Invasive aspergillosis in immunocompromised patients is associated with significant morbidity and mortality (3,4). In view of the well-known problems with the established agents, it is clear that newer antifungal therapies with improved efficacy and reduced toxicity are needed. FK463 is a new, parenterally administered antifungal drug candidate undergoing clinical development. This compound is a novel water-soluble lipopeptide derived by semisynthetic modification of FR901379, a naturally occurring cyclic hexapeptide with a fatty acyl side chain, similar in structure to echinocandins and pneumocandins (T. Iwamoto, N. Sakamoto, M. Yamashita, M. Ezaki, S. Hashimoto, T. Furuta, M. Okuhara, and M. Kohsaka, Prog. Abstr. 33rd Intersci. Conf. Antimicrob. Agents Chemother., abstr. 371, 1993 F141, p. 268, 1998). In mouse models of disseminated candidiasis and aspergillosis, FK463 showed good efficacy (S. Matsumoto, Y. Wakai, K. Maki, E. Watabe, T. Ushitani, K. Otomo, T. Nakai, Y. Watanabe, F. Ikeda, S. Tawara, T. Goto, F. Matsumoto, and S. Kuwahara, Abstr. 38th Intersci. Conf. Antimicrob. Agents Chemother., abstr. F142, p. 268, 1998). In the study described in this report, the activity of FK463 was evaluated in mouse models of pulmonary aspergillosis. MATERIALS AND METHODS Compound and animals. FK463 was synthesized in Fujisawa PharmaceuticalCo., Ltd. Amphotericin B (AMPH-B) and fluconazole (FLCZ) were purchased from Bristol-Myers Squibb (Tokyo, Japan) and Pfizer (Tokyo, Japan), respectively. FK463 and AMPH-B were formulated in sterile saline and 5% glucose, respectively, for intravenous injection and injected as 10mL/kg. Male Slc-ICR strain mice (4 weeks old) were purchased from SLC Japan (Shizuoka, Japan).Organisms and media. Inocula of Aspergillus fumigatus TIMM0063, IFM40814, and IFM41209 were prepared by culturing the test organisms on potato dextrose agar. Conidia were collected in s...

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The protective effect of an immunoactive peptide, D-lactoyl-L-alanyl-r-D-glutamyl-(L)meso-diaminopimelyl-(L)-glycine (FK-156) and a related compound, heptanoyl-r-D-glutamyl-(L)meso-diaminopimelyl-(D)-alanine (FK-565) was determined in mice with various kinds of microbial infections. FK-156 and FK-565 were given to mice either subcutaneously or orally before challenge. The drugs enhanced significantly the defense of mice against acute systemic infections induced by various extracellular and facultative intracellular organisms, and subcutaneous abscess by Staphylococcus aureus. The protective effect of these drugs against Escherichia coli infection differed considerably depending on the route of administration; FK-156 was only effective by the parenteral route; however, FK-565 was effective by both parenteral and oral routes. After subcutaneous dosing with FK-156, the enhancement of host defense of mice against E. coli infection was more rapid than against Listeria infection. The enhancing effects of FK-156 and FK-565 on host defense of mice against pseudomonal infection was more potent than other immunoactive drugs.

␤-Lactams have been considered ineffective against organisms growing inside mammalian cells because of their poor penetration into cells. However, cefixime has been shown to be clinically effective against typhoid fever. The probable mechanism of therapeutic effectiveness of cefixime against typhoid fever was investigated using Salmonella enterica serovar Typhimurium instead of S. enterica serovar Typhi both in a cellular and in a mouse infection model. Cefixime was able to inhibit the growth of serovar Typhimurium inhabiting monocytederived THP-1 cells. Elongation of serovar Typhimurium in THP-1 cells was observed microscopically. Apparent morphological changes of serovar Typhimurium in THP-1 cells were also observed by electron microscopy. The concentration of cefixime inside THP-1 cells was almost half (46 to 48%) of the concentration outside the cells when serovar Typhimurium coexisted in the solution. The length of time after oral dosing (8 mg/kg) that cefixime was present-calculated from levels in serum-at a concentration above the MIC at which 90% of the serovar Typhi organisms inside human cells were inhibited was presumed to be more than 12 h. Cefixime also showed excellent activity in the mouse systemic and oral infection models based on infections caused by serovar Typhimurium. It is concluded that a fair amount of cefixime can enter mammalian cells and inhibit the growth of bacteria inside cells when the bacteria are sensitive enough to cefixime, as are serovars Typhimurium and Typhi.

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