Previous Article | Next Article 
Antimicrobial Agents and Chemotherapy, October 2000, p. 2883-2886, Vol. 44, No. 10
Medical Microbiology Division, Department of
Pathology, University of Iowa College of Medicine, Iowa City, Iowa
52242
Received 9 May 2000/Returned for modification 29 May 2000/Accepted 20 July 2000
The in vitro activities of the new triazole, ravuconazole
(BMS-207147), were compared to those of fluconazole and itraconazole against 541 clinical isolates of Cryptococcus neoformans.
Isolates were obtained from cerebrospinal fluid (396), blood (116), and miscellaneous clinical specimens (29). Overall, ravuconazole (MIC at
which 90% of the isolates are inhibited [MIC90], 0.25 µg/ml) was more active than either itraconazole (MIC90,
0.5 µg/ml) or fluconazole (MIC90, 8 µg/ml). Among the
isolates inhibited by Cryptococcus neoformans
has a worldwide distribution and is one of the most important agents of
life-threatening infection among the community-acquired opportunistic
fungal pathogens (4). Since the 1980s the incidence of
Cryptococcus infections in some countries has increased
dramatically as a result of AIDS (4, 7). In the United
States, the majority of studies report a prevalence of C. neoformans infection among human immunodeficiency virus
(HIV)-infected patients in the 5 to 10% range (4, 11). For
the treatment of cryptococcal meningitis, fluconazole has been
primarily used for maintenance therapy or prophylaxis (15). However, concerns regarding fluconazole-resistant strains of C. neoformans have been expressed by several investigators (2, 3). Itraconazole has been found to be less effective than
fluconazole in the treatment of cryptococcal meningitis in HIV-infected
patients (17). For these reasons, investigation of the
activities of newer antifungal agents against C. neoformans
is desired.
Ravuconazole (BMS-207147) is a novel triazole antifungal agent (1,
18) with a broad antifungal spectrum and potent activities against major pathogenic fungi such as Aspergillus
fumigatus, C. neoformans, Candida spp., and
dermatophytes (5, 6, 8, 9). Although its activity against
C. neoformans is promising, earlier investigations included
limited numbers of clinical isolates, and there is a lack of
comparative data with other azole agents. This study provides
comparative in vitro susceptibility data for three triazole antifungal
agents against a large number of clinical isolates of C. neoformans.
A total of 541 clinical isolates of C. neoformans from
geographically diverse locations were selected for this study. The collection included 396 isolates from cerebrospinal fluid cultures, 116 from blood cultures, and 29 from miscellaneous clinical specimens (pleural fluid, urine, etc.). All isolates were stored as suspensions in sterile distilled water at room temperature until the study was
performed. Prior to testing, each isolate was subcultured at least
twice on potato dextrose agar plates (Remel, Lenexa, Kans.) to ensure
purity and optimal growth.
Standard antifungal powders of ravuconazole (Bristol-Myers Squibb),
fluconazole (Pfizer), and itraconazole (Janssen) were obtained from
their respective manufacturers. Stock solutions were prepared in water
(fluconazole) or dimethyl sulfoxide (ravuconazole and itraconazole).
Antifungal agents were diluted with RPMI 1640 medium (Sigma, St. Louis,
Mo.), buffered to pH 7.0 with 0.165 M morpholinepropanesulfonic acid
(MOPS) buffer (Sigma), and dispensed into 96-well microdilution trays.
Trays containing an aliquot of 0.1 ml in each well were sealed and
frozen at Broth microdilution MICs were determined by the NCCLS method (12,
17). The yeast inoculum was adjusted spectrophotometrically to a
concentration of 0.5 × 103 to 2.5 × 103 cells/ml in RPMI 1640 medium, and an aliquot of 0.1 ml
was added to each well of the microdilution tray. The final
concentrations of the antifungal agents ranged from 0.007 to 8 µg/ml
for ravuconazole and itraconazole and from 0.125 to 128 µg/ml for
fluconazole. In each case, the inoculum size was verified by colony
counting. The microdilution trays were incubated at 35°C. The MIC
endpoints were read visually following 48 and 72 h of incubation,
and MIC results read at 72 h are reported herein. The MIC of each
triazole was defined as the lowest concentration that produced an 80%
reduction in growth (prominent decrease in turbidity) compared with
that of drug-free growth control (3, 5, 12, 14, 17).
Candida parapsilosis ATCC 22019 and C. krusei
ATCC 6258 were used as quality control organisms and were included each
time that a set of isolates was tested (5, 12, 14). Since
interpretive breakpoints for antifungal susceptibility testing of
C. neoformans have not yet been established, for simplicity
we adopted the fluconazole breakpoint values proposed by the NCCLS for
Candida spp. (12, 16) to Cryptococcus.
Ravuconazole and itraconazole were compared with respect to the
fluconazole susceptibility category (Table 1; also see Fig. 2).
0066-4804/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
In Vitro Activities of Ravuconazole (BMS-207147)
against 541 Clinical Isolates of Cryptococcus
neoformans
ABSTRACT
Top
Abstract
Text
References
16 µg of fluconazole/ml, 90.2% were
inhibited by 
1 µg of ravuconazole/ml. On the basis of our findings
and the favorable pharmacokinetic properties of ravuconazole, we
suggest that ravuconazole may be useful for the treatment of infectious
diseases due to C. neoformans and that further clinical
studies to confirm these promising in vitro results are warranted.
TEXT
Top
Abstract
Text
References
70°C until needed.
TABLE 1.
In vitro susceptibilities of 541 clinical isolates of
C. neoformans to ravuconazole and itraconazole stratified by
fluconazole susceptibility category
Figure 1 shows the cumulative
distribution of MICs for each of the three antifungal agents. The
isolates were generally susceptible to all three azole agents.
Fluconazole had MICs of 8 µg/ml for 90.6% of the C. neoformans isolates tested, 16 to 32 µg/ml for 8.1%, and 64
µg/ml for 1.3% of these isolates. Overall, ravuconazole was the most
active agent (MIC at which 90% of the isolates are inhibited
[MIC90], 0.25 µg/ml). Itraconazole and fluconazole
showed MIC90 at 0.5 and 8 µg/ml, respectively (Table 1
and Fig. 1). The Wilcoxon signed-rank test was used to compare
ravuconazole and itraconazole MICs. The difference in MICs was
statistically significant (P < 0.001), with the
ravuconazole MIC being lower than that of itraconazole for 432 of the
541 isolates.
|
), itraconazole (
), and fluconazole
(×).
Figure 2 shows the cumulative
distribution of ravuconazole and itraconazole MICs for the isolates
categorized as fluconazole susceptible (fluconazole MIC, 8 µg/ml)
and fluconazole susceptible-dose dependent (fluconazole MIC, 16 to 32 µg/ml). Among the isolates inhibited by 8 µg of fluconazole/ml,
ravuconazole was more potent than itraconazole (MIC50 and
MIC90, 0.12 and 0.25 µg/ml versus 0.25 and 0.5 µg/ml,
respectively). Out of 490 strains in this category, 96.3% (472 strains) were inhibited by 0.25 µg of ravuconazole/ml, and 72.9%
(357 strains) were inhibited by the same concentration of itraconazole
(P < 0.001).
|
Among the isolates inhibited by 16 to 32 µg of fluconazole/ml, ravuconazole remained slightly more active than itraconazole (P < 0.001). Since a significant shift toward higher ravuconazole and itraconazole MICs was observed in this category (Fig. 2), the independent sample t test was used to compare mean itraconazole and ravuconazole MICs between fluconazole-susceptible and fluconazole-susceptible-dose-dependent isolates. In each case, the mean MIC was lower for fluconazole-susceptible isolates than for fluconazole-susceptible-dose-dependent isolates (mean itraconazole MICs, 0.28 versus 0.65 µg/ml [P < 0.01]; mean ravuconazole MICs, 0.125 versus 0.45 µg/ml [P < 0.001]).
Seven isolates required 64 µg of fluconazole/ml to inhibit growth
in vitro. For these isolates, MICs ranged from 0.12 to 4.0 µg/ml for
ravuconazole and 0.12 to 2.0 µg/ml for itraconazole. Three isolates
were inhibited by 0.5 µg of either ravuconazole or itraconazole/ml.
These results support and extend findings reported previously (6,
8). We found that ravuconazole was more active than either
itraconazole or fluconazole against clinical isolates of C. neoformans. Both ravuconazole and itraconazole appeared most active against isolates exhibiting the greatest susceptibility to
fluconazole (MIC of fluconazole, 8 µg/ml). Although other investigators have evaluated the activity of ravuconazole against fluconazole-susceptible isolates (6, 8), we also examined the activity of ravuconazole against C. neoformans isolates
for which the fluconazole MICs were elevated. As the fluconazole MICs increased, stepwise increases in the MICs of both ravuconazole and
itraconazole were noted. However, a greater percentage of isolates
inhibited by 16 to 32 µg of fluconazole/ml remained susceptible (MIC,
0.25 µg/ml) to ravuconazole (56.8%) than to itraconazole (6.8%).
Out of seven isolates for which fluconazole MICs were 64 µg/ml,
four isolates required 1 µg of itraconazole/ml and 2 µg of
ravuconazole/ml to inhibit growth in vitro. These MICs for both
itraconazole and ravuconazole are considerably higher than the results
from other reports (6, 8, 13) and might represent
itraconazole- and ravuconazole-resistant strains of C. neoformans. Nonetheless, it is noteworthy that 3 isolates in the
fluconazole-resistant category were inhibited by 0.5 µg of either
ravuconazole or itraconazole/ml.
In vivo studies with animal models have demonstrated that ravuconazole has efficacy comparable to that of fluconazole and is more effective than itraconazole against systemic cryptococcosis (8), pulmonary cryptococcosis (9), and intracranial cryptococcosis (9). In addition, pharmacokinetic studies with humans using a 400-mg/day oral multiple dosing regimen (14 days) demonstrated peak plasma concentrations of 6.02 µg/ml, an area under the plasma concentration-time curve of 119.12 µg · h/ml, and a terminal half-life of 115 h (D. M. Grasela, S. J. Olsen, V. Mummaneni, P. Rolan, L. Christopher, J. Norton, O. H. Hadjilambris, and M. R. Marino, 40th Intersci. Conf. Antimicrob. Agents Chemother., abstr., 2000). By comparison, a 400-mg daily dose of itraconazole capsules (200 mg twice a day for 15 days) provides a maximum drug concentration of 2.28 µg/ml, an area under the concentration-time curve of 22.56 µg · h/ml, and a terminal half-life of 64 h (10). Plasma concentrations of ravuconazole have been maintained at more than four times the MIC90 for C. neoformans (0.25 µg/ml) from day 4 to day 31 following administration of 400 mg daily for 14 days (D. M. Grasela et al., 40th ICAAC).
In summary, we have demonstrated ravuconazole to be more potent than fluconazole and itraconazole against clinical isolates of C. neoformans in vitro. Although ravuconazole, like itraconazole, is highly protein bound (98%), the favorable pharmacokinetic properties and greater potency suggest that ravuconazole may be useful for the treatment of infectious diseases due to C. neoformans. Further clinical trials to confirm these promising in vitro results are warranted.
| |
ACKNOWLEDGMENTS |
|---|
We thank Daniel Diekema for help in statistical analysis.
Toshiaki Yamazumi is partly supported by a grant from the Japan Clinical Pathology Foundation for International Exchanges. This study was supported by a grant from Bristol-Myers Squibb Company.
| |
FOOTNOTES |
|---|
* Corresponding author. Mailing address: Medical Microbiology Division, Department of Pathology, 606GH, University of Iowa College of Medicine, Iowa City, IA 52242. Phone: (319) 384-9566. Fax: (319) 356-4916. E-mail: michael-pfaller{at}uiowa.edu.
Present address: Department of Clinical Pathology, Kinki University
School of Medicine, Ohnohigashi, Osakasayama, Osaka, Japan 589-8511.
| |
REFERENCES |
|---|
|
Top
Abstract
Text
References
|
|---|
| 1. | Bartroli, J., E. Turmo, M. Alguero, E. Boncompte, M. L. Vericat, L. Conte, J. Ramis, M. Merlos, J. Garcia-Rafanell, and J. Forn. 1998. New azole antifungals. 3. Synthesis and antifungal activity of 3-substituted-4(3H)-quinazolinones. J. Med. Chem. 41:1869-1882[CrossRef][Medline]. |
| 2. | Berg, J., C. J. Clancy, and M. H. Nguyen. 1998. The hidden danger of primary fluconazole prophylaxis for patients with AIDS. Clin. Infect. Dis. 26:186-187[Medline]. |
| 3. | Brandt, M. E., M. A. Pfaller, R. A. Hajjeh, E. A. Graviss, J. Rees, E. D. Spitzer, R. W. Pinner, L. W. Mayer, and the Cryptoccocal Disease Active Surveillance Group. 1996. Molecular subtypes and antifungal susceptibilities of serial Cryptococcus neoformans isolates in human immunodeficiency virus associated cryptococcosis. J. Infect. Dis. 174:812-820[Medline]. |
| 4. | Casadevall, A., and J. R. Perfect. 1998. Cryptococcus neoformans, p. 351-380. American Society for Microbiology, Washington, D.C. |
| 5. |
Diekema, D. J.,
M. A. Pfaller,
S. A. Messer,
A. Houston,
R. J. Hollis,
G. V. Doern,
R. N. Jones, and the SENTRY Participants Group.
1999.
In vitro activities of BMS-207147 against over 600 contemporary clinical blood stream isolates of Candida species from the SENTRY antimicrobial surveillance program in North America and Latin America.
Antimicrob. Agents Chemother.
43:2236-2239 |
| 6. |
Fung-Tomc, J. C.,
E. Huczko,
B. Minassian, and D. P. Bonner.
1998.
In vitro activity of a new oral triazole, BMS-207147 (ER-30346).
Antimicrob. Agents Chemother.
42:313-318 |
| 7. |
Hajjeh, R. A.,
M. E. Brandt, and R. W. Pinner.
1995.
Emergence of cryptococcal disease: epidemiologic perspectives 100 years after its discovery.
Epidemiol. Rev.
17:303-320 |
| 8. | Hata, K., J. Kimura, H. Miki, T. Toyosawa, T. Nakamura, and K. Katsu. 1996. In vitro and in vivo activities of ER-30346, a novel oral triazole with a broad antifungal spectrum. Antimicrob. Agents Chemother. 40:2237-2242[Abstract]. |
| 9. | Hata, K., J. Kimura, H. Miki, T. Toyosawa, M. Moriyama, and K. Katsu. 1996. Efficacy of ER-30346, a novel oral triazole antifungal agent, in experimental models of aspergillosis, candidiasis, and cryptococcosis. Antimicrob. Agents Chemother. 40:2243-2247[Abstract]. |
| 10. | Medical Economics Co., Inc. 2000. Physicians' desk reference, 54th ed., p. 1457-1460. Medical Economics Co., Inc., Oradell, N.J. |
| 11. |
Mitchell, T. G., and J. R. Perfect.
1995.
Cryptococcosis in the era of AIDS 100 years after the discovery of Cryptococcus neoformans.
Clin. Microbiol. Rev.
8:515-548[Abstract].
|
| 12. | National Committee for Clinical Laboratory Standards. 1997. Reference method for broth dilution antifungal susceptibility testing of yeasts. Approved standard M27-A. National Committee for Clinical Laboratory Standards, Wayne, Pa. |
| 13. |
Nguyen, M. H., and C. Y. Yu.
1998.
In vitro comparative efficacy of voriconazole and itraconazole against fluconazole-susceptible and resistant Cryptococcus neoformans isolates.
Antimicrob. Agents Chemother.
42:471-472 |
| 14. |
Pfaller, M. A.,
J. Zhang,
S. A. Messer,
M. E. Brandt,
R. A. Hajjeh,
C. J. Jessup,
M. Tumberland,
E. K. Mbidde, and M. A. Ghannoum.
1999.
In vitro activities of voriconazole, fluconazole, and itraconazole against 566 clinical isolates of Cryptococcus neoformans from the United States and Africa.
Antimicrob. Agents Chemother.
43:169-171 |
| 15. |
Powderly, W. G.,
D. M. Finkelstein,
J. Feinberg,
P. T. Frame,
W. He,
C. M. van der Horst,
S. L. Koletar,
M. E. Eyster,
J. Carey,
H. A. Waskin,
T. M. Hooton,
N. E. Hyslop,
S. A. Spector, and S. A. Bozzette.
1995.
A randomized trial comparing fluconazole with clotrimazole troches for the prevention of fungal infections in patients with advanced human immunodeficiency virus infection.
N. Engl. J. Med.
332:700-705 |
| 16. | Rex, J. H., M. A. Pfaller, J. N. Galgiani, M. S. Bartlett, A. Espinel-Ingroff, M. A. Ghannoum, M. Lancaster, F. C. Odds, M. G. Rinaldi, T. J. Walsh, and A. L. Barry. 1997. Development of interpretive breakpoints for antifungal susceptibility testing: conceptual framework and analysis of in vitro-in vivo correlation data for fluconazole, itraconazole, and Candida infections. Clin. Infect. Dis. 24:235-247[Medline]. |
| 17. | Sanati, H., S. A. Messer, M. A. Pfaller, M. Witt, R. Larsen, A. Espinel-Ingroff, and M. Ghannoum. 1996. Multicenter evaluation of broth microdilution method for susceptibility testing of Cryptococcus neoformans against fluconazole. J. Clin. Microbiol. 34:1280-1282[Abstract]. |
| 18. | Tsuruoka, A., Y. Kaku, H. Kakinuma, I. Tsukada, M. Yanagisawa, K. Nara, and T. Naito. 1998. Synthesis and antifungal activity of novel thiazole-containing triazole antifungals. II. Optically active ER-30346 and its derivatives. Chem. Pharm. Bull. 46:623-630. |
Antimicrobial Agents and Chemotherapy, October 2000, p. 2883-2886, Vol. 44, No. 10
0066-4804/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
This article has been cited by other articles:
-
Sionov, E., Chang, Y. C., Garraffo, H. M., Kwon-Chung, K. J.
(2009). Heteroresistance to Fluconazole in Cryptococcus neoformans Is Intrinsic and Associated with Virulence. Antimicrob. Agents Chemother.
53: 2804-2815
[Abstract] [Full Text] -
Yan, J.-H., Marino, M. R., Smith, R. A., Kanamaluru, V., O'Mara, E. M., Grasela, D. M.
(2006). The Effect of Ravuconazole on the Pharmacokinetics of Nelfinavir in Healthy Male Volunteers. J Clin Pharmacol
46: 193-200
[Abstract] [Full Text] -
Cuenca-Estrella, M., Gomez-Lopez, A., Mellado, E., Garcia-Effron, G., Monzon, A., Rodriguez-Tudela, J. L.
(2005). In Vitro Activity of Ravuconazole against 923 Clinical Isolates of Nondermatophyte Filamentous Fungi. Antimicrob. Agents Chemother.
49: 5136-5138
[Abstract] [Full Text] -
Groll, A. H., Mickiene, D., Petraitis, V., Petraitiene, R., Kelaher, A., Sarafandi, A., Wuerthwein, G., Bacher, J., Walsh, T. J.
(2005). Compartmental pharmacokinetics and tissue distribution of the antifungal triazole ravuconazole following intravenous administration of its di-lysine phosphoester prodrug (BMS-379224) in rabbits. J Antimicrob Chemother
56: 899-907
[Abstract] [Full Text] -
Pfaller, M. A., Messer, S. A., Boyken, L., Rice, C., Tendolkar, S., Hollis, R. J., Doern, G. V., Diekema, D. J.
(2005). Global Trends in the Antifungal Susceptibility of Cryptococcus neoformans (1990 to 2004). J. Clin. Microbiol.
43: 2163-2167
[Abstract] [Full Text] -
Trilles, L., Fernandez-Torres, B., dos Santos Lazera, M., Wanke, B., Guarro, J.
(2004). In Vitro Antifungal Susceptibility of Cryptococcus gattii. J. Clin. Microbiol.
42: 4815-4817
[Abstract] [Full Text] -
Cuenca-Estrella, M., Gomez-Lopez, A., Mellado, E., Garcia-Effron, G., Rodriguez-Tudela, J. L.
(2004). In Vitro Activities of Ravuconazole and Four Other Antifungal Agents against Fluconazole-Resistant or -Susceptible Clinical Yeast Isolates. Antimicrob. Agents Chemother.
48: 3107-3111
[Abstract] [Full Text] -
Pfaller, M. A., Messer, S. A., Boyken, L., Rice, C., Tendolkar, S., Hollis, R. J., Diekema, D. J.
(2004). Cross-Resistance between Fluconazole and Ravuconazole and the Use of Fluconazole as a Surrogate Marker To Predict Susceptibility and Resistance to Ravuconazole among 12,796 Clinical Isolates of Candida spp.. J. Clin. Microbiol.
42: 3137-3141
[Abstract] [Full Text] -
Petraitiene, R., Petraitis, V., Lyman, C. A., Groll, A. H., Mickiene, D., Peter, J., Bacher, J., Roussillon, K., Hemmings, M., Armstrong, D., Avila, N. A., Walsh, T. J.
(2004). Efficacy, Safety, and Plasma Pharmacokinetics of Escalating Dosages of Intravenously Administered Ravuconazole Lysine Phosphoester for Treatment of Experimental Pulmonary Aspergillosis in Persistently Neutropenic Rabbits. Antimicrob. Agents Chemother.
48: 1188-1196
[Abstract] [Full Text] -
Maxwell, M. J., Messer, S. A., Hollis, R. J., Diekema, D. J., Pfaller, M. A.
(2003). Evaluation of Etest Method for Determining Voriconazole and Amphotericin B MICs for 162 Clinical Isolates of Cryptococcus neoformans. J. Clin. Microbiol.
41: 97-99
[Abstract] [Full Text] -
Yamazumi, T., Pfaller, M. A., Messer, S. A., Houston, A. K., Boyken, L., Hollis, R. J., Furuta, I., Jones, R. N.
(2003). Characterization of Heteroresistance to Fluconazole among Clinical Isolates of Cryptococcus neoformans. J. Clin. Microbiol.
41: 267-272
[Abstract] [Full Text] -
Clemons, K. V., Martinez, M., Calderon, L., Stevens, D. A.
(2002). Efficacy of Ravuconazole in Treatment of Systemic Murine Histoplasmosis. Antimicrob. Agents Chemother.
46: 922-924
[Abstract] [Full Text] -
Kirkpatrick, W. R., Perea, S., Coco, B. J., Patterson, T. F.
(2002). Efficacy of ravuconazole (BMS-207147) in a guinea pig model of disseminated aspergillosis. J Antimicrob Chemother
49: 353-357
[Abstract] [Full Text]
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Copyright © 2009 by the American Society for Microbiology. For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»