The use of echinocandins for the treatment of invasive zygomycosis (IZ) is controversial since they show no activity in vitro. Although posaconazole shows promising results, the mortality rates of patients with IZ treated with posaconazole, at 30%, remain unacceptably high. The use of new therapies with alternative agents alone or combined is mandatory.
This is the first study to evaluate the in vitro effect of combining posaconazole (Schering-Plough, Kenilworth, NJ) and caspofungin (MSD, Merck Research Laboratories, Rahway, NJ) against 12 clinical zygomycetes. Zygomycetes were identified by conventional morphological methods (8).
Drug combinations were tested by using a checkerboard modification of the guidelines presented in CLSI document M38-A. The final concentrations of posaconazole and caspofungin were 0.06 to 4 μg/ml and 0.25 to 256 μg/ml, respectively. The adjusted suspensions of sporangiospores had an optical density of 0.15 to 0.17 (68 to 70% transmittance). The MIC for each drug alone and its combinations was defined as the lowest concentration that produced complete visual inhibition of fungal growth (6). Drug interactions were defined as synergistic if the lowest fractional inhibitory concentration index was ≤0.5 (6). Although the caspofungin MICs were all above serum drug levels, the combination proved to be synergistic in all strains (Table 1).
TABLE 1.
In vitro activities of posaconazole and caspofungin alone and in combination against 12 zygomycete isolates obtained by a checkerboard modification of the CLSI M38-A procedure
| Zygomycete strain no. | MIC (μg/ml) of drug:
|
Lowest FICIa for combination | ||
|---|---|---|---|---|
| Alone
|
In combination (caspofungin/posaconazole) | |||
| Caspofungin | Posaconazole | |||
| Rhizopus | ||||
| 0 | >256 | 1 | 32/0.25 | 0.312 |
| 3 | >256 | 1 | 16/0.25 | 0.281 |
| 8 | >256 | 1 | 8/0.25 | 0.265 |
| Absidia | ||||
| 11 | >256 | 1 | 4/0.25 | 0.258 |
| 17 | 256 | 1 | 16/0.25 | 0.281 |
| 18 | >256 | 0.5 | 64/0.125 | 0.375 |
| Cunninghamella sp. strain 22 | >256 | 1 | 128/0.25 | 0.5 |
| Snycephalastrum sp. strain 23 | >256 | 1 | 8/0.125 | 0.187 |
| Rhizomucor sp. strain 25 | 128 | 2 | 32/0.5 | 0.375 |
| Mucor | ||||
| 28 | 256 | 0.5 | 32/0.125 | 0.375 |
| 39 | >256 | 1 | 64/0.125 | 0.25 |
| 43 | 256 | 0.5 | 32/0.125 | 0.375 |
Fractional inhibitory concentration index.
Caspofungin exhibits minimal activity against zygomycetes when tested in vitro, although this information should be interpreted with caution (10). The mechanisms underlying the lack of activity of echinocandins against zygomycetes are believed to be attributable to differences in their cell wall compositions, as these organisms largely contain 1,3-α-glucan and glycuronomannoproteins instead of 1,3-β-d-glucan. However, Rhizopus oryzae expressed the target enzyme for caspofungin (FKS1), and this drug presented limited activity in a murine model of disseminated IZ (3).
Combinations of two drugs have been studied mainly for the treatment of cryptococcal meningitis (5). The combination of caspofungin and posaconazole has been evaluated in other fungi and shows synergy or indifference but never antagonism (1, 7). The combination of two antifungal agents has been shown to be effective against zygomycetes in vitro and in vivo (2). In animal models, the combination of caspofungin and amphotericin B lipid complex was synergistic, improving the survival of mice (9). Few cases of IZ responded partially to the echinocandins, although we have so little clinical experience with caspofungin for the treatment of IZ (4, 11). These data suggest that echinocandins may have a role as a second agent in combinations.
The precise mechanism of synergy found between posaconazole and caspofungin must be shown. A possible explanation may be that the inhibition of the synthesis of ergosterol by posaconazole may be changing the membrane environment and making the enzyme FKS1 more accessible or sensitive to inhibition by caspofungin. The result is that the active agent posaconazole is potentiating the activity of caspofungin. Another possibility is that the effect of caspofungin interfering with the structure of the cell wall may facilitate the penetration of posaconazole into the cell and make this agent more active.
However, the clinical value of these in vitro data is unknown. In vitro-in vivo correlation is needed for this combination.
Acknowledgments
We thank Thomas O'Boyle for help in the translation of the article.
This study does not present any conflict of interest for its authors.
This study was partially financed by grants from CIBER RES CD06/06/0058 and from Fundación Mutua Madrileña. Jesús Guinea is under contract (no. CM05/00171) by the Fondo de Investigación Sanitaria (FIS).
Footnotes
Published ahead of print on 18 June 2007.
REFERENCES
- 1.Cacciapuoti, A., J. Halpern, C. Mendrick, C. Norris, R. Patel, and D. Loebenberg. 2006. Interaction between posaconazole and caspofungin in concomitant treatment of mice with systemic Aspergillus infection. Antimicrob. Agents Chemother. 50:2587-2590. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 2.Dannaoui, E., J. Afeltra, J. F. G. M. Meis, P. E. Verweij, and The Eurofung Network. 2002. In vitro susceptibilities of zygomycetes to combinations of antimicrobial agents. Antimicrob. Agents Chemother. 46:2708-2711. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Ibrahim, A. S., J. C. Bowman, V. Avanessian, K. Brown, B. Spellberg, J. E. Edwards, Jr., and C. M. Douglas. 2005. Caspofungin inhibits Rhizopus oryzae 1,3-β-d-glucan synthase, lowers burden in brain measured by quantitative PCR, and improves survival at a low but not a high dose during murine disseminated zygomycosis. Antimicrob. Agents Chemother. 49:721-727. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4.Jacobs, P., L. Wood, A. Du Toit, and K. Esterhuizen. 2003. Eradication of invasive mucormycosis. Effectiveness of the echinocandin FK463. Hematology 8:119-123. [DOI] [PubMed] [Google Scholar]
- 5.Mukherjee, P. K., D. J. Sheehan, C. A. Hitchcock, and M. A. Ghannoum. 2005. Combination treatment of invasive fungal infections. Clin. Microbiol. Rev. 18:163-194. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Odds, F. C. 2003. Synergy, antagonism, and what the chequerboard puts between them. J. Antimicrob. Chemother. 52:1. [DOI] [PubMed] [Google Scholar]
- 7.Oliveira, E. R., A. W. Fothergill, W. R. Kirkpatrick, B. J. Coco, T. F. Patterson, and S. W. Redding. 2005. In vitro interaction of posaconazole and caspofungin against clinical isolates of Candida glabrata. Antimicrob. Agents Chemother. 49:3544-3545. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8.Richardson, M. D., and G. S. Shankland. 1999. Rhizopus, Rhizomucor, Absidia, and other agents of systemic and subcutaneous zygomycoses, p. 1242-1258. In P. R. Murray, E. J. Baron, M. A. Pfaller, F. C. Tenover, and R. H. Yolken (ed.), Manual of clinical microbiology, 7th ed. ASM Press, Washington, DC.
- 9.Spellberg, B., Y. Fu, J. E. Edwards, Jr., and A. S. Ibrahim. 2005. Combination therapy with amphotericin B lipid complex and caspofungin acetate of disseminated zygomycosis in diabetic ketoacidotic mice. Antimicrob. Agents Chemother. 49:830-832. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 10.Torres-Narbona, M., J. Guinea, J. Martínez-Alarcón, T. Peláez, and E. Bouza. 2007. In vitro activities of amphotericin B, caspofungin, itraconazole, posaconazole, and voriconazole against 45 clinical isolates of zygomycetes: comparison of CLSI M38-A, Sensititre YeastOne, and the Etest. Antimicrob. Agents Chemother. 51:1126-1129. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 11.Voitl, P., C. Scheibenpflug, T. Weber, O. Janata, and A. M. Rokitansky. 2002. Combined antifungal treatment of visceral mucormycosis with caspofungin and liposomal amphotericin B. Eur. J. Clin. Microbiol. Infect. Dis. 21:632-634. [DOI] [PubMed] [Google Scholar]