Articulo Microbiologia

Medical Mycology October 2003, 41, 417 Á/425

Antifungal activity of fluconazole in combination with lovastatin and their effects on gene expression in the ergosterol and prenylation pathways in Candida albicans
JIA L. SONG, CHRIS N. LYONS, SCOTT HOLLEMAN, BRIAN G. OLIVER & THEODORE C. WHITE* Department of Pathobiology, School of Public Health and Community Medicine, University of Washingtonand Seattle Biomedical Research Institute, Seattle, Washington, USA

The sterol pathway in Candida albicans is the target for several classes of antifungal drugs. Intermediates in the sterol pathway are involved in ergosterol synthesis, prenylation and dolichol synthesis. This study examines gene expression of the sterol pathway in response to lovastatin, an inhibitor of HMG-CoA reductase (Hmg1p),and fluconazole, an inhibitor of 14 a-lanosterol demethylase (Erg11p). Minimum inhibitory concentration (MIC) studies indicated that lovastatin acts synergistically with fluconazole in vitro. Semi-quantitative reverse transcriptaseÁ polymerase chain reaction (RT-PCR) results indicated that genes in the early part of the sterol pathway, such as HMG1 and ERG20, did not alter expression in thepresence of both lovastatin and fluconazole, whereas genes in the later part of the sterol pathway, such as ERG9 and ERG11, had increased expression in response to these drugs in mid-logarithmic growth. Genes involved in prenylation, such as RAM1 and RAM2, also respond to these drugs in mid-logarithmic growth, although another prenylation gene, CDC43, was not affected. After 24 h of growth, therelative expression of ERG20, ERG9, and ERG11 remained unchanged or increased in the presence of both drugs, while all other genes decreased in expression under all drug treatments.
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Keywords

Candida albicans, drugs, ergosterol, prenylation

Introduction
Candida albicans is an opportunistic pathogen that can cause oral, esophageal, and systemic infections in immunocompromised patients [1]. Itcan also cause vaginal infections in healthy women. The major drugs used to treat candidiasis include the fungicidal polyenes and fungistatic azoles. Both classes of drugs are directed against ergosterol, which is the main sterol of the fungal plasma membrane. Polyenes are effective antifungal drugs, but their use is limited by their toxicity and insolubility. Fluconazole is widely usedReceived August 2002; Final version received 31 December 2002 Correspondence: Theodore C. White, Seattle Biomedical Research Institute, 4 Nickerson Street, Suite 200, Seattle, WA 98109-1651. Tel.: (206) 284 8846 (ext. 344); Fax: (206) 284 0313. E-mail: tedwhite@u.washington.edu

for treatment because of its effectiveness, oral bioavailability and negligible toxicity. The emergence of azole drugresistance has been increasing, and it has been estimated that 20Á/30% of oral isolates from HIV patients are resistant to azoles [2]. The sterol pathway in C. albicans is a target for several classes of antifungal drugs in addition to polyenes and azoles. This study examines how drug inhibition of the sterol pathway may affect the regulation of its genes and of genes involved in other cellularprocesses. An understanding of the sterol pathway and how the fungal cell responds to various drugs may further our knowledge on the development of drug resistance in C. albicans. The intermediates in the sterol pathway are precursors for molecules in many important cellular processes, such as ergosterol biosynthesis, protein prenylation and dolichol synthesis [3]. The sterol pathway is initiated withacetyl-CoA and ends in ergosterol
DOI: 10.1080/1369378031000137233

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[4]. The conversion of acetyl-coenzyme A (CoA) to farnesyl diphosphate (FPP) is commonly referred to as the isoprenoid pathway, in which b-hydroxy- b-methylglutaryl(HMG)-CoA reductase is the major site of regulation [5]. The ERG20 gene encodes geranyl and farnesyl diphosphate synthase, which...