Efflux Pump Mediated Azole Resistance in Candida Albicans

Study Overview
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Brief Summary
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Candida albicans is the major fungal pathogen causing infections in humans, ranging from superficial mucosal infection to systemic mycoses. In recent years, Candida infections have increased disproportionately as a result of the increased number of compromised host populations, such as patients with AIDS, diabetes and various cancers, and organ-transplant recipients. Severe oro-pharyngeal candidiasis afflicts many AIDS patients and is a significant infection in cancer patients being treated with chemotherapy and/or radiotherapy.

Detailed Description
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In cancer patients, the increased incidence of oro-pharyngeal candidiasis results both from the debilitating effects of the cancer itself and from the immuno-suppressive treatment for the cancer. Administration of broad-spectrum antibiotics for the management of bacterial infections in these patients may further predispose them to oro-pharyngeal candidiasis . Systemic fungal infections are often hard to diagnose, which contributes to their high attributable mortality. In addition, there are far fewer classes of antifungal agent than antibacterial drugs, limiting therapeutic options. The azole antifungals are commonly used to treat fungal infections, as they are conveniently administered and have few side effects . the number of drug-resistant Candida strains has also increased dramatically owing to the increased use of antifungal agents. The major mechanism responsible for high-level azole resistance in clinical Candida isolates is overexpression of plasma membrane efflux pumps . There are two main families of efflux proteins, the ATP-binding cassette pumps and the major facilitator superfamily transporters . Azole resistance calls for the use of alternative antifungal drugs like echinocandins, voriconazole, posaconazole, ravuconazole and Amphotericin B. But constraints like high costs or adverse effects associated with these agents often limit their usage. Some recent pioneering studies have reported the modulating effect of verapamil, oestradiol, progesterone and ibuprofen on resistance of Candida isolates. While fluconazole MIC decreased in most strains after exposure to these modulators, this effect was particularly remarkable for Ibuprofen. The molecular basis of this reversal of resistance has also been demonstrated recently. Thus, drugs capable of reversing fluconazole resistance might offer novel breakthroughs in the treatment of resistant Candida infections. As investigators have previously observed a high prevalence of fluconazole resistance among Candida isolates recovered in our centre. consequently ,implementing the concept of combination therapy using an efflux pump inhibitor seems an adequate strategy for overcoming resistance. in this study investigators were interested in exploring if the resistant phenotype could be reversed in a proportion of these isolates by the use of Ibuprofen .Expression levels of the target genes associated with antifungal resistance of C.albicans (CDR1, CDR2 and MDR1) were assessed by quantitative real-time The relative quantities of the target genes were normalized against ACT1 housekeeping gene expression and analysed using the comparative DDCt method, taking the amplification efficiency into consideration .

Official Title
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Efflux Pump Mediated Azole Resistance in Candida Albicans Among Neutropenic Patients With Haematological Malignancies

Conditions
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Fungal Infection

Intervention / Treatment
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* Drug: Azole Antifungal


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: Patients suffered from neutropenia. Patients with absolute neutrophiles count less than 500 cells / ml . 7 days or more from start of chemotherapy. Also neutropenic patients presented during relapse after first remission of hematological malignancies. Presence of resistance to at least one azole derivatives. Exclusion Criteria: Patient who have contraindication to Azoles
Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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No

Study Plan
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How is the study designed?
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Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| neutropenic cancer patient recieving azoles .<br>neutropenic cancer patients that undergoing chemotherapy with prophylaxis with azoles and exhibit recurrent fungal infection so appropriate clinical specimen will taken for isolation & identification of causative agent & it's antimicrobial profile then identification of the mechanism of resistance by invitro technique then confirmed by real time pcr. | Drug: Azole Antifungal<br>* oral gel & injections<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| characterize the clinical Candida albicans strains isolated from neutropenic patients with Hematological Malignancies. | isolation & identification of of Candida albicans by culturing on Sabouraud's Dextrose Agar (SDA) and CHROM Agar Candida then determination of antifungal susceptibility testing by Disk Diffusion test and determination of MIC by broth microdilution method to identify resistant Candida albicans strains to decrease hazard to these high risk group of patients. | 3 months | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Determination if Ibuprofen could elicit a reversal of fluconazole resistance by interference with efflux pump activity. | reverting of resistance to fluconazole using Ibuprofen as efflux pump inhibitor detected by redetermination of MIC values in presence of a subinhibitory concentration of Ibuprofen. | 2month | 
| Testing new NS-IDs other than Ibuprofen as efflux pump inhibitor | resistant Candida albicans strains will subject to broth microdilution in presence and absence of other NS-IDs. | 3 months | 
| Uncover the molecular base of antifungal resistance in C. albicans clinical strains that could be reverted by ibuprofen | quantification of gene expression of genes encoding for efflux pump (CDR1, CDR2 and MDR1) by real time pcrRT-PCR(q RT-PCR) The relative quantities of the target genes were normalized against ACT1 housekeeping gene expression and analysed using the comparative DDCt method, taking the amplification efficiency into consideration. | 4 months |