scholarly journals Whole-Genome Approach to Understanding the Mechanism of Action of a Histatin 5-Derived Peptide

2019 ◽  
Vol 64 (3) ◽  
Author(s):  
Cody B. Bullock ◽  
David S. McNabb ◽  
Inés Pinto
Keyword(s):  
Mechanism Of Action ◽  
Fungal Infections ◽  
Antifungal Drugs ◽  
Content Type ◽  
Histatin 5 ◽  
Amino Acid Peptide ◽  
Killing Activity ◽  
Transport Chain ◽  
Mitochondrial Functions ◽  
Increased Sensitivity

ABSTRACT The incidence of opportunistic fungal infections that threaten immunocompromised patients, along with the limited arsenal of antifungal drugs, calls for renewed efforts to develop novel antifungal therapies. Antimicrobial peptides have garnered interest as potential therapeutics. Among naturally occurring peptides, histatin 5 is a well-characterized 24-amino-acid peptide with strong antifungal activity. Our lab has identified a smaller histatin derivative, KM29, with stronger activity against multiple Candida spp., prompting us to investigate its fungicidal mechanism. A genetic screen was developed to test the Saccharomyces cerevisiae genomewide deletion collection for mutants with increased or decreased peptide sensitivity. The goal was to identify genes that would reveal insights into the mechanism of action of KM29, to be assessed in Candida albicans. Several biological processes yielded increased sensitivity, with endosomal transport and vacuolar function appearing at high frequencies. Among the pathways involved in increased resistance, mitochondrial function showed the highest normalized genome frequency; hence, we focused on characterizing this pathway. KM29 localizes to mitochondria, and the killing activity depends on a functional electron transport chain. In addition, KM29 triggered reactive oxygen species (ROS) production, which was responsible for some cell death but insufficient to account for the complete killing activity. In agreement with this finding, we found that KM29 induced mitochondrial fragmentation and a mild loss of mitochondrial membrane potential. Furthermore, respiratory mutants exhibited severely diminished KM29 uptake. We confirmed this behavior in a C. albicans respiratory mutant. Taking our findings together, this work delineates the mitochondrial functions associated with KM29 fungicidal activity and provides additional pathways for further characterization in Candida spp.

scholarly journals Derivatives of the Antimalarial Drug Mefloquine Are Broad-Spectrum Antifungal Molecules with Activity against Drug-Resistant Clinical Isolates

2020 ◽  
Vol 64 (3) ◽  
Author(s):  
Marhiah C. Montoya ◽  
Sarah Beattie ◽  
Kathryn M. Alden ◽  
Damian J. Krysan
Keyword(s):  
Antifungal Activity ◽  
Mechanism Of Action ◽  
Broad Spectrum ◽  
Antimalarial Drug ◽  
Fungal Infections ◽  
Antifungal Drugs ◽  
Clinical Isolates ◽  
Content Type ◽  
Recent Emergence ◽  
Derivatives Of

ABSTRACT The antifungal pharmacopeia is critically small, particularly in light of the recent emergence of multidrug-resistant pathogens, such as Candida auris. Here, we report that derivatives of the antimalarial drug mefloquine have broad-spectrum antifungal activity against pathogenic yeasts and molds. In addition, the mefloquine derivatives have activity against clinical isolates that are resistant to one or more of the three classes of antifungal drugs currently used to treat invasive fungal infections, indicating that they have a novel mechanism of action. Importantly, the in vitro toxicity profiles obtained using human cell lines indicated that the toxicity profiles of the mefloquine derivatives are very similar to those of the parent mefloquine, despite being up to 64-fold more active against fungal cells. In addition to direct antifungal activity, subinhibitory concentrations of the mefloquine derivatives inhibited the expression of virulence traits, including filamentation in Candida albicans and capsule formation/melanization in Cryptococcus neoformans. Mode/mechanism-of-action experiments indicated that the mefloquine derivatives interfere with both mitochondrial and vacuolar function as part of a multitarget mechanism of action. The broad-spectrum scope of activity, blood-brain barrier penetration, and large number of previously synthesized analogs available combine to support the further optimization and development of the antifungal activity of this general class of drug-like molecules.

scholarly journals The Phosphoenolpyruvate:Sugar Phosphotransferase System Is Involved in Sensitivity to the Glucosylated Bacteriocin Sublancin

2015 ◽  
Vol 59 (11) ◽  
pp. 6844-6854 ◽  
Author(s):  
C. V. Garcia De Gonzalo ◽  
E. L. Denham ◽  
R. A. T. Mars ◽  
J. Stülke ◽  
W. A. van der Donk ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Mechanism Of Action ◽  
Mode Of Action ◽  
Phosphotransferase System ◽  
Content Type ◽  
Gram Positive Bacteria ◽  
Distinct Mechanism ◽  
Genomic Studies ◽  
Increased Sensitivity ◽  
Bacterial Sensitivity

ABSTRACTThe mode of action of a group of glycosylated antimicrobial peptides known as glycocins remains to be elucidated. In the current study of one glycocin, sublancin, we identified the phosphoenolpyruvate:sugar phosphotransferase system (PTS) ofBacillusspecies as a key player in bacterial sensitivity. Sublancin kills several Gram-positive bacteria, such asBacillusspecies andStaphylococcus aureus, including methicillin-resistantS. aureus(MRSA). Unlike other classes of bacteriocins for which the PTS is involved in their mechanism of action, we show that the addition of PTS-requiring sugars leads to increased resistance rather than increased sensitivity, suggesting that sublancin has a distinct mechanism of action. Collectively, our present mutagenesis and genomic studies demonstrate that the histidine-containing phosphocarrier protein (HPr) and domain A of enzyme II (PtsG) in particular are critical determinants for bacterial sensitivity to sublancin.

scholarly journals Transcription Factor ADS-4 Regulates Adaptive Responses and Resistance to Antifungal Azole Stress

2015 ◽  
Vol 59 (9) ◽  
pp. 5396-5404 ◽  
Author(s):  
Kangji Wang ◽  
Zhenying Zhang ◽  
Xi Chen ◽  
Xianyun Sun ◽  
Cheng Jin ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Efflux Pump ◽  
Fungal Infections ◽  
Antifungal Drugs ◽  
Bzip Transcription Factor ◽  
Azole Resistance ◽  
Homologous Gene ◽  
Adaptive Responses ◽  
Transcriptional Responses ◽  
Content Type

ABSTRACTAzoles are commonly used as antifungal drugs or pesticides to control fungal infections in medicine and agriculture. Fungi adapt to azole stress by rapidly activating the transcription of a number of genes, and transcriptional increases in some azole-responsive genes can elevate azole resistance. The regulatory mechanisms that control transcriptional responses to azole stress in filamentous fungi are not well understood. This study identified a bZIP transcription factor, ADS-4 (antifungaldrugsensitive-4), as a new regulator of adaptive responses and resistance to antifungal azoles. Transcription ofads-4inNeurospora crassacells increased when they were subjected to ketoconazole treatment, whereas the deletion ofads-4resulted in hypersensitivity to ketoconazole and fluconazole. In contrast, the overexpression ofads-4increased resistance to fluconazole and ketoconazole inN. crassa. Transcriptome sequencing (RNA-seq) analysis, followed by quantitative reverse transcription (qRT)-PCR confirmation, showed that ADS-4 positively regulated the transcriptional responses of at least six genes to ketoconazole stress inN. crassa. The gene products of four ADS-4-regulated genes are known contributors to azole resistance, including the major efflux pump CDR4 (Pdr5p ortholog), an ABC multidrug transporter (NcAbcB), sterol C-22 desaturase (ERG5), and a lipid transporter (NcRTA2) that is involved in calcineurin-mediated azole resistance. Deletion of theads-4-homologous gene Afads-4inAspergillus fumigatuscaused hypersensitivity to itraconazole and ketoconazole, which suggested that ADS-4 is a functionally conserved regulator of adaptive responses to azoles. This study provides important information on a new azole resistance factor that could be targeted by a new range of antifungal pesticides and drugs.

scholarly journals Acylhydrazones as Antifungal Agents Targeting the Synthesis of Fungal Sphingolipids

2018 ◽  
Vol 62 (5) ◽  
Author(s):  
Cristina Lazzarini ◽  
Krupanandan Haranahalli ◽  
Robert Rieger ◽  
Hari Krishna Ananthula ◽  
Pankaj B. Desai ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Antifungal Agents ◽  
Fungal Infections ◽  
Antifungal Drugs ◽  
Fungal Resistance ◽  
Content Type ◽  
Highly Active ◽  
Pharmacokinetic Properties ◽  
Pass Through

ABSTRACTThe incidence of invasive fungal infections has risen dramatically in recent decades. Current antifungal drugs are either toxic, likely to interact with other drugs, have a narrow spectrum of activity, or induce fungal resistance. Hence, there is a great need for new antifungals, possibly with novel mechanisms of action. Previously our group reported an acylhydrazone called BHBM that targeted the sphingolipid pathway and showed strong antifungal activity against several fungi. In this study, we screened 19 derivatives of BHBM. Three out of 19 derivatives were highly active againstCryptococcus neoformansin vitroand had low toxicity in mammalian cells. In particular, one of them, called D13, had a high selectivity index and showed better activity in an animal model of cryptococcosis, candidiasis, and pulmonary aspergillosis. D13 also displayed suitable pharmacokinetic properties and was able to pass through the blood-brain barrier. These results suggest that acylhydrazones are promising molecules for the research and development of new antifungal agents.

scholarly journals Candida albicans Cek1 Mitogen-Activated Protein Kinase Signaling Enhances Fungicidal Activity of Salivary Histatin 5

2015 ◽  
Vol 59 (6) ◽  
pp. 3460-3468 ◽  
Author(s):  
Rui Li ◽  
Sumant Puri ◽  
Swetha Tati ◽  
Paul J. Cullen ◽  
Mira Edgerton
Keyword(s):  
Candida Albicans ◽  
Protein Kinase ◽  
Antifungal Drugs ◽  
Mitogen Activated Protein Kinase ◽  
Fungal Cell ◽  
Content Type ◽  
Histatin 5 ◽  
Mitogen Activated Protein ◽  
Hyphal Formation ◽  
Sensor Proteins

ABSTRACTCandida albicansis a major etiological organism for oropharyngeal candidiasis (OPC), while salivary histatin 5 (Hst 5) is a human fungicidal protein that protects the oral cavity from OPC.C. albicanssenses its environment by mitogen-activated protein kinase (MAPK) activation that can also modulate the activity of some antifungal drugs, including Hst 5. We found that phosphorylation of the MAPK Cek1, induced either byN-acetylglucosamine (GlcNAc) or serum, or its constitutive activation by deletion of its phosphatase Cpp1 elevated the susceptibility ofC. albicanscells to Hst 5. Cek1 phosphorylation but not hyphal formation was needed for increased Hst 5 sensitivity. Interference with the Cek1 pathway by deletion of its head sensor proteins, Msb2 and Sho1, or by addition of secreted aspartyl protease (SAP) cleavage inhibitors, such as pepstatin A, reduced Hst 5 susceptibility under Cek1-inducing conditions. Changes in fungal cell surface glycostructures also modulated Hst 5 sensitivity, and Cek1-inducing conditions resulted in a higher uptake rate of Hst 5. These results show that there is a consistent relationship between activation of Cek1 MAPK and increased Hst 5 susceptibility inC. albicans.

First Line Antifungal Treatment with Caspofungin of Oncohemopatic Patients. A Single Centre Experience.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 5271-5271 ◽  
Author(s):  
Alessandro Bonini ◽  
Alessia Tieghi ◽  
Luigi Gugliotta
Keyword(s):  
Adverse Events ◽  
Fungal Infection ◽  
Ct Scan ◽  
Mechanism Of Action ◽  
Fungal Infections ◽  
Lymphoblastic Leukemia ◽  
Antifungal Drugs ◽  
Severe Neutropenia ◽  
First Line ◽  
The Mean

Abstract Infections are the main complication for patients (pts) with hematologic diseases and severe neutropenia and among them fungal infections are the most difficult to treat and a major cause of mortality. The availability of a new class of antifungal drugs (echinocandins) could improve the chance of cure. Caspofungin (Caspo) is the first drug which is able to destroy the fungal cell wall. Since January 2004 we have treated 28 consecutive adult oncohemopatic and neutropenic pts with Caspo as first line therapy. In case of persistent fever (4 days) despite broad spectrum antibiotic therapy (association of Tazobactam/Piperacillin, Amikacin with or without Vancomycin) with negative blood cultures, a high-resolution CT-scan of the lungs, an abdomen US-scan, swabs from pharynx, nose and rectum, galactomannan test (this test is available at our Centre since February 2006) were performed. In the presence of any other sign or symptom we performed any other test according to the physicyan’s choice. In case of possible, probable or proven fungal infection (according to the EORTC criteria) Caspo was administered at the dosage of 70 mg i.v. on the first day followed by 50 mg i.v. in 1 hour daily. The pts were 15 males and 13 females; the mean age was 46 yrs (range 18–66 yrs). The diagnoses were: acute myeloid leukemia 13, acute lymphoblastic leukemia 5, multiple myeloma 2, lymphoma 8; the disease’s phases were: onset 9, complete remission 11, relapse 2, resistant 6. Six pts received an allogeneic and 4 an autologous BMT; the other pts received an induction or consolidation or rescue chemotherapy course. All the pts had severe neutropenia and the fungal infections were proven in 3 cases (2 aspergillus spp and 1 aspergillus fumigatus), probable in 3 cases and possible in 22 cases. The first site of infection was the lung in 27 pts and paranasal sinuses in 1 patient. CT scan was positive (halo sign, air-crescent sign or cavitation) in all the pts with a lung localization. The mean time of treatment was 18 days (range 6–21 days). The treatment was not discontinued for anyone because of adverse events and no modifications of the dosage were necessary. All the pts submitted to an allogeneic BMT received concomitant therapy with Cyclosporine A and we had not to change the dosage and we did not found any renal or liver alterations. No adverse events during the infusion of Caspo were seen and it was not necessary to administer any drug before the infusion as premedication. No breakthrough fungal infections were found. The infection was cured in 24/28 pts; 4 pts died for fungal infection progression (3 with a progression to the brain and in 1 case the infection remained in the lungs). For all the cured pts there was a concomitant recovery of neutrophils so also in our experience this appears to be a crucial fact for the resolution of the infection. Among the 24 cured patients 8 died later: 5 for hematologic disease and 3 for sepsis during malignant disease recurrence. In 2 cases there was the recurrence of the fungal infection despite the secondary prophylaxis with Caspo. In conclusion we can say we have a new treatment option for fungal infections in neutropenic pts with a new mechanism of action; this option seems safe, it does not preclude any other treatment (such as Cyclosporine), it is well tolerated and the resolution rate of the infections is very high, probably because of the new mechanism of action of the drug. Moreover the cost is lower than other antifungal treatments.

scholarly journals Screening of Chemical Libraries for New Antifungal Drugs against Aspergillus fumigatus Reveals Sphingolipids Are Involved in the Mechanism of Action of Miltefosine

mBio ◽  
2021 ◽  
Author(s):  
Thaila Fernanda dos Reis ◽  
Maria Augusta Crivelente Horta ◽  
Ana Cristina Colabardini ◽  
Caroline Mota Fernandes ◽  
Lilian Pereira Silva ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Mechanism Of Action ◽  
Filamentous Fungus ◽  
Antifungal Drugs ◽  
Clinical Isolates ◽  
Content Type ◽  
Chemical Libraries

The filamentous fungus Aspergillus fumigatus causes a group of diseases named aspergillosis, and their development occurs after the inhalation of conidia dispersed in the environment. Very few classes of antifungal drugs are available for aspergillosis treatment, e.g., azoles, but the emergence of global resistance to azoles in A. fumigatus clinical isolates has increased over recent decades.

scholarly journals Synergistic Effects of Amiodarone and Fluconazole on Candida tropicalis Resistant to Fluconazole

2013 ◽  
Vol 57 (4) ◽  
pp. 1691-1700 ◽  
Author(s):  
Cecília Rocha da Silva ◽  
João Batista de Andrade Neto ◽  
José Júlio Costa Sidrim ◽  
Maria Rozzelê Ferreira Ângelo ◽  
Hemerson Iury Ferreira Magalhães ◽  
...  
Keyword(s):  
Candida Tropicalis ◽  
Inhibitory Concentration ◽  
Fungal Infections ◽  
Synergistic Effects ◽  
Antifungal Drugs ◽  
Cross Resistance ◽  
Combination Drug ◽  
Content Type ◽  
Effective Strategies ◽  
Drug Of Choice

ABSTRACTThere have recently been significant increases in the prevalence of systemic invasive fungal infections. However, the number of antifungal drugs on the market is limited in comparison to the number of available antibacterial drugs. This fact, coupled with the increased frequency of cross-resistance, makes it necessary to develop new therapeutic strategies. Combination drug therapies have become one of the most widely used and effective strategies to alleviate this problem. Amiodarone (AMD) is classically used for the treatment of atrial fibrillation and is the drug of choice for patients with arrhythmia. Recent studies have shown broad antifungal activity of the drug when administered in combination with fluconazole (FLC). In the present study, we induced resistance to fluconazole in six strains ofCandida tropicalisand evaluated potential synergism between fluconazole and amiodarone. The evaluation of drug interaction was determined by calculating the fractional inhibitory concentration and by performing flow cytometry. We conclude that amiodarone, when administered in combination with fluconazole, exhibits activity against strains ofC. tropicalisthat are resistant to fluconazole, which most likely occurs via changes in the integrity of the yeast cell membrane and the generation of oxidative stress, mitochondrial dysfunction, and DNA damage that could lead to cell death by apoptosis.

scholarly journals Analytical and Clinical Evaluation of the PathoNostics AsperGenius Assay for Detection of Invasive Aspergillosis and Resistance to Azole Antifungal Drugs Directly from Plasma Samples

2017 ◽  
Vol 55 (8) ◽  
pp. 2356-2366 ◽  
Author(s):  
P. Lewis White ◽  
Raquel B. Posso ◽  
Rosemary A. Barnes
Keyword(s):  
Clinical Study ◽  
Invasive Aspergillosis ◽  
Clinical Performance ◽  
Fungal Infections ◽  
Antifungal Drugs ◽  
European Organization ◽  
Content Type ◽  
Enhanced Sensitivity ◽  
Resistant Disease ◽  
Single Positive

ABSTRACT With the proposal to include Aspergillus PCR in the revised European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) definitions for fungal disease, commercially manufactured assays may be required to provide standardization and accessibility. The PathoNostics AsperGenius assay represents one such test that has the ability to detect a range of Aspergillus species as well as azole resistance in Aspergillus fumigatus . Its performance has been validated on bronchoalveolar lavage (BAL) fluid and serum specimens, but recent evidence suggests that testing of plasma may have enhanced sensitivity over that with serum. We decided to evaluate the analytical and clinical performances of the PathoNostics AsperGenius assay for testing of plasma. For the analytical evaluations, plasma was spiked with various concentrations of Aspergillus genomic DNA before extraction following international recommendations, using two automated platforms. For the clinical study, 211 samples from 10 proven/probable invasive aspergillosis (IA) and 2 possible IA cases and 27 controls were tested. The limits of detection for testing of DNA extracted using the bioMérieux EasyMag and Qiagen EZ1 extractors were 5 and 10 genomes/0.5-ml sample, respectively. In the clinical study, true positivity was significantly greater than false positivity ( P < 0.0001). The sensitivity and specificity obtained using a single positive result as significant were 80% and 77.8%, respectively. If multiple samples were required to be positive, specificity was increased to 100%, albeit sensitivity was reduced to 50%. The AsperGenius assay provided good clinical performance, but the predicted improvement of testing with plasma was not seen, possibly as a result of target degradation attributed to sample storage. Prospective testing is required to determine the clinical utility of this assay, particularly for the diagnosis of azole-resistant disease.

scholarly journals Amino Acid-Derived 1,2-Benzisothiazolinone Derivatives as Novel Small-Molecule Antifungal Inhibitors: Identification of Potential Genetic Targets

2012 ◽  
Vol 56 (9) ◽  
pp. 4630-4639 ◽  
Author(s):  
Deepu Alex ◽  
Francoise Gay-Andrieu ◽  
Jared May ◽  
Linta Thampi ◽  
Dengfeng Dou ◽  
...  
Keyword(s):  
Amino Acid ◽  
Fungal Pathogens ◽  
Structural Similarity ◽  
Homozygous Deletion ◽  
Antifungal Drugs ◽  
Respiratory Pathway ◽  
Phenotypic Data ◽  
Content Type ◽  
Mitochondrial Functions ◽  
Phenotype Analysis

ABSTRACTWe have identified four synthetic compounds (DFD-VI-15, BD-I-186, DFD-V-49, and DFD-V-66) from an amino acid-derived 1,2-benzisothiazolinone (BZT) scaffold that have reasonable MIC50values against a panel of fungal pathogens. These compounds have no structural similarity to existing antifungal drugs. Three of the four compounds have fungicidal activity againstCandidaspp.,Cryptococcus neoformans, and several dermatophytes, while one is fungicidal toAspergillus fumigatus. The kill rates of our compounds are equal to those in clinical usage. The BZT compounds remain active against azole-, polyene-, and micafungin-resistant strains ofCandidaspp. A genetics-based approach, along with phenotype analysis, was used to begin mode of action (MOA) studies of one of these compounds, DFD-VI-15. The genetics-based screen utilized a homozygous deletion collection of approximately 4,700Saccharomyces cerevisiaemutants. We identified mutants that are both hypersensitive and resistant. Using FunSpec, the hypersensitive mutants and a resistantace2mutant clustered within a category of genes related directly or indirectly to mitochondrial functions. InCandida albicans, the functions of the Ace2p transcription factor include the regulation of glycolysis. Our model is that DFD-VI-15 targets a respiratory pathway that limits energy production. Supporting this hypothesis are phenotypic data indicating that DFD-VI-15 causes increased cell-reactive oxidants (ROS) and a decrease in mitochondrial membrane potential. Also, the same compound has activity when cells are grown in a medium containing glycerol (mitochondrial substrate) but is much less active when cells are grown anaerobically.

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