A Combination Approach to Treating Fungal Infections

Azoles are antifungal drugs used to treat fungal infections such as candidiasis in humans. Their extensive use has led to the emergence of drug resistance, complicating antifungal therapy for yeast infections in critically ill patients. Combination therapy has become popular in clinical practice as a potential strategy to fight resistant fungal isolates. Recently, amphiphilic tobramycin analogues, C12 and C14, were shown to display antifungal activities. Herein, the antifungal synergy of C12 and C14 with four azoles, fluconazole (FLC), itraconazole (ITC), posaconazole (POS), and voriconazole (VOR), was examined against seven Candida albicans strains. All tested strains were synergistically inhibited by C12 when combined with azoles, with the exception of C. albicans 64124 and MYA-2876 by FLC and VOR. Likewise, when combined with POS and ITC, C14 exhibited synergistic growth inhibition of all C. albicans strains, except C. albicans MYA-2876 by ITC. The combinations of FLC-C14 and VOR-C14 showed synergistic antifungal effect against three C. albicans and four C. albicans strains, respectively. Finally, synergism between C12/C14 and POS were confirmed by time-kill and disk diffusion assays. These results suggest the possibility of combining C12 or C14 with azoles to treat invasive fungal infections at lower administration doses or with a higher efficiency

Multifunctional Donepezil Analogues as Cholinesterase and BACE1 Inhibitors

A series of 22 donepezil analogues were synthesized through alkylation/benzylation and compared to donepezil and its 6-O-desmethyl adduct. All the compounds were found to be potent inhibitors of both acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), two enzymes responsible for the hydrolysis of the neurotransmitter acetylcholine in Alzheimer’s disease patient brains. Many of them displayed lower inhibitory concentrations of EeAChE (IC50 = 0.016 ± 0.001 µM to 0.23 ± 0.03 µM) and EfBChE (IC50 = 0.11 ± 0.01 µM to 1.3 ± 0.2 µM) than donepezil. One of the better compounds was tested against HsAChE and was found to be even more active than donepezil and inhibited HsAChE better than EeAChE. The analogues with the aromatic substituents were generally more potent than the ones with aliphatic substituents. Five of the analogues also inhibited the action of β-secretase (BACE1) enzyme

Amphiphilic Tobramycin Analogues as Antibacterial and Antifungal Agents

In this study, we investigated the in vitro antifungal activities, cytotoxicities, and membrane-disruptive actions of amphiphilic tobramycin (TOB) analogues. The antifungal activities were established by determination of MIC values and in time-kill studies. Cytotoxicity was evaluated in mammalian cell lines. The fungal membrane-disruptive action of these analogues was studied by using the membrane-impermeable dye propidium iodide. TOB analogues bearing a linear alkyl chain at their 6″-position in a thioether linkage exhibited chain length-dependent antifungal activities. Analogues with C12 and C14 chains showed promising antifungal activities against tested fungal strains, with MIC values ranging from 1.95 to 62.5 mg/liter and 1.95 to 7.8 mg/liter, respectively. However, C4, C6, and C8 TOB analogues and TOB itself exhibited little to no antifungal activity. Fifty percent inhibitory concentrations (IC50s) for the most potent TOB analogues (C12 and C14) against A549 and Beas 2B cells were 4- to 64-fold and 32- to 64-fold higher, respectively, than their antifungal MIC values against various fungi. Unlike conventional aminoglycoside antibiotics, TOB analogues with alkyl chain lengths of C12 and C14 appear to inhibit fungi by inducing apoptosis and disrupting the fungal membrane as a novel mechanism of action. Amphiphilic TOB analogues showed broad-spectrum antifungal activities with minimal mammalian cell cytotoxicity. This study provides novel lead compounds for the development of antifungal drugs

Membrane lipid-modulated mechanism of action and non-cytotoxicity of novel fungicide aminoglycoside FG08

A novel aminoglycoside, FG08, that differs from kanamycin B only by a C8 alkyl chain at the 4″-O position, was previously reported. Unlike kanamycin B, FG08 shows broad-spectrum fungicidal but not anti-bacterial activities. To understand its specificity for fungi, the mechanism of action of FG08 was studied using intact cells of the yeast Saccharomyces cerevisiae and small unilamellar membrane vesicles. With exposure to FG08 (30 µg mL−1), 8-fold more cells were stained with fluorescein isothiocyanate, cells had 4 to 6-fold higher K+ efflux rates, and 18-fold more cells were stained with SYTOX Green in comparison to exposure to kanamycin B (30 µg mL−1). Yeast mutants with aberrant membrane sphingolipids (no sphingoid base C4 hydroxyl group, truncated very long fatty acid chain, or lacking the terminal phosphorylinositol group of mannosyl-diinositolphosphorylphytoceramide were 4 to 8-fold less susceptible to growth inhibition with FG08 and showed 2 to 10-fold lower SYTOX Green dye uptake rates than did the isogenic wild-type strain. FG08 caused leakage of pre-loaded calcein from 50% of small unilamellar vesicles with glycerophospholipid and sterol compositions that mimic the compositions of fungal plasma membranes. Less than 5 and 10% of vesicles with glycerophospholipid and sterol compositions that mimic bacterial and mammalian cell plasma membranes, respectively, showed calcein leakage. In tetrazolium dye cytotoxicity tests, mammalian cell lines NIH3T3 and C8161.9 showed FG08 toxicity at concentrations that were 10 to 20-fold higher than fungicidal minimal inhibitory concentrations. It is concluded that FG08’s growth inhibitory specificity for fungi lie in plasma membrane permeability changes involving mechanisms that are modulated by membrane lipid composition

Subcutaneous administration of TC007 reduces disease severity in an animal model of SMA

Background Spinal Muscular Atrophy (SMA) is the leading genetic cause of infantile death. It is caused by the loss of functional Survival Motor Neuron 1 (SMN1). There is a nearly identical copy gene, SMN2, but it is unable to rescue from disease due to an alternative splicing event that excises a necessary exon (exon 7) from the majority of SMN2-derived transcripts. While SMNΔ7 protein has severely reduced functionality, the exon 7 sequences may not be specifically required for all activities. Therefore, aminoglycoside antibiotics previously shown to suppress stop codon recognition and promote translation read-through have been examined to increase the length of the SMNΔ7 C-terminus. Results Here we demonstrate that subcutaneous-administration of a read-through inducing compound (TC007) to an intermediate SMA model (Smn-/-; SMN2+/+; SMNΔ7) had beneficial effects on muscle fiber size and gross motor function. Conclusion Delivery of the read-through inducing compound TC007 reduces the disease-associated phenotype in SMA mice, however, does not significantly extend survival

Differential Effects of Linkers on the Activity of Amphiphilic Tobramycin Antifungals

As the threat associated with fungal infections continues to rise and the availability of antifungal drugs remains a concern, it becomes obvious that the need to bolster the antifungal armamentarium is urgent. Building from our previous findings of tobramycin (TOB) derivatives with antifungal activity, we further investigate the effects of various linkers on the biological activity of these aminoglycosides. Herein, we analyze how thioether, sulfone, triazole, amide, and ether functionalities affect the antifungal activity of alkylated TOB derivatives against 22 Candida, Cryptococcus, and Aspergillus species. We also evaluate their impact on the hemolysis of murine erythrocytes and the cytotoxicity against mammalian cell lines. While the triazole linker appears to confer optimal activity overall, all of the linkers incorporated into the TOB derivatives resulted in compounds that are very effective against the Cryptococcus neoformans species, with MIC values ranging from 0.48 to 3.9 μg/mL

Zinc Transporters YbtX and ZnuABC Are Required for the Virulence of \u3cem\u3eYersinia pestis\u3c/em\u3e in Bubonic and Pneumonic Plague in Mice

A number of bacterial pathogens require the ZnuABC Zinc (Zn2+) transporter and/or a second Zn2+ transport system to overcome Zn2+ sequestration by mammalian hosts. Previously we have shown that in addition to ZnuABC, Yersinia pestis possesses a second Zn2+ transporter that involves components of the yersiniabactin (Ybt), siderophore-dependent iron transport system. Synthesis of the Ybt siderophore and YbtX, a member of the major facilitator superfamily, are both critical components of the second Zn2+ transport system. Here we demonstrate that a ybtX znu double mutant is essentially avirulent in mouse models of bubonic and pneumonic plague while a ybtX mutant retains high virulence in both plague models. While sequestration of host Zn is a key nutritional immunity factor, excess Zn appears to have a significant antimicrobial role in controlling intracellular bacterial survival. Here, we demonstrate that ZntA, a Zn2+ exporter, plays a role in resistance to Zn toxicity in vitro, but that a zntA zur double mutant retains high virulence in both pneumonic and bubonic plague models and survival in macrophages. We also confirm that Ybt does not directly bind Zn2+in vitro under the conditions tested. However, we detect a significant increase in Zn2+-binding ability of filtered supernatants from a Ybt+ strain compared to those from a strain unable to produce the siderophore, supporting our previously published data that Ybt biosynthetic genes are involved in the production of a secreted Zn-binding molecule (zincophore). Our data suggest that Ybt or a modified Ybt participate in or promote Zn-binding activity in culture supernatants and is involved in Zn acquisition in Y. pestis

Multifunctional Donepezil Analogues as Cholinesterase and BACE1 Inhibitors

A series of 22 donepezil analogues were synthesized through alkylation/benzylation and compared to donepezil and its 6-O-desmethyl adduct. All the compounds were found to be potent inhibitors of both acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), two enzymes responsible for the hydrolysis of the neurotransmitter acetylcholine in Alzheimer’s disease patient brains. Many of them displayed lower inhibitory concentrations of EeAChE (IC50 = 0.016 ± 0.001 µM to 0.23 ± 0.03 µM) and EfBChE (IC50 = 0.11 ± 0.01 µM to 1.3 ± 0.2 µM) than donepezil. One of the better compounds was tested against HsAChE and was found to be even more active than donepezil and inhibited HsAChE better than EeAChE. The analogues with the aromatic substituents were generally more potent than the ones with aliphatic substituents. Five of the analogues also inhibited the action of β-secretase (BACE1) enzyme

Quantitative structure activity relationship analysis of antibacterial cationic anthraquinone analogs using Hansch and Fujita models

A quantitative structure–activity relationship (QSAR) of a library of eighteen cationic anthraquinone analogs was done using the Hansch and Fujita models. This offers a clear understanding of the structural parameter that could be used to explain the antibacterial activity of this class of compounds. The physicochemical parameters taken into consideration here were the Mlog P, partial atomic charge, dipole moment, and the HOMO. These parameters correlate well (q2 \u3e 0.8) with the observed antibacterial activity which provides valuable guidelines for new structural designs to further improve the antibacterial activity