Mitochondrial Cytochrome Bc1 Complex Research Articles

Mitochondrial Cytochrome bc1 Complex as Validated Drug Target: A Structural Perspective.

Mitochondrial respiratory chain Complex III, also known as cytochrome bc1 complex or cyt bc1, is a validated target not only for antibiotics but also for pesticides and anti-parasitic drugs. Although significant progress has been made in understanding the mechanisms of cyt bc1 function and inhibition by using various natural and synthetic compounds, important issues remain in overcoming drug resistance in agriculture and in evading cytotoxicity in medicine. In this review, we look at these issues from a structural perspective. After a brief description of the essential and common structural features, we point out the differences among various cyt bc1 complexes of different organisms, whose structures have been determined to atomic resolution. We use a few examples of cyt bc1 structures determined via bound inhibitors to illustrate both conformational changes observed and implications to the Q-cycle mechanism of cyt bc1 function. These structures not only offer views of atomic interactions between cyt bc1 complexes and inhibitors, but they also provide explanations for drug resistance when structural details are coupled to sequence changes. Examples are provided for exploiting structural differences in evolutionarily conserved enzymes to develop antifungal drugs for selectivity enhancement, which offer a unique perspective on differential interactions that can be exploited to overcome cytotoxicity in treating human infections.

Cryo-EM structure of the four-subunit Rhodobacter sphaeroides cytochrome bc1 complex in styrene maleic acid nanodiscs

Cytochrome bc1 complexes are ubiquinol:cytochrome c oxidoreductases, and as such, they are centrally important components of respiratory and photosynthetic electron transfer chains in many species of bacteria and in mitochondria. The minimal complex has three catalytic components, which are cytochrome b, cytochrome c1, and the Rieske iron-sulfur subunit, but the function of mitochondrial cytochrome bc1 complexes is modified by up to eight supernumerary subunits. The cytochrome bc1 complex from the purple phototrophic bacterium Rhodobactersphaeroides has a single supernumerary subunit called subunit IV, which is absent from current structures of the complex. In this work we use the styrene-maleic acid copolymer to purify the R. sphaeroides cytochrome bc1 complex in native lipid nanodiscs, which retains the labile subunit IV, annular lipids, and natively bound quinones. The catalytic activity of the four-subunit cytochrome bc1 complex is threefold higher than that of the complex lacking subunit IV. To understand the role of subunit IV, we determined the structure of the four-subunit complex at 2.9 Å using single particle cryogenic electron microscopy. The structure shows the position of the transmembrane domain of subunit IV, which lies across the transmembrane helices of the Rieske and cytochrome c1 subunits. We observe a quinone at the Qo quinone-binding site and show that occupancy of this site is linked to conformational changes in the Rieske head domain during catalysis. Twelve lipids were structurally resolved, making contacts with the Rieske and cytochrome b subunits, with some spanning both of the two monomers that make up the dimeric complex.

Knockout of the Complex III subunit Uqcrh causes bioenergetic impairment and cardiac contractile dysfunction

Ubiquinol cytochrome c reductase hinge protein (UQCRH) is required for the electron transfer between cytochrome c1 and c of the mitochondrial cytochrome bc1 Complex (CIII). A two-exon deletion in the human UQCRH gene has recently been identified as the cause for a rare familial mitochondrial disorder. Deletion of the corresponding gene in the mouse (Uqcrh-KO) resulted in striking biochemical and clinical similarities including impairment of CIII, failure to thrive, elevated blood glucose levels, and early death. Here, we set out to test how global ablation of the murine Uqcrh affects cardiac morphology and contractility, and bioenergetics. Hearts from Uqcrh-KO mutant mice appeared macroscopically considerably smaller compared to wildtype littermate controls despite similar geometries as confirmed by transthoracic echocardiography (TTE). Relating TTE-assessed heart to body mass revealed the development of subtle cardiac enlargement, but histopathological analysis showed no excess collagen deposition. Nonetheless, Uqcrh-KO hearts developed pronounced contractile dysfunction. To assess mitochondrial functions, we used the high-resolution respirometer NextGen-O2k allowing measurement of mitochondrial respiratory capacity through the electron transfer system (ETS) simultaneously with the redox state of ETS-reactive coenzyme Q (Q), or production of reactive oxygen species (ROS). Compared to wildtype littermate controls, we found decreased mitochondrial respiratory capacity and more reduced Q in Uqcrh-KO, indicative for an impaired ETS. Yet, mitochondrial ROS production was not generally increased. Taken together, our data suggest that Uqcrh-KO leads to cardiac contractile dysfunction at 9 weeks of age, which is associated with impaired bioenergetics but not with mitochondrial ROS production.Graphical abstractGlobal ablation of the Uqcrh gene results in functional impairment of CIII associated with metabolic dysfunction and postnatal developmental arrest immediately after weaning from the mother. Uqcrh-KO mice show dramatically elevated blood glucose levels and decreased ability of isolated cardiac mitochondria to consume oxygen (O2). Impaired development (failure to thrive) after weaning manifests as a deficiency in the gain of body mass and growth of internal organ including the heart. The relative heart mass seemingly increases when organ mass calculated from transthoracic echocardiography (TTE) is normalized to body mass. Notably, the heart shows no signs of collagen deposition, yet does develop a contractile dysfunction reflected by a decrease in ejection fraction and fractional shortening.

Baseline sensitivity and control efficacy of a new QiI fungicide, florylpicoxamid, against Botrytis cinerea.

Gray mold caused by Botrytis cinerea is an airborne plant pathogen with a necrotrophic lifestyle that infects more than 200 crops worldwide. Florylpicoxamid is a second-generation picolinamide fungicide inspired by a natural product. Florylpicoxamid targets the Qi site of the mitochondrial cytochrome bc1 complex and is currently being registered in China for the control of gray mold in a variety of crops. Although a broad spectrum of activity and attributes have been reported for florylpicoxamid, little is known about its effectiveness against gray mold or its protective and curative properties. Florylpicoxamid exhibited substantial inhibitory activity against 12 tested species of plant-pathogenic fungi, with effective concentration for 50% growth inhibition (EC50 ) values ranging from 0.017 to 2.096 μg ml-1 . A total of 129 isolates of B. cinerea from ten regions were tested for their sensitivity to florylpicoxamid, and the mean EC50 value was 0.04 ± 0.017 μg ml-1 . Furthermore, florylpicoxamid was observed to substantially inhibit all developmental stages of B. cinerea, with mycelial development, sclerotium germination, germ tube elongation and conidial germination being restrained with an EC50 value of 0.051 ± 0.0072, 0.012 ± 0.0069, 0.019 ± 0.0041 and 0.0062 ± 0.0007 μg ml-1 , respectively. No cross-resistance was observed between florylpicoxamid and quinone outside inhibitor (QoI), methyl benzimidazole carbamates or succinate dehydrogenase inhibitor. Florylpicoxamid also exhibited protective and curative activity against the development of B. cinerea infection in tests on tomato fruits. At application rates of 90, 112.5 and 135 g a.i. ha-1 , florylpicoxamid was also observed to provide more-effective control than boscalid (300 g a.i. ha-1 ). This study demonstrated that the novel fungicide florylpicoxamid exhibits strong inhibitory activity against B. cinerea, regardless of the resistance profiles of those isolates to tested fungicides with different modes of action. This makes florylpicoxamid a powerful new solution to optimize gray mold control and manage fungicide resistance. © 2022 Society of Chemical Industry.

Targeting the Ubiquinol-Reduction (Qi) Site of the Mitochondrial Cytochrome bc1 Complex for the Development of Next Generation Quinolone Antimalarials.

Simple SummaryMalaria is a life-threatening disease which infects millions of people a year via mosquito bites, particularly in developing countries. Although many malaria drugs are available in the market today, all of them have been challenged by drug-resistant variants. Developing a new drug to fight mutated malaria is extremely critical. Cytochrome bc1 complex of malaria parasites is an important drug target focused on by several antimalarial development programs. One of those is the 4(1H)-quinolone series which inhibits cytochrome bc1 and effectively kills drug-resistant malaria parasites. However, some of these compounds have unexpected toxicity due to cross-species inhibition of human cytochrome bc1. In this work, we explore by experimental and computational studies how 4(1H)-quinolone compounds work with human and parasite cytochrome bc1. This information reveals the key to improved selectivity between human and parasite cytochrome bc1 and helps drug developers to design new compounds with better therapeutic efficiency and safety.Antimalarials targeting the ubiquinol-oxidation (Qo) site of the Plasmodium falciparum bc1 complex, such as atovaquone, have become less effective due to the rapid emergence of resistance linked to point mutations in the Qo site. Recent findings showed a series of 2-aryl quinolones mediate inhibitions of this complex by binding to the ubiquinone-reduction (Qi) site, which offers a potential advantage in circumventing drug resistance. Since it is essential to understand how 2-aryl quinolone lead compounds bind within the Qi site, here we describe the co-crystallization and structure elucidation of the bovine cytochrome bc1 complex with three different antimalarial 4(1H)-quinolone sub-types, including two 2-aryl quinolone derivatives and a 3-aryl quinolone analogue for comparison. Currently, no structural information is available for Plasmodial cytochrome bc1. Our crystallographic studies have enabled comparison of an in-silico homology docking model of P. falciparum with the mammalian’s equivalent, enabling an examination of how binding compares for the 2- versus 3-aryl analogues. Based on crystallographic and computational modeling, key differences in human and P. falciparum Qi sites have been mapped that provide new insights that can be exploited for the development of next-generation antimalarials with greater selective inhibitory activity against the parasite bc1 with improved antimalarial properties.

Florylpicoxamid, a new picolinamide fungicide with broad spectrum activity.

Following the introduction of fenpicoxamid, a natural product-based fungicide targeting the Qi site of mitochondrial cytochrome bc1 complex, a second generation fully synthetic picolinamide, florylpicoxamid, was discovered and its biological activity and attributes were characterized. In vitro fungal growth inhibition assays and in planta glasshouse biological activity evaluations showed florylpicoxamid was active against 21 different plant pathogenic fungi within the phyla Ascomycota and Basidiomycota. Among the pathogens evaluated, florylpicoxamid was most potent against Zymoseptoria tritici, the causal organism of wheat leaf blotch, providing 80% growth inhibition in vitro at 0.0046 mg L-1 and 80% disease control in planta at 0.03 mg L-1 when applied as a preventative treatment. Florylpicoxamid was more efficacious than epoxiconazole, fluxapyroxad, and benzovindiflupyr versus a Z. tritici wild-type isolate when applied as curative and preventative treatments, with superior 10-day curative reachback activity. Analytical studies and in planta tests demonstrated that florylpicoxamid partitioned into plants quickly and showed good systemicity and translaminar activity on both monocot and dicot plants. No cross-resistance was observed between florylpicoxamid and strobilurin or azole fungicides. Florylpicoxamid exerts its preventative effect by preventing spore germination on the leaf surface and curative activity by arresting mycelial growth and pycnidia development in leaf tissue. With strong broad spectrum fungicidal activity, florylpicoxamid delivers an innovative solution for growers to sustain high productivity and quality of many crops, and also provides a new option for developing effective strategies for fungicide resistance management. © 2021 Society of Chemical Industry.

Novel Endochin-Like Quinolones Exhibit Potent In Vitro Activity against Plasmodium knowlesi but Do Not Synergize with Proguanil.

Quinolones, such as the antimalarial atovaquone, are inhibitors of the malarial mitochondrial cytochrome bc1 complex, a target critical to the survival of both liver- and blood-stage parasites, making these drugs useful as both prophylaxis and treatment. Recently, several derivatives of endochin have been optimized to produce novel quinolones that are active in vitro and in animal models. While these quinolones exhibit potent ex vivo activity against Plasmodium falciparum and Plasmodium vivax, their activity against the zoonotic agent Plasmodium knowlesi is unknown. We screened several of these novel endochin-like quinolones (ELQs) for their activity against P. knowlesiin vitro and compared this with their activity against P. falciparum tested under identical conditions. We demonstrated that ELQs are potent against P. knowlesi (50% effective concentration, <117 nM) and equally effective against P. falciparum We then screened selected quinolones and partner drugs using a longer exposure (2.5 life cycles) and found that proguanil is 10-fold less potent against P. knowlesi than P. falciparum, while the quinolones demonstrate similar potency. Finally, we used isobologram analysis to compare combinations of the ELQs with either proguanil or atovaquone. We show that all quinolone combinations with proguanil are synergistic against P. falciparum However, against P. knowlesi, no evidence of synergy between proguanil and the quinolones was found. Importantly, the combination of the novel quinolone ELQ-300 with atovaquone was synergistic against both species. Our data identify potentially important species differences in proguanil susceptibility and in the interaction of proguanil with quinolones and support the ongoing development of novel quinolones as potent antimalarials that target multiple species.

Mitochondrial complex III in larval stage of Echinococcus multilocularis as a potential chemotherapeutic target and in vivo efficacy of atovaquone against primary hydatid cysts

Echinococcus multilocularis employs aerobic and anaerobic respiration pathways for its survival in the specialized environment of the host. Under anaerobic conditions, fumarate respiration has been identified as a promising target for drug development against E. multilocularis larvae, although the relevance of oxidative phosphorylation in its survival remains unclear. Here, we focused on the inhibition of mitochondrial cytochrome bc1 complex (complex III) and evaluated aerobic respiratory activity using mitochondrial fractions from E. multilocularis protoscoleces. An enzymatic assay revealed that the mitochondrial fractions possessed NADH-cytochrome c reductase (mitochondrial complexes I and III) and succinate-cytochrome c reductase (mitochondrial complexes II and III) activities in the aerobic pathway. Enzymatic analysis showed that atovaquone, a commercially available anti-malarial drug, inhibited mitochondrial complex III at 1.5 nM (IC50). In addition, culture experiments revealed the ability of atovaquone to kill protoscoleces under aerobic conditions, but not under anaerobic conditions, indicating that protoscoleces altered their respiration system to oxidative phosphorylation or fumarate respiration depending on the oxygen supply. Furthermore, combined administration of atovaquone with atpenin A5, a quinone binding site inhibitor of complex II, completely killed protoscoleces in the culture. Thus, inhibition of both complex II and complex III was essential for strong antiparasitic effect on E. multilocularis. Additionally, we demonstrated that oral administration of atovaquone significantly reduced primary alveolar hydatid cyst development in the mouse liver, compared with the untreated control, indicating that complex III is a promising target for development of anti-echinococcal drug.

1895. The Anti-Parasitic Drug Atovaquone Inhibits Arbovirus Replication

BackgroundAtovaquone, a hydroxynaphthoquinone, FDA Pregnancy category C, used for the treatment and prevention of pneumocysits jirovecii pneumonia (PCP), toxoplasmosis, babesiosis and malaria has in vitro activity against Zika virus (ZIKV). The mechanism of action against Plasmodium spp. and other parasites is based in the inhibition of mitochondrial cytochrome bc1 complex which further collapses parasite–mitochondrial membrane potential. But to date, antiviral activity of this drug has not been described.MethodsVero cells (monkey kidney epithelial cells) were seeded. At 24 hours of incubation, the cells were pretreated with ribavirin and brequinar (known antiviral drugs) and atovaquone at different concentrations for 1 hour and then infected with ZIKV Brazilian strain and Ugandan strain, and subsequently treated with drugs again. After incubation for 72 hours virus antigen Env-protein production was quantified by immunodetection. The concentration of atovaquone that decreased the level of Env-protein production by 50% was calculated by non-linear regression analysis (CC50). Cell viability was measured using the CellTiter 96 aqueous one solution cell proliferation assay (Promega, Madison, WI), according to the manufacturers protocol. Viral infection was rescued adding uracil to Vero cells pre-treated with ribavirin, brequinar and atovaquone. Experiment was repeated with Chikungunya virus (CHIKV).ResultsWe found that atovaquone inhibits ZIKV infection in Vero cells at smaller concentration (CC50 = 0.52 μM) than those used for parasitic killing. The effect is more prominent in the Brazilian strain when compared with the Ugandan strain. No cytotoxic effect was found in Vero cells up to 15 μM; above this concentration atovaquone formed crystals. Uracil rescues ZIKV infection after treatment with atovaquone. Atovaquone also inhibited CHIKV infection in Vero cells. ConclusionAtovaquone has antiviral activity against ZIKV likely via depletion of nucleotides blocking pyrimidine biosynthesis. Furthermore, the antiviral effect is applicable to other arboviruses which makes atovaquone a broad-spectrum antiviral drug and a potential attractive candidate for the treatment of ZIKV infection in vulnerable population such pregnant women and children.Disclosures All authors: No reported disclosures.

The binding mode of the natural compound ilicicolin H to the Qi site of the mitochondrial cytochrome bc1 complex

The binding mode of the natural compound ilicicolin H to the Qi site of the mitochondrial cytochrome bc1 complex

The antimalarial compound ELQ-400 is an unusual inhibitor of the bc1 complex, targeting both Qo and Qi sites.

Inhibitors of the mitochondrial respiratory chain cytochrome bc1 complex, such as the antimalarial atovaquone and ELQ-300, and many well-studied compounds, are classified as either Qo or Qi site inhibitors based on their site of action. Here, we investigated the site of action of ELQ-400 that showed an unusual behaviour, being effective against parasites resistant to the Qo site inhibitor atovaquone or to the Qi site inhibitor ELQ-300. Analysis of yeast mutants and comparison with atovaquone and other ELQs strongly suggest that ELQ-400 targets both Qo and Qi sites. Dual site inhibition would be particularly efficient as it would lower the risk of acquired resistance. However, such compounds are seldom found, which could be explained by structural and mechanistic differences between the sites.

Atovaquone and ELQ-300 Combination Therapy as a Novel Dual-Site Cytochrome bc1 Inhibition Strategy for Malaria.

Antimalarial combination therapies play a crucial role in preventing the emergence of drug-resistant Plasmodium parasites. Although artemisinin-based combination therapies (ACTs) comprise the majority of these formulations, inhibitors of the mitochondrial cytochrome bc1 complex (cyt bc1) are among the few compounds that are effective for both acute antimalarial treatment and prophylaxis. There are two known sites for inhibition within cyt bc1: atovaquone (ATV) blocks the quinol oxidase (Qo) site of cyt bc1, while some members of the endochin-like quinolone (ELQ) family, including preclinical candidate ELQ-300, inhibit the quinone reductase (Qi) site and retain full potency against ATV-resistant Plasmodium falciparum strains with Qo site mutations. Here, we provide the first in vivo comparison of ATV, ELQ-300, and combination therapy consisting of ATV plus ELQ-300 (ATV:ELQ-300), using P. yoelii murine models of malaria. In our monotherapy assessments, we found that ATV functioned as a single-dose curative compound in suppressive tests whereas ELQ-300 demonstrated a unique cumulative dosing effect that successfully blocked recrudescence even in a high-parasitemia acute infection model. ATV:ELQ-300 therapy was highly synergistic, and the combination was curative with a single combined dose of 1 mg/kg of body weight. Compared to the ATV:proguanil (Malarone) formulation, ATV:ELQ-300 was more efficacious in multiday, acute infection models and was equally effective at blocking the emergence of ATV-resistant parasites. Ultimately, our data suggest that dual-site inhibition of cyt bc1 is a valuable strategy for antimalarial combination therapy and that Qi site inhibitors such as ELQ-300 represent valuable partner drugs for the clinically successful Qo site inhibitor ATV.

Comparative iTRAQ proteomics revealed proteins associated with spermatogenic arrest of cattleyak

Male infertility of cattleyak due to spermatogenic arrest greatly restricts their effective utilization in yak breeding. Although much work has been done to investigate the mechanisms of spermatogenic arrest, there is no information regarding the differences of protein composition between cattleyak and yak testis. Comparative investigation of testis proteomes between cattleyak and yak using iTRAQ proteomics identified 256 differentially abundant proteins with fold change values higher than ±1.5. Most of the differentially abundant proteins were involved in extracellular matrix organization, response to stimulus, metabolic and cellular process, in which a large number of the cattleyak predominant proteins were associated with various stresses, cell adhesion and germ cell migration. Such upregulated proteins as integrins and their ligands in the extracellular matrix involved in ECM-receptor interaction pathway may help germ cells to endure pulling forces and impede their migration. In contrast, down-regulated proteins in cattleyak were associated with defects in various metabolic processes and cellular processes during spermatogenesis. Such Downregulated proteins as the subunits of mitochondrial cytochrome Bc1 complex involved in the Alzheimer's disease and oxidative phosphorylation pathways may lead to mitochondrial dysfunction and cell death in cattleyak testis. Biological significanceSpermatogenic arrest of cattleyak involves defects of both germ cells and their micro-environment in testis. In this study, Dozens of proteins possibly associated with spermatogenic arrest of cattleyak were identified by comparative iTRAQ proteomics, in which a large number of the cattleyak predominant proteins may act in response to various stresses (especially inflammatory stresses), enhance cell adhesion and impeded germ cell migration, while down-regulated proteins in cattleyak were associated with defects in various metabolic processes and cellular processes during spermatogenesis. Therefore, germ cells in testis of cattleyak may be constantly in a stress state (similar to inflammation) due to accumulation of some toxic intermediate products resulted from metabolic disturbances. Metabolic defects of germ cells and their deleterious micro-environment in testis of cattleyak may be the root of all other problems during spermatogenesis.

Structural analysis of mitochondrial cytochrome bc1 complex with atovaquone bound reveals the molecular basis of antimalarial drug action

Results We determined the 3.0-A resolution X-ray structure of mitochondrial cyt bc1 from Saccharomyces cerevisiae with atovaquone bound in the catalytic Qo site [1]. The drug, which has a pKa of 6.9, forms a polarized H-bond between its ionized hydroxyl group and His181 of the Rieske protein subunit. Multiple non-polar interactions with side chains of cytochrome b residues stabilize hydroxynaphthoquinone and chlorophenyl-cyclohexyl groups. The cytochrome b sequence analysis showed that the majority of the interacting residues are conserved, so that atovaquone binding to the yeast cyt bc1 is likely to resemble the binding to the complexes of the target organisms. Conclusion The binding mode provides detailed insights in the molecular basis of broad target spectrum, species-specific efficacies and acquired resistances. This may aid drug development to control the spread of drug-resistant parasites.

Structural analysis of atovaquone-inhibited cytochrome bc1 complex reveals the molecular basis of antimalarial drug action.

Atovaquone, a substituted hydroxynaphthoquinone, is a potent antimalarial drug that acts by inhibiting the parasite's mitochondrial cytochrome bc1 complex (cyt bc1). Mutations in cyt bc1 confer atovaquone resistance. Here we describe the X-ray structure of mitochondrial cyt bc1 from Saccharomyces cerevisiae with atovaquone bound in the catalytic Qo site, at 3.0-Å resolution. A polarized H-bond to His181 of the Rieske protein in cyt bc1 traps the ionized hydroxyl group of the drug. Side chains of highly conserved cytochrome b residues establish multiple non-polar interactions with the napththoquinone group, whereas less-conserved residues are in contact with atovaquone's cyclohexyl-chlorophenyl tail. Our structural analysis reveals the molecular basis of atovaquone's broad target spectrum, species-specific efficacies and acquired resistances, and may aid drug development to control the spread of resistant parasites.

AS2077715 is a selective inhibitor of fungal mitochondrial cytochrome bc₁ complex.

AS2077715 is a novel antifungal metabolite produced by the newly isolated fungal strain Capnodium sp. 339855. This compound has an analogous structure to funiculosin, an inhibitor of mitochondrial cytochrome bc1 complex (complex III). AS2077715 inhibited ubiquinol-cytochrome c reductase activity of Trichophyton mentagrophytes complex III with an IC50 of 0.9 ng ml(-1), while 6000-20,000 ng ml(-1) AS2077715 was required to obtain comparable inhibition of mammalian complex III. This inhibitor also suppressed the growth of T. mentagrophytes with a MIC of 0.08 μg ml(-1), while cytotoxicity for mammalian cells was >6 μg ml(-1). These results indicate that AS2077715 is a selective inhibitor of fungal mitochondrial complex III. AS2077715 in doses of 1 μg ml(-1) or greater showed fungicidal activity against T. mentagrophytes within 2 h of incubation. This early-onset effect of fungicidal activity was also exhibited by other complex III inhibitors. These results suggest that inhibition of complex III is a promising strategy for designing anti-Trichophyton agents and that AS2077715 can be a potential drug candidate for treating Trichophyton infections.

Therapeutic efficacy of AS2077715 against experimental tinea pedis in guinea pigs in comparison with terbinafine.

AS2077715 is a novel antifungal metabolite produced by the newly isolated fungal strain Capnodium sp. 339855. This compound has potent inhibitory activity against Trichophyton mentagrophytes mitochondrial cytochrome bc1 complex (complex III) and potent fungicidal activity against T. mentagrophytes, as measured in vitro. Here, we compared the effects of AS2077715 and terbinafine in a guinea pig model of tinea pedis. In a treatment regimen started from the day 7 after infection, 10 daily oral doses of 10 and 20 mg kg(-1) AS2077715 and 20 mg kg(-1) of terbinafine significantly decreased fungal colony-forming units (CFUs) in foot pad skin. In a treatment regimen started from the day 11 after infection, 20 mg kg(-1) AS2077715 significantly reduced fungal CFUs in foot pad skin after 7 daily doses in comparison with 20 mg kg(-1) terbinafine-treated guinea pigs. Our findings suggest that in vivo potency and efficacy of AS2077715 are equal to or greater than that of terbinafine, positioning AS2077715 as a good candidate for use in treating trichophytosis.

AS2077715: a novel antifungal antibiotic produced by Capnodium sp. 339855.

A novel antifungal agent, AS2077715, was isolated from the fermentation broth of a fungal strain (339855) identified as a new Capnodium species based on morphological characteristics and large-subunit ribosomal DNA sequencing. AS2077715 was isolated as a white powder via solvent extraction, HP-20 and ODS-B column chromatography and crystallization, and was determined to have the molecular formula C25H41NO7. AS2077715 has a structure related to that of funiculosin, an inhibitor of mitochondrial cytochrome bc1 complex (complex III), and showed antifungal activity against Trichophyton species.

Cyclosporin A and tacrolimus induce renal Erk1/2 pathway via ROS-induced and metalloproteinase-dependent EGF-receptor signaling

We previously demonstrated that the widely used immunosuppressive drugs cyclosporin A (CsA) and tacrolimus (FK506), independent of immunophilin binding, can activate profibrogenic transforming growth factor β (TGFβ)/Smad signaling cascades in rat renal mesangial cells (MC). Here we report that both peptidyl-prolyl cis/trans isomerase (PPIase) inhibitors activate the extracellular-signaling regulated kinase (ERK) a member of the mitogen activated protein kinase (MAPK) and induce a rapid and transient increase in ERK phosphorylation. The MEK inhibitor U0126, the reactive oxygen species (ROS) scavenger N-acetyl-cysteine (NAC), a cell-permeant superoxide dismutase (SOD) and stigmatellin, an inhibitor of mitochondrial cytochrome bc1 complex strongly attenuated the increase in ERK1/2 phosphorylation triggered by PPIase inhibitors. Moreover, neutralizing antibodies against heparin binding-epidermal growth factor (HB-EGF), and inhibition of the EGF receptor by either small interfering (si)RNA or AG1478, demonstrate that ERK activation by both PPIase inhibitors is mediated via HB-EGF-induced EGF receptor (EGFR) tyrosine kinase activation. The strong inhibitory effects achieved by GM6001 and TAPI-2 furthermore implicate the involvement of a desintegrin and metalloproteinase 17 (ADAM17). Concomitantly, the PPIase inhibitor-induced ADAM17 secretase activity was significantly reduced by SOD and stigmatellin thus suggesting that mitochondrial ROS play a primary role in PPIase inhibitor-induced and ADAM17-mediated HB-EGF shedding. Functionally, both immunosuppressants caused a strong increase in MC proliferation which was similarly impeded when cells were treated in the presence of NAC, TAPI-2 or AG1478, respectively. Our data suggest that CsA and FK506, via ROS-dependent and ADAM17-catalyzed HB-EGF shedding induce the mitogenic ERK1/2 signaling cascade in renal MC.

The role of the ubiquinone pool in modulating the superoxide production by the mitochondrial cytochrome bc1 complex

The role of the ubiquinone pool in modulating the superoxide production by the mitochondrial cytochrome bc1 complex