Search

Nature challenges healthy mammals with constant risk of infection by a wide variety of pathogens and with degradation of healthy tissue and control systems. I have been interested in drug delivery and improved delivery of therapeutics primarily to attack pathogens but with incidental value in assessing vital functions of a healthy mammal. My recent work uses protein design to approach such problems. Delivering biomacromolecules remains important both for therapeutics and in discerning and shaping functions of cells.My first thesis project focused on designing a better glomerular filtration rate (GFR) marker to facilitate assessment of renal function and seeking a marker reflecting water distribution in the body, which is relevant to distribution of highly soluble drugs [1]. Tritiated polyethylene glycol (PEG) was known to clear the kidneys effectively and correlated well with a GFR standard assay. A better radioactive label for PEG would allow for easy detection, including imaging. At the time of the project, only one biological conjugation with PEG was reported. Attaching an iodinatable moiety to polyethylene glycol (PEG) polymers of different sizes enabled tracking the compound using radioactive iodine. I made a series of related compounds and studied the pharmacokinetics (PK) and pharmacodynamics (PD) of these in rodents and ultimately in a dog. Using relatively long PEG polymers of molecular weight (MW) 5,000 to 6,000 daltons, the PEG dominated the behavior of the compounds, clearing rapidly through the kidneys. With shorter PEG polymers, the chemistry of the iodinatable group was more significant and the compounds were more likely to clear through the bile, to a degree making them unsuitable for a GFR marker but possibly useful to study liver function. Chapter 1, the published manuscript from my first thesis project, is cited in 20 scientific publications and 51 issued US patents. Variations on the design principles of my project have been used widely in the ph

Fungal infections, once considered a rare disease, have become an everyday problem in modern societies, posing major challenges to global health. It is estimated that more than one billion people are affected by fungal infections and 1.6 million people succumb to these diseases every year. Of the 600 species of fungi capable of causing infections in humans, species of the genus Candida cause more than 85% of infections, especially C. albicans, which has become a serious threat to human health in immunocompromised and immunosuppressed individuals. Unfortunately, the current arsenal of clinical drugs relies on only four classes of approved drugs (polyenes, azoles, echinocandins and allylamines), which are only partially effective, resulting in incomplete eradication of the fungal infection. In addition, the serious side effects, ranging from systemic or organ-specific toxicity to poor bioavailability and low activity, significantly hamper the clinical use of antifungals. These problems call for new effective and safe antifungal agents,but also for appropriate preclinical models to accurately study potential adverse effects on the human population and test their efficacy against fungal infections. In this sense, zebrafish (Danio rerio) embryos have become one of the most powerful preclinical animal models in infection biology and drug discovery, offering the unique opportunity to simultaneously monitor the safety and efficacy of the applied molecule in real time. With the aim of providing a preclinical platform for the identification of new safe antifungal drugs to effectively control C. albicans infection, we comprehensively tested the toxicity of 13 clinical antifungal drugs in the zebrafish embryo model. The 21 toxicity endpoints, including survival, teratogenicity, cardiotoxicity and hepatotoxicity, were evaluated and compared with adverse effects described in rats and humans. Of the clinical drugs, the efficacy of fluconazole and voriconazole was evaluated in the

Antifungal resistance is a growing concern as it poses a significant threat to public health. Fungal infections are a significant cause of morbidity and mortality, especially in immunocompromised individuals. The limited number of antifungal agents and the emergence of resistance have led to a critical need to understand the mechanisms of antifungal drug resistance. This review provides an overview of the importance of antifungal resistance, the classes of antifungal agents, and their mode of action. It highlights the molecular mechanisms of antifungal drug resistance, including alterations in drug modification, activation, and availability. In addition, the review discusses the response to drugs via the regulation of multidrug efflux systems and antifungal drug–target interactions. We emphasize the importance of understanding the molecular mechanisms of antifungal drug resistance to develop effective strategies to combat the emergence of resistance and highlight the need for continued research to identify new targets for antifungal drug development and explore alternative therapeutic options to overcome resistance. Overall, an understanding of antifungal drug resistance and its mechanisms will be indispensable for the field of antifungal drug development and clinical management of fungal infections.

Invasive pulmonary aspergillosis (IPA) is typically considered a disease of immunocompromised patients, but, recently, many cases have been reported in patients without typical risk factors. The aim of our study is to develop a risk predictive model for IPA through machine learning techniques (decision trees) in patients with influenza. We conducted a retrospective observational study analyzing data regarding patients diagnosed with influenza hospitalized at the University Hospital “Umberto I” of Rome during the 2018-2019 season. We collected five IPA cases out of 77 influenza patients. Although the small sample size is a limit, the most vulnerable patients among the influenza-infected population seem to be those with evidence of lymphocytopenia and those that received corticosteroid therapy., Interactive Intelligence

The risk of airborne transmission of SARS-CoV-2 by patients with Coronavirus disease 2019 (Covid-19) is unclear. However, theAerosoltherapy workgroup(GAT) ofthe Sociétéde Pneumologie de Langue Franc¸ aise decided at SARS-CoV2 pandemic preparedness to suggest avoiding a drug delivery via nebulization to reduce the risk of spreading the virus by this way. [...]

Invasive fungal infections are associated with significant morbidity and mortality, and their management is restricted to a variety of agents from five established classes of antifungal medication. In practice, existing antifungal agents are often constrained by dose-limiting toxicities, drug interactions, and the routes of administration. An increasing prevalence of invasive fungal infections along with rising rates of resistance and the practical limitations of existing agents has created a demand for the development of new antifungals, particularly those with novel mechanisms of action. This article reviews antifungal agents currently in various stages of clinical development. New additions to existing antifungal classes will be discussed, including SUBA-itraconazole, a highly bioavailable azole, and amphotericin B cochleate, an oral amphotericin formulation, as well as rezafungin, a long-acting echinocandin capable of once-weekly administration. Additionally, novel first-in-class agents such as ibrexafungerp, an oral glucan synthase inhibitor with activity against various resistant fungal isolates, and olorofim, a pyrimidine synthesis inhibitor with a broad spectrum of activity and oral formulation, will be reviewed. Various other innovative antifungal agents and classes, including MGCD290, tetrazoles, and fosmanogepix, will also be examined.

Coccidioidomycosis (Valley fever) is a fungal disease caused by the inhalation of Coccidioides posadasii or C. immitis. This neglected disease occurs in the desert areas of the western United States, most notably in California and Arizona, where infections continue to rise. Clinically, coccidioidomycosis ranges from asymptomatic to severe pulmonary disease and can disseminate to the brain, skin, bones, and elsewhere. New estimates suggest as many as 350,000 new cases of coccidioidomycosis occur in the United States each year. Thus, there is an urgent need for the development of a vaccine and new therapeutic drugs against Coccidioides infection. In this review, we discuss the battle against Coccidioides including the development of potential vaccines, the quest for new therapeutic drugs, and our current understanding of the protective host immune response to Coccidioides infection.

Candida glabrata is one of the most prevalent pathogenic Candida species in dental plaque on tooth surfaces. Candida biofilms exhibit an enhanced resistance against most antifungal agents. Thus, the development of alternative more potent and effective antimicrobials is required to overcome this resistance. In this study, three novel fluorinated derivatives and nine selenoester compounds were screened as novel antifungal and antibiofilm agents against C. krusei, C. parapsilosis, and C. glabrata (N = 81 dental isolates). C. glabrata strains were susceptible only to fluorinated compounds while C. krusei, C. parapsilosis, and C. glabrata were susceptible to the action of the selenoesters. The evaluated symmetrical selenoester compounds presented very good antifungal activity against all the tested C. glabrata dental isolates (1–4 µg/mL of minimum inhibitory concentration-MIC). The most active compound (Se-5) was able to inhibit and disperse C. glabrata biofilms. These results demonstrated that selenoesters may be novel and promising biocide agents against C. glabrata clinical dental isolates.

White and opaque cells of Candida albicans have the same genome but differ in gene expression patterns, metabolic profiles, and host niche preferences. We tested whether these differences, which include the differential expression of drug transporters, resulted in different sensitivities to 27 antifungal agents. The analysis was performed in two different strain backgrounds; although there was strain-to-strain variation, only terbinafine hydrochloride and caspofungin showed consistent, 2-fold differences between white and opaque cells across both strains.

In this study, the in vitro interactions of amphotericin B (AmB), voriconazole (VRC) and anidulafungin (AFG) in double and triple combinations against four species of multiresistant fungi (Fusarium solani, Lomentospora prolificans, Scopulariopsis brevicaulis and Scopulariopsis brumptii) were evaluated. In general, AmB combined with AFG was the most synergistic, especially against F. solani (7/8; 87.5%) when low concentrations of AmB were used, i.e. 0.125-0.5 µg/mL. The least active combination was AmB + VRC, with the lowest percentage of synergy against S. brevicaulis (2/11; 18.2%) and, in general, high concentrations of both antifungals were needed to achieve synergy. The triple combination was also highly synergistic against F. solani and S. brevicaulis, especially when the lowest concentrations of AmB were used, suggesting that use of combined therapies would reduce the toxicity of therapy. The triple combination was more effective than the double combinations in some cases, but not against all strains, suggesting that administration of three drugs is not always useful in the treatment of infections due to multiresistant fungi.

Marketing research of the modern range of antifungal drugs has been conducted according to the data of the State Expert Center of the Ministry of Public Health of Ukraine and Morion information retrieval program. The share of manufacturing countries of these drugs, and the ratio of different dosage forms and drug subgroups of the drugs studied by group D 01A have been determined according to the unified anatomic-therapeutic and chemical classification system ATC (Anatomical Therapeutic Chemical)., Проведены маркетинговые исследования современного ассортимента лекарственных средств противогрибкового действия по данным Государственного экспертного центра МОЗ Украины и информационно-поисковой программы «Морион». Установлено долю стран-производителей этих лекарственных средств, соотношение различных лекарственных форм и лекарственных подгрупп в соответствии с унифицированной анатомо-терапевтической и химической классификационной системой АТС (Anatomical Therapeutic Chemical) исследуемых ЛС по группе D 01A., Проведено маркетингові дослідження сучасного асортименту лікарських засобів протигрибкової дії за даними Державного експертного центру МОЗ України та інформаційно-пошукової програми «Моріон». Встановлено частку країн виробників цих лікарських засобів, співвідношення різних лікарських форм та лікарських підгруп відповідно до уніфікованої анатомо-терапевтичної та хімічної класифікаційної системи АТС (Anatomical Therapeutic Chemical) досліджуваних ЛЗ за групою D 01A.

Incidences of fungal infections are on the rise in immunosuppressed people. Predominant causative agents for these mycoses are species of the genus Candida, including Candida albicans, Candida glabrata and Candida dublieniensis. Despite a wide range of emerging pathogens, C. albicans remains the leading cause. According to recent epidemiological studies, blood stream infections with C. albicans cause annually ~55% mortality in approximately 300,000 patients from intensive care units worldwide. Furthermore, the percentage of morbidity linked to oral, esophageal and vulvovaginal mycoses cause by C. albicans reach up to 90%. Reasons for these medical concerns are the lack of efficient diagnostics and antifungal therapy. Here, we therefore sought to find novel antifungal strategies inspired by innate immune cells, such as neutrophils. These phagocytes are able to block the fungal pathogenicity. Neutrophils are bloodstream leukocytes serving as the first line of defense against pathogenic microbes. It has been shown that neutrophils have a strong antifungal activity by impairing the conversion of the dimorphic C. albicans from yeast to hyphal form (Y-H). Consequently, we raised the question whether other immune cells, such as mast cells, with less phagocytic cabapilities may have similar activity to neutrophils. Mast cells are tissue-dwelling cells. Mucosal tissue is rich in mast cells and usually constitutes the entry ports for fungal pathogens into the human body. A contribution of mast cells in antifungal defense is, thus, very likely. We human explored mast cell functions upon encounter with fungal pathogens. Interestingly, human mast cells show a transient potential to impair fungal viability. To understand the mechanism behind this impairment we analyzed the human mast cell functions in more detail. We found that human mast cells challenged with C. albicans, immediately degranulate and secrete distinct cytokines and chemokines in an orchestrated manner. The chemoki

Lipophilicity is one of the most frequently used physicochemical properties that affects compound solubility, determines its passive transport through biological membranes, influences biodistribution, metabolism and pharmacokinetics. We compared, ranked and grouped chromatographic lipophilicity indices and computationally estimated logP-s by sensitive and robust non-parametric approaches: sum of ranking differences (SRD) and generalized pairwise correlation method (GPCM). Chromatographic indices of fourteen neurotoxins and twenty one 1,2,4-triazole compounds have been derived from typical reversed-phase thin-layer chromatography and micellar chromatography. They were compared with in silico estimated logP-s. Under typical reversed-phase conditions, octadecyl-, octyl-, and cyanopropyl-modified silica have clear advantage over ethyl-, aminopropyl, and diol-modified beds, i.e., the preferable choice of the stationary phase follows this order: octadecyl gt octyl gt cyanopropyl gt ethyl gt octadecyl wettable gt aminopropyl gt diol. Many of these indices outperform the majority of computationally estimated logP-s. Clear distinction can be made based on cross-validation and statistical tests. Oppositely, micellar chromatography may not be successfully used for the lipophilicity assessment, since retention parameters obtained from the typical reversed-phase conditions outperform the parameters obtained by micellar chromatography. Both ranking approaches, SRD and GPCM, although based on different background, provide highly similar variable ordering and grouping leading to the same, above mentioned conclusions. However, GPCM results in more degeneracy, i.e., in some cases it cannot distinguish the lipophilicity parameters whereas SRD and its cross-validated version can. On the other hand GPCM produces a more characteristic grouping. Both methods can be successfully used for selection of the most and least appropriate lipophilicity measures.

The antifungal armamentarium for the treatment of systemic fungal infections has increased in recent years. Although very helpful to control/eliminate fungal infections, the available antifungal drugs do have some limitations such as antifungal drug resistance. As an example, azole resistance is an issue in patients with chronic mucocutaneous candidiasis caused by C. albicans in the context of HIV-infected individuals with recurrent oropharyngeal and esophageal candidiasis. A similar trend in vaginal isolates of C. albicans has been seen in women prone to recurrent vaginal candidiasis exposed to long-term fluconazole (Bulik et al., 2009),(Shahid and Sobel, 2009). In the latter scenario fortunately relatively rare to date therapeutic options available for oral management of fluconazole-reduced susceptibility C. albicans are few, resulting in the inconvenient use of long-term topical imidazoles. These facts have generated greater interest in the development of new antifungal drugs using various synthetic and naturally occurring antimicrobial molecules. Natural antimicrobial peptides, such as defensins produced by epithelial cells, showed a broad range of antibacterial activity and could play a role in preventing microbial infections(Decanis et al., 2009),(Zaslof, 2002). These antimicrobial peptides generally exhibit selective toxicity for microorganisms and show fewer propensities to induce microbial resistance., The original document contains color images.

Fungal cell wall is a rigid structure mainly composed of polysaccharides (up to 90 %) and glycoproteins. It is essential for survival of the fungal cells, because it protects them against bursting caused by internal turgor pressure and against mechanical injury. Because of its absence in mammalian cells, it is an attractive target for antifungal agents. Thus, for various reasons, it might be important to know how the cell wall is synthesized, and how to analyze its composition. We provide here information about in vitro analysis of the biosynthetic activities of the main fungal wall and describe some methods for rapid analysis of cell wall composition by using specific enzymatic degradations. We also describe some additional methods that can be occasionally used to analyze fungal wall properties or composition. These methods provide powerful tools to evaluate changes in fungal cell walls and will be useful for screening new compounds for antifungal activity that might cause inhibition of cell wall biosynthesis and/or alter the structure of the fungal cell wall.

Antibiotics (AB) and antifungals are large, heterogeneous, and commonly prescribed groups of drugs. Even thou, their use in therapy are considered to be safe, during concomitant use with other drugs the result can be clinically important interaction. Certain classes of AB or antifungal drugs are known to interact with many other drugs, but the interaction potential of them is not uniform among members of the class. Therefore, choosing the appropriate drug allows the possibility to avoid potentially dangerous drug-drug interactions., Antibiotici (AB) i antimikotici predstavljaju velike, heterogene i često propisivane grupe lekova. Iako se smatraju za relativno bezbedne lekove, pri istovremenoj primeni sa drugim lekovima mogu da stupe u klinički značajne interakcije. Određene grupe AB i antimikotika su poznate da stupaju u interakcije, ali potencijal za stupanje u interakcije unutar istih klasa lekova je različit, što pruža mogućnost da se odgovarajućim odabirom AB odnosno antimikotika izbegnu potencijalno štetne lek-lek interakcije.

In this work 20 clinical and 3 environmental yeast isolates were characterized by classical morphological and physiological methods, as well as molecular methods based on PCR of the ITS1-5.8S rDNA-ITS2 region. The characteristic morphology and biochemical profiles observed in these samples correspond to those described for the Pichia genera, more specifically to P. anomala. The profiles of susceptibility to five antifungal drugs were determined by two broth dilution methods. The results obtained by both methods were comparable and showed that clinical isolates presented more resistance to azoles, amphotericin B, and 5-fluorocytosine, than environmental ones did. By amplification and sequencing of internal transcribed spacers (ITS1 and ITS2) and the ribosomal 5.8S DNA, the yeast samples were divided into four groups, where the strains within each group had the same sequence. Of the analyzed yeast isolates, 78% were identified as Pichia anomala. Using RAPD analysis with seven different Operon primers, polymorphism was observed within the four groups. Our study highlights the growing importance of P. anomala in fungemic episodes in premature neonates. Furthermore, the methodologies used provide a powerful tool to identify and determine differences in similar strains of this yeast

SCOPUS: ar.j, info:eu-repo/semantics/published

SCOPUS: ar.j, info:eu-repo/semantics/published

Humane steroidogene Hormone spielen eine essentielle Rolle in der Wachstums-Regulation, Metabolismus, Geschlechter-Differenzierung, Reproduktion, Entzündungsreaktionen, Erhaltung des Blut-Volumens, Regulierung des Blutzucker-Levels und der Regulierung des Immunsystems innerhalb des Körpers. Während ihre Aktivität durch spezifische Rezeptoren reguliert wird, werden die Prä-Rezeptor-Konzentrationen durch die Enzyme steroidogenes Zytochrom P450 (CYP450) und Hydroxysteroid Dehydrogenasen (HSDs) gesteuert. Humanes CYP11B1 und 11β-HSDs regulieren die intrazellulären Konzentration von Glucocorticoiden. CYP11B2 erhält die Mineralocorticoid-Konzentrationen. Humanes CYP17A1, CYP19A1 und 17β-HSDs katalysieren die lokale Synthese von geschlechtsspezifischen Steroiden. Aufgrund deren essentieller Funktionen stellt die Inhibierung dieser Enzyme eine attraktive Möglichkeit dar um verschiedenste Krankheiten wie Metabolisches Syndrom, Bluthochdruck, kongestive Herzinsuffizienz, Kastrations-resistenter Prostatakrebs, Brustkrebs, Typ II Diabetes und Osteoporose zu behandeln. Jedoch kann der Ausfall deren katalytischer Funktion zu endokrinen Fehlfunktionen wie zum Beispiel Nebennieren-Insuffizienz, fehlerhafte Entwicklung des Geschlechts, Unfruchtbarkeit und kardiovaskulären Komplikationen führen. In dieser Arbeit wurden menschliches CYP11B1/2, CYP17A1, CYP19A1, 11β-HSD1/2, 17β-HSD2, CYP2D6 und 5α-Reduktase als potentielle Ziel- und Anti-Ziel-Enzyme untersucht. Die durchgeführte Studie hatte zwei große Ziele; 1) Identifizierung von neuen Inhibitoren der Ziel-Enzyme innerhalb von bestehenden ‚small molecule‘ Datenbanken und 2) die Identifizierung von endokrinen Disruptoren (EDs) der Ziel-Enzyme aufgrund von Exposition gegenüber Umwelt-Chemikalien. Die experimentell validierten Pharmakophor-Modelle der 11β-HSD2 Inhibierung wurden für ein virtuelles Screening von Anti-Ziel-Strukturen basierend auf bereits zugelassenen Medikamenten herangezogen. Dadurch konnten zwei potente anti-Pilz Medik, Human steroidogenic hormones have a crucial role in growth regulation, metabolism, sexual differentiation, reproduction, inflammation, maintenance of blood volume, regulation of blood sugar level, and immune system regulation in the body. They regulate their actions through target receptors; however, their pre-receptor concentrations are monitored by steroidogenic cytochrome P450 (CYP450) enzymes and hydroxysteroid dehydrogenases (HSDs). Human CYP11B1 and 11β-HSDs regulate the intracellular concentrations of glucocorticoids. CYP11B2 maintains the concentrations of mineralocorticoids. Human CYP17A1, CYP19A1, and 17β-HSDs catalyze the local synthesis of sex steroids. Due to their crucial functions, the inhibition of these enzymes offers an attractive way for the treatment of multiple disorders such as metabolic syndrome, hypertension, congestive heart failure, castration-resistant prostate cancer, breast cancer, type II diabetes, and osteoporosis. On the other hand, the failure of their catalytic functions can lead to the endocrine disruptions causing adrenal insufficiency, impaired sexual development, infertility, and cardiovascular complications. In this thesis, the human CYP11B1/2, CYP17A1, CYP19A1, 11β-HSD1/2, 17β-HSD2, CYP2D6, and 5α-reductase were studied as target and anti-target enzymes. The research performed for this study had two broader aims; 1) identification of the novel inhibitors of target enzymes amongst the small molecule databases, 2) and to find the endocrine disrupting chemicals (EDCs) of the target enzymes from exposure to the environmental chemicals. The experimentally validated pharmacophore models of 11β-HSD2 inhibition were employed for anti-target virtual screening of approved drugs. This study found two potent antifungal drugs itraconazole and posaconazole as EDCs of 11β-HSD2. Pharmacophore-based virtual screening for 17β-HSD2 inhibitors discovered twelve new potential lead compounds. Further, in vivo studies are required to confirm the inh, vorgelegt von M.Sc., PharmD. Muhammad Akram, Kumulative Dissertation aus acht Artikeln, Zusammenfassung in deutscher Sprache, Dissertation Universität Innsbruck 2019