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Background: Dietary habits may play a non-negligible role in the development, duration, and severity of acne, as shown in past critical review articles on such association., Methods: The aim of this systematic review is to supplement data available on scientific literature spanning the last 10 years by inserting the keywords "acne" or "acne vulgaris" and "diet", "nutrition", "food", "chocolate", "dairy", "whey protein", "fatty acid", or "drink" in the timeframe "January 2009-April 2020" within the PubMed database., Results: Fifty-three reviewed articles met eligibility criteria. They included 11 interventional clinical trials (seven randomized controlled trials and four uncontrolled open label studies) and 42 observational studies (17 case-control and 22 cross-sectional studies, and three descriptive studies)., Conclusions: This review reinforces the notion of a rapidly growing exponential trend of interest in this subject by the scientific community. Acne-promoting factors include high GI/GL food, dairy products, fat food, and chocolate, whereas acne-protective factors include fatty acids, fruit, and vegetable intake. The role played by specific dietary components pertaining to different foods, as done for milk (full-fat/whole, reduced-fat, low-fat/skim milk), dairy products (milk cream, ice cream, yogurt, cheese, etc.), or chocolate (cocoa, dark/milk chocolate), remains an unsolved issue and objective of future research., (© 2021 the International Society of Dermatology.)

TRIM proteins form a protein family that is characterized by a conserved tripartite motif domain comprising a RING domain, one or two B-box domains and a coiled-coil region. Members of this large protein family are important regulators of numerous cellular functions including innate immune responses, transcriptional regulation and apoptosis. Key to their cellular role is their E3 ligase activity which is conferred by the RING domain. Self-association is an important characteristic of TRIM protein activity and is mediated by homodimerization via the coiled-coil region, and in some cases higher order association via additional domains of the tripartite motif. In many of the TRIM family proteins studied thus far, RING dimerization is an important prerequisite for E3 ligase enzymatic activity though the propensity of RING domains to dimerize differs significantly between different TRIMs and can be influenced by other regions of the protein., (© 2020 The Author(s).)

Keratin extracted from wool fibers has recently gained attention as an abundant source of renewable, biocompatible material for tissue engineering and drug delivery applications. However, keratin extraction and processing generally require a copious use of chemicals, not only bearing consequences for the environment but also possibly compromising the envisioned biological outcome. In this study, we present, for the first time, keratin-PVP biocomposite fibers obtained via an all-water co-electrospinning process and explored their properties modulation as a result of different thermal crosslinking treatments. The protein-based fibers featured homogenous morphologies and average diameters in the range of 170-290 nm. The thermomechanical stability and response to a wet environment can be tuned by acting on the curing time; this can be achieved without affecting the 3D fibrous network nor the intrinsic hydrophilic behavior of the material. More interestingly, our protein-based membranes treated at 170 °C for 18 h successfully sustained the attachment and growth of primary human dermal fibroblasts, a cellular model which can recapitulate more faithfully the physiological human tissue conditions. Our proposed approach can be viewed as pivotal in designing tunable protein-based scaffolds for the next generation of skin tissue growth devices., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Aims: Optimal management of redundant or malfunctioning leads is controversial. We aimed to assess safety and efficacy of mechanical transvenous lead extraction (TLE) in patients with abandoned leads., Methods and Results: Consecutive TLE procedures performed in our centre from January 2009 to December 2017 were considered. We evaluated the safety and efficacy of mechanical TLE in patients with abandoned (Group 1) compared to non-abandoned (Group 2) leads. We analysed 1210 consecutive patients that required transvenous removal of 2343 leads. Group 1 accounted for 250 patients (21%) with a total of 617 abandoned leads (26%). Group 2 comprised 960 patients (79%) with 1726 leads (74%). The total number of leads (3.0 vs. 2.0), dwelling time of the oldest lead (108.00 months vs. 60.00 months) and infectious indications for TLE were higher in Group 1. Clinical success was achieved in 1168 patients (96.5%) with a lower rate in Group 1 (90.4% vs. 98.1%; P < 0.001). Major complications occurred in only 9 patients (0.7%), without significant differences among the two groups. The presence of one or more abandoned leads [odds ratio (OR) 3.47; 95% confidence interval (CI) 1.07-11.19; P = 0.037] and dwelling time of the oldest lead (OR 1.01 for a month; 95% CI 1.01-1.02; P < 0.001) were associated with a higher risk of clinical failure., Conclusion: Transvenous mechanical lead extraction is a safe procedure also in high-risk settings, as patients with abandoned leads. Success rate resulted a bit lower, especially in the presence of abandoned leads with long implantation time., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2020. For permissions, please email: journals.permissions@oup.com.)

The Candida parapsilosis genome encodes for five agglutinin-like sequence (Als) cell-wall glycoproteins involved in adhesion to biotic and abiotic surfaces. The work presented here is aimed at analyzing the role of the two still uncharacterized ALS genes in C. parapsilosis, CpALS4790 and CpALS0660 , by the generation and characterization of C pALS4790 and CpALS066 single mutant strains. Phenotypic characterization showed that both mutant strains behaved as the parental wild type strain regarding growth rate in liquid/solid media supplemented with cell-wall perturbing agents, and in the ability to produce pseudohyphae. Interestingly, the ability of the CpALS0660 null mutant to adhere to human buccal epithelial cells (HBECs) was not altered when compared with the wild-type strain, whereas deletion of CpALS4790 led to a significant loss of the adhesion capability. RT-qPCR analysis performed on the mutant strains in co-incubation with HBECs did not highlight significant changes in the expression levels of others ALS genes. In vivo experiments in a murine model of vaginal candidiasis indicated a significant reduction in CFUs recovered from BALB/C mice infected with each mutant strain in comparison to those infected with the wild type strain, confirming the involvement of CpAls4790 and CpAls5600 proteins in C. parapsilosis vaginal candidiasis in mice.

Background: Managing elderly patients with infection or malfunction deriving from a cardiac implantable electronic device (CIED) may be challenging. The aim of this study was to evaluate safety and efficacy of mechanical transvenous lead extraction (TLE) in elderly patients., Methods: Patients who had undergone TLE in single tertiary referral center were divided in two groups (group 1: ≥ 80 years; group 2: < 80 years) and their acute and chronic outcomes were compared. All patients were treated with manual traction or mechanical dilatation., Results: Our analysis included 1316 patients (group 1: 202; group 2: 1114 patients), with a total of 2513 leads extracted. Group 1 presented more comorbidities, more pacemakers than implantable cardioverter-defibrillators, whereas the dwelling time of the oldest lead and the number of leads were similar, irrespective of patient's age. In group 1, the radiological success rate for lead was higher (99.0% vs 95.9%; P < .001) and the fluoroscopy time lower (13.0 vs 15.0 min; P = .04) than in group 2. Clinical success was reached in 1273 patients (96.7%), without significant differences between groups (group 1: 98.0% vs group 2: 96.4%; P = .36). Major complications occurred in 10 patients (0.7%) without significative differences between patients with more or less than 80 years (group 1: 1.5% vs group 2: 0.6%; P = .24) and with no procedure-related deaths in elderly group., Conclusions: Mechanical TLE in elderly patients is a safe and effective procedure. In the over-80s, a comparable incidence of major complications with younger patients was observed, with at least a similar efficacy of the procedure., (© 2020 Wiley Periodicals, Inc.)

Flavin-containing monooxygenases (FMOs) are ubiquitous in all domains of life and metabolize a myriad of xenobiotics, including toxins, pesticides and drugs. However, despite their pharmacological importance, structural information remains bereft. To further our understanding behind their biochemistry and diversity, we used ancestral-sequence reconstruction, kinetic and crystallographic techniques to scrutinize three ancient mammalian FMOs: AncFMO2, AncFMO3-6 and AncFMO5. Remarkably, all AncFMOs could be crystallized and were structurally resolved between 2.7- and 3.2-Å resolution. These crystal structures depict the unprecedented topology of mammalian FMOs. Each employs extensive membrane-binding features and intricate substrate-profiling tunnel networks through a conspicuous membrane-adhering insertion. Furthermore, a glutamate-histidine switch is speculated to induce the distinctive Baeyer-Villiger oxidation activity of FMO5. The AncFMOs exhibited catalysis akin to human FMOs and, with sequence identities between 82% and 92%, represent excellent models. Our study demonstrates the power of ancestral-sequence reconstruction as a strategy for the crystallization of proteins.

Monooxygenases (MOs) face the challenging reaction of an organic target, oxygen and a cofactor - most commonly heme or flavin. To correctly choreograph the substrates spatially and temporally, MOs evolved a variety of strategies, which involve structural flexibility. Besides classical domain and loop movements, flavin-containing MOs feature conformational changes of their flavin prosthetic group and their nicotinamide cofactor. With similar mechanisms emerging in various subclasses, their generality and involvement in selectivity are intriguing questions. Cytochrome P450 MOs are often inherently plastic and large movements of individual segments throughout the entire structure occur. As these complicated and often unpredictable movements are largely responsible for substrate uptake, engineering strategies for these enzymes were mostly successful when randomly mutating residues across the entire structure., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The flavin-containing monooxygenases (FMOs) play an important role in drug metabolism but they also have a high potential in industrial biotransformations. Among the hitherto characterized FMOs, there was no thermostable representative, while such biocatalyst would be valuable for FMO-based applications. Through a targeted genome mining approach, we have identified a gene encoding for a putative FMO from Nitrincola lacisaponensis, an alkaliphilic extremophile bacterium. Herein, we report the biochemical and structural characterization of this newly discovered bacterial FMO (NiFMO). NiFMO can be expressed as active and soluble enzyme at high level in Escherichia coli (90-100 mg/L of culture). NiFMO is relatively thermostable (melting temperature (T m ) of 51 °C), displays high organic solvent tolerance, and accepts a broad range of substrates. The crystal structure of NiFMO was solved at 1.8 Å resolution, which allows future structure-based enzyme engineering. Altogether, NiFMO represents an interesting newly discovered enzyme with the appropriate features to develop into an industrially applied biocatalyst.

CYP153s are bacterial class I P450 enzymes traditionally described as alkane hydroxylases with a high terminal regioselectivity. They have been more recently shown to also catalyze hydroxylations at nonactivated carbon atoms of small heterocycles. The aim of our work was to perform an extensive characterization of this subfamily in order to deliver a toolbox of CYP153 enzymes for further development as biocatalysts. Through the screening of recently sequenced bacterial genomes, 20 CYP153s were selected, comprising 17 single monooxygenase domains and three multidomain variants, where the monooxygenase domain is naturally fused to its redox partners in a single polypeptide chain. The 20 novel variants were heterologously expressed, and their activity was screened toward octane and small heterocycles. A more extended substrate characterization was then performed on three representative candidates, and their crystal structures were unveiled and compared with those of the known CYP153A7 and CYP153A33. The tested enzymes displayed a wide range of activities, ranging from Ω and Ω-1 hydroxylations of lauric acid to indigo-generating indole modification. The comparative analysis highlighted a conserved architecture and amino acid composition of the catalytic core close to the heme, while showing a huge degree of structural plasticity and flexibility in those regions hosting the substrate recognition sites. Although dealing with this type of conformational variability adds a layer of complexity and difficulty to structure-based protein engineering, such diversity in substrate acceptance and recognition promotes the investigated CYP153s as a prime choice for tailoring designer hydroxylases.

Background Pharmacological inhibition of polo-like kinase 1 (PLK1) represents a new approach for the treatment of solid tumors. This study was aimed at determining the first cycle dose-limiting toxicities (DLTs) and related maximum tolerated dose (MTD) of NMS-1286937, a selective ATP-competitive PLK1-specific inhibitor. Secondary objectives included evaluation of its safety and pharmacokinetic (PK) profile in plasma, its antitumor activity, and its ability to modulate intracellular targets in biopsied tissue. Methods This was a Phase I, open-label, dose-escalation trial in patients with advanced/metastatic solid tumors. A treatment cycle comprised 5 days of oral administration followed by 16 days of rest, for a total of 21 days (3-week cycle). Results Nineteen of 21 enrolled patients with confirmed metastatic disease received study medication. No DLTs occurred at the first 3 dose levels (6, 12, and 24 mg/m 2 /day). At the subsequent dose level (48 mg/m 2 /day), 2 of 3 patients developed DLTs. An intermediate level of 36 mg/m 2 /day was therefore investigated. Four patients were treated and two DLTs were observed. After further cohort expansion, the MTD and recommended phase II dose (RP2D) were determined to be 24 mg/m 2 /day. Disease stabilization, observed in several patients, was the best treatment response observed. Hematological toxicity (mostly thrombocytopenia and neutropenia) was the major DLT. Systemic exposure to NMS-1286937 increased with dose and was comparable between two cycles of treatment following oral administration of the drug. Conclusions This study successfully identified the MTD and DLTs for NMS-1286937 and characterized its safety profile.

Flavin-containing monooxygenases (FMOs) are emerging as effective players in oxidative drug metabolism. Until recently, the functions of the five human FMO isoforms were mostly linked to their capability of oxygenating molecules containing soft N- and S-nucleophiles. However, the human FMO isoform 5 was recently shown to feature an atypical activity as Baeyer-Villiger monooxygenase. With the aim of evaluating such an alternative entry point in the metabolism of active pharmaceutical ingredients, we selected and tested drug molecules bearing a carbonyl group on an aliphatic chain. Nabumetone and pentoxifylline, two widely used pharmaceuticals, were thereby demonstrated to be efficiently oxidized in vitro by FMO5 to the corresponding acetate esters with high selectivity. The proposed pathways explain the formation of a predominant plasma metabolite of pentoxifylline as well as the crucial transformation of the pro-drug nabumetone into the pharmacologically active compound. Using the recombinant enzyme, the ester derivatives of both drugs were obtained in milligram amounts, purified, and fully characterized. This protocol can potentially be extended to other FMO5 candidate substrates as it represents an effective and robust bench-ready platform applicable to API screening and metabolite synthesis.

Background: An extremely elevated erythrosedimentation rate (ESR), defined as equal or higher than 100 mm/h, has been linked to serious underlying conditions, such as infections, connective tissue and oncologic disease., Aim: To analyze a group of patients in order to determine the underlying diagnosis and the characteristics associated with extremely elevated ESR in our environment., Methods: Cross-sectional study of adult patients, who presented with at least one ESR equal or higher than 100 mm/h at Hospital Italiano, in Buenos Aires (Buenos Aires, Argentina) between January 2002 and August 2014., Results: During the previously stated period of time, we analyzed the results of 879 patients. All patients were over 18 years of age. The median for the ESR results was 111 mm/h (interquartile range 105-120). The most prevalent etiology of an elevated ESR was infectious (41.64%), followed by malignancies (21.62%) and autoimmune / inflammatory diseases (12.97%). The most frequent individual diagnosis found was pneumonia (11.49%), followed by undetermined causes (5.92%)., Conclusion: When comparing inpatient versus outpatient populations, the most frequent cause was infectious in the former group, while malignancies were the most frequent diagnosis in the latter.

Background: A clinical registry encompasses a selective set of rigorously collected and stored clinical data focused on a specific condition. Hip fracture is a common complication of osteoporosis in elderly patients. Hip fracture substantially increases the risk of death and major morbidity in the elderly patients. Limited data regarding hip fracture are available from Latin America and Argentina. The purpose of this project is to create an institutional registry of elderly patients with hip fracture in order to obtain data that reveal the impact of this disease in our environment, allowing us to evaluate different strategies of patient's care and clinical outcomes., Objective: To describe the implementation of an institutional registry of elderly patients with hip fracture in Argentina., Methods: In this article, we described the creation, implementation, and data management of a prospective registry of elderly patients with hip fracture. The registry contains information on baseline demographics, comorbidities, laboratory, and radiological data. Follow-up at 3 and 12 months postfracture is done by phone interview to assess physical function, readmissions, and morbi-mortality. Clinical Trials registry number NCT02279550., Conclusion: In this project, we have created a hip fracture registry. We hope that this registry will provide valuable data that can lead us to new lines of research, addressed to answer questions raised in clinical practice.

Psoriasis is an inflammatory disease frequently associated with psychiatric disturbances and sleep disorders. The aim of the study was to assess the prevalence of depression, interaction anxiety, audience anxiety, and sleep quality in psoriatic patients. One hundred and two psoriatic patients were enrolled and underwent the following questionnaires: Zung Self-Rating Depression Scale (SDS), Interaction Anxiousness Scale (IAS), Audience Anxiousness Scale (AAS), Pittsburgh Sleep Quality Index (PSQI). The severity of skin lesions was assessed by Psoriasis Area Severity Index (PASI). The presence of a link between clinical variables and with demographic data has been investigated. Psoriasis was linked to depression, interaction and audience anxiety, as well as to poor sleep quality; 37.5% of patients were depressed, 46.1% scored above 37 at the IAS, 47.1% scored above 33 at the AAS. Thirty-nine subjects (38.2%) presented a PSQI ≥ 5. An association between interaction anxiety and lower limbs psoriasis-related erythema as well as between PSQI and head psoriasis-related erythema was found, particularly among male patients. Hence, psoriatic patients should be assessed from a holistic point of view, in order to identify associated disorders that could benefit from targeted treatments.

Flavin-containing mono-oxygenases are known as potent drug-metabolizing enzymes, providing complementary functions to the well-investigated cytochrome P450 mono-oxygenases. While human FMO isoforms are typically involved in the oxidation of soft nucleophiles, the biocatalytic activity of human FMO5 (along its physiological role) has long remained unexplored. In this study, we demonstrate the atypical in vitro activity of human FMO5 as a Baeyer-Villiger mono-oxygenase on a broad range of substrates, revealing the first example to date of a human protein catalyzing such reactions. The isolated and purified protein was active on diverse carbonyl compounds, whereas soft nucleophiles were mostly non- or poorly reactive. The absence of the typical characteristic sequence motifs sets human FMO5 apart from all characterized Baeyer-Villiger mono-oxygenases so far. These findings open new perspectives in human oxidative metabolism.

Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase responsible for the development of different tumor types. Despite the remarkable clinical activity of crizotinib (Xalkori), the first ALK inhibitor approved in 2011, the emergence of resistance mutations and of brain metastases frequently causes relapse in patients. Within our ALK drug discovery program, we identified compound 1, a novel 3-aminoindazole active on ALK in biochemical and in cellular assays. Its optimization led to compound 2 (entrectinib), a potent orally available ALK inhibitor active on ALK-dependent cell lines, efficiently penetrant the blood-brain barrier (BBB) in different animal species and highly efficacious in in vivo xenograft models. Moreover, entrectinib resulted to be strictly potent on the closely related tyrosine kinases ROS1 and TRKs recently found constitutively activated in several tumor types. Entrectinib is currently undergoing phase I/II clinical trial for the treatment of patients affected by ALK-, ROS1-, and TRK-positive tumors.

In this study we report the effects of fungal metabolites isolated from cultures of the fungus Trichoderma citrinoviride ITEM 4484 on the feeding preference of the aphid Rhopalosiphum padi, a major pest of cereal crops. Different phagodeterrent metabolites were purified by a combination of direct and reverse phase column chromatography and thin-layer chromatography. Chemical investigations, by spectroscopic and chemical methods, led to the identification of different long chain primary alcohols (LCOHs) of the general formula R-OH, wherein R is a long, unbranched, unsubstituted, linear aliphatic group. LCOHs have been reported as components of lepidopteran pheromone blends, but their phagodeterrent effect to aphids is herein reported for the first time. The effects of LCOHs on R. padi were studied by behavioral and electrophysiological bioassays. Feeding preference tests that were carried out with winged and wingless morphs of R. padi showed that LCOHs had high phagodeterrent activity and restrained aphids from settling on treated leaves at a concentration as low as 0.15 mM (0.036 g/l). The results of different electrophysiological analyses indicated that taste receptor neurons located on the aphid tarsomeres were involved in the LCOHs perception. Behavioral assays carried out with some commercial agrochemicals, including azadirachtin A, pyrethrum and a mineral oil-based product, in combination with 1-hexadecanol, the LCOH most abundantly produced by T. citrinoviride ITEM 4484, showed that these different active principles could be applied together, resulting in a useful increase of the phagodeterrent effect. The data shown indicate that these compounds can be profitably utilized for novel applications in biotechnical control of aphid pests. Furthermore, the tested LCOHs have no chiral centers and therefore can be obtained with good yield and at low cost through chemical synthesis, as well as from natural sources.

Rationale: Neutrophils play a fundamental role in a number of chronic lung diseases. Among the mediators of their recruitment to the lung, CXCL8 (IL-8) is considered to be one of the major players. CXCL8 exerts its chemotactic activity by binding to its GPCR receptors (CXCR1/R2) located on neutrophils, as well as through interactions with glycosaminoglycans (GAGs) on cell surfaces including those of the microvascular endothelium. Binding to GAG co-receptors is required to generate a solid-phase haptotactic gradient and to present IL-8/CXCL8 in a proper conformation to its receptors on circulating neutrophils., Methods: We have engineered increased GAG-binding affinity into human CXCL8, thereby obtaining a competitive inhibitor that displaces wild-type IL-8/CXCL8 from GAGs. By additionally knocking-out the GPCR binding domain of the chemokine, we generated a dominant negative protein (dnCXCL8; PA401) with potent anti-inflammatory characteristics proven in vivo in a murine model of LPS-induced lung inflammation (Adage et al., 2015). Here we have further investigated PA401 activity in this pulmonary model by evaluating plasma changes induced by LPS on white blood cells (WBC) and a broad range of inflammatory markers, especially chemokines, by addressing immediate effects of PA401 on these parameters in healthy and LPS exposed mice., Results: Aerosolized LPS induced a significant increase in bronchoalveolar lavage (BAL) neutrophils after 3 and 7h, as well as an increase in total WBC and changes in 21 of the 59 measured plasma markers, mostly belonging to the chemokine family. PA401 treatment in saline exposed mice didn't induce major changes in any of the measured parameters. When administered to LPS aerosolized mice, PA401 caused a significant normalization of KC/mCXCL1 and other inflammatory markers, as well as of blood WBC count. In addition, BAL neutrophils were significantly reduced, confirming the previously observed lung anti-inflammatory activity of PA401 in this experiment., Conclusions: PA401 is a new promising biologic therapeutic with a novel and unique mechanism of action for interfering with neutrophilic lung inflammation, that also normalizes plasma inflammatory markers., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Background: Respiratory infection caused by Pneumocystis jiroveci is a common opportunistic infection in patients with human immunodeficiency virus (HIV) with CD4 counts < 200 cells/mm(3). However, it has also been reported in patients with other causes of immunosuppression., Objectives: To compare the characteristics, severity and mortality of respiratory infection by P. jiroveci in patients with and without HIV infection., Methods: Retrospective cohort follow-up of adult patients admitted to our hospital with infection by P. jiroveci since 2006 to 2013., Results: We included 82 patients with respiratory infection by P. jiroveci of which 55% (45) were not infected with HIV. In this group, 68.8% (31) had diagnosis of cancer and 20% (9) received solid-organ transplant. 57.9% (26) were hospitalized in an intensive care unit. 42.2% (19) suffered multiple organ failure (MOF), 46.7% (21) required mechanical ventilation (MV) and 40.9% (18) inotropic drugs. Mortality was 33.3% (15). Statistically significant differences were observed between groups in age (p < 0.001), requirement of MV (p < 0.001) inotropic drugs (p 0.001) and MOF (p < 0.001). Mortality was higher in the HIV-positive group, reaching statistical significance (p 0.007)., Conclusion: Pneumocystis pneumonia mortality was higher in patients without HIV, who suffered more complications and progression to respiratory failure with MOF.

Aim: The aim of the present study was to investigate the relationship between maximal aerobic capacity (VO(2max)) and repeated sprint ability (RSA) in a group of professional soccer players., Methods: Forty-one professional soccer players (age 23 ± 4 yrs, height 180.0 ± 5.3 cm, weight 79.6 ± 5.3 kg) were required to perform tests to assess RSA and VO(2max) on two separate days with at least 48 hr rest between testing sessions. Each player performed a treadmill test to determine their VO(2max) and a test for RSA involving the players completing 6 × 40 m sprints (turn after 20 m) with 20 s active recovery between each sprint., Results: There was a significant negative correlation between body mass normalised VO(2max) and mean sprint time (RSAmean) (r = -0.655; P < 0.01) and total sprint time (RSAtotal) (r = -0.591, P < 0.01)., Conclusion: Results of the current study indicate that VO(2max) is one important factor aiding soccer players in the recovery from repeated sprint type activities.

As neuroinflammation is an early event in the pathogenesis of Alzheimer' s disease, new selective antiinflammatory drugs could lead to promising preventive strategies. We evaluated the safety, tolerability, pharmacokinetics and pharmacodynamics of CHF5074, a new microglial modulator, in a 12-week, double-blind, placebo-controlled, parallel groups, ascending dose study involving 96 MCI patients. Subjects were allocated into three successive study cohorts to receive ascending, titrated doses of CHF5074 (200, 400 or 600 mg/day) or placebo. Vital signs, cardiac safety, neuropsychological performance and safety clinical laboratory parameters were assessed on all subjects. Plasma samples were collected throughout the study for measuring drug concentrations, soluble CD40 ligand (sCD40L) and TNF-α. At the end of treatment, cerebrospinal fluid (CSF) samples were optionally collected after the last dose to measure drug levels, β- amyloid1-42 (Aβ42), tau, phospho-tau181, sCD40L and TNF-α. Ten patients did not complete the study: one in the placebo group (consent withdrawn), two in the 200-mg/day treatment group (consent withdrawn and unable to comply) and seven in the 400-mg/day treatment group (five AEs, one consent withdrawn and one unable to comply). The most frequent treatment-emergent adverse events were diarrhea, dizziness and back pain. There were no clinically significant treatmentrelated clinical laboratory, vital sign or ECG abnormalities. CHF5074 total body clearance depended by gender, age and glomerular filtration rate. CHF5074 CSF concentrations increased in a dose-dependent manner. At the end of treatment, mean sCD40L and TNF-α levels in CSF were found to be inversely related to the CHF5074 dose (p=0.037 and p=0.001, respectively). Plasma levels of sCD40L in the 600-mg/day group were significantly lower than those measured in the placebo group (p=0.010). No significant differences between treatment groups were found in neuropsychological tests but a positive dose-response trend was found on executive function in APOE4 carriers. This study shows that CHF5074 is well tolerated in MCI patients after a 12-week titrated treatment up to 600 mg/day and dose-dependently affects central nervous system biomarkers of neuroinflammation.

CHF5074 has been shown to inhibit brain β-amyloid deposition and attenuate memory deficits in different transgenic mice models of Alzheimer disease. We evaluated the safety, pharmacokinetics, and pharmacodynamics of 3 ascending dose regimens of CHF5074 (200, 400, and 600 mg/d for 14 d) in a double-blind, placebo-controlled, parallel group study involving 48 healthy subjects. Plasma, urine, and cerebrospinal fluid (CSF) samples were collected for measuring drug and main metabolite concentrations and potential biomarkers of pharmacodynamic activity (β-amyloid1-40, β-amyloid1-42, soluble CD40 ligand, and tumor necrosis factor-α). All subjects completed the study, and no serious or severe adverse events were reported. The maximum tolerated dose was close to 600 mg/d with mild diarrhea being the most frequent adverse event at this dose. CHF5074 reached peak plasma levels 2 to 3 hours after drug administration and then was slowly eliminated (t(1/2z)=30 h) in the urine as glucoronide. Systemic exposure to the drug appeared to be dose-proportional with a 2-fold accumulation ratio at steady state. Metabolite plasma levels peaked at 4 to 5 hours and accounted for about 25% of the parent compound. Drug levels in the CSF were dose-proportional. The drug dose-dependently lowered the levels of the soluble CD40 ligand, a marker of microglia activation, in both plasma and CSF samples.

Previous work has established the existence of dystrophin-nitric oxide (NO) signaling to histone deacetylases (HDACs) that is deregulated in dystrophic muscles. As such, pharmacological interventions that target HDACs (that is, HDAC inhibitors) are of potential therapeutic interest for the treatment of muscular dystrophies. In this study, we explored the effectiveness of long-term treatment with different doses of the HDAC inhibitor givinostat in mdx mice--the mouse model of Duchenne muscular dystrophy (DMD). This study identified an efficacy for recovering functional and histological parameters within a window between 5 and 10 mg/kg/d of givinostat, with evident reduction of the beneficial effects with 1 mg/kg/d dosage. The long-term (3.5 months) exposure of 1.5-month-old mdx mice to optimal concentrations of givinostat promoted the formation of muscles with increased cross-sectional area and reduced fibrotic scars and fatty infiltration, leading to an overall improvement of endurance performance in treadmill tests and increased membrane stability. Interestingly, a reduced inflammatory infiltrate was observed in muscles of mdx mice exposed to 5 and 10 mg/kg/d of givinostat. A parallel pharmacokinetic/pharmacodynamic analysis confirmed the relationship between the effective doses of givinostat and the drug distribution in muscles and blood of treated mice. These findings provide the preclinical basis for an immediate translation of givinostat into clinical studies with DMD patients.

Purpose: This phase I trial assessed the safety, maximally tolerated dose (MTD) and pharmacokinetics of TRKA/CDK inhibitor PHA-848125AC in adult patients with advanced/metastatic solid tumors., Patients and Methods: Patients with relapsed or refractory solid tumors, for which no standard therapy existed, were eligible. PHA-848125AC was administered orally in two schedules: daily for 7 consecutive days in 2-week cycles (i.e. 7 days on/7 days off q2wks; S1) or daily for 4 consecutive days a week for 3 weeks in 4-week cycles (i.e. 4 days on/3 days off x 3wks q4wks; S2)., Results: Thirty-seven patients were treated in this study, 22 in S1 and 15 in S2. The recommended phase II dose (RP2D) was 150 mg/day for either schedule. The dose-limiting toxicities (DLTs) in S1 included ataxia (Grade 2-4) and tremors (Grade 2-3). In S2, DLTs included tremors (Grade 2-3), elevated lipase (Grade 3), increased creatinine (Grade 2), and nausea and vomiting (Grade 3). These events were all reversible. In S2, out of 14 patients evaluable for efficacy, 2 patients with thymic carcinoma, showed partial response and stable disease was observed in 3 patients. Stable disease was observed in 6 out 14 patients evaluable for efficacy on S1. Drug pharmacokinetics demonstrated a half-life of approximately 33 h, and dose-proportionality with accumulation by a factor of 3 after repeated administrations., Conclusion: The RP2D of PHA-848125AC was 150 mg/day on both schedules. Based on the responses noted in thymic carcinoma, a phase II study for patients with that disease is currently enrolling.

Polo-like kinase 1 (PLK1) is a serine/threonine protein kinase considered to be the master player of cell-cycle regulation during mitosis. It is indeed involved in centrosome maturation, bipolar spindle formation, chromosome separation, and cytokinesis. PLK1 is overexpressed in a variety of human tumors and its overexpression often correlates with poor prognosis. Although five different PLKs are described in humans, depletion or inhibition of kinase activity of PLK1 is sufficient to induce cell-cycle arrest and apoptosis in cancer cell lines and in xenograft tumor models. NMS-P937 is a novel, orally available PLK1-specific inhibitor. The compound shows high potency in proliferation assays having low nanomolar activity on a large number of cell lines, both from solid and hematologic tumors. NMS-P937 potently causes a mitotic cell-cycle arrest followed by apoptosis in cancer cell lines and inhibits xenograft tumor growth with clear PLK1-related mechanism of action at well-tolerated doses in mice after oral administration. In addition, NMS-P937 shows potential for combination in clinical settings with approved cytotoxic drugs, causing tumor regression in HT29 human colon adenocarcinoma xenografts upon combination with irinotecan and prolonged survival of animals in a disseminated model of acute myelogenous leukemia in combination with cytarabine. NMS-P937, with its favorable pharmacologic parameters, good oral bioavailability in rodent and nonrodent species, and proven antitumor activity in different preclinical models using a variety of dosing regimens, potentially provides a high degree of flexibility in dosing schedules and warrants investigation in clinical settings., (©2012 AACR.)

The discovery and characterization of two new chemical classes of potent and selective Polo-like kinase 1 (PLK1) inhibitors is reported. For the most interesting compounds, we discuss the biological activities, crystal structures and preliminary pharmacokinetic parameters. The more advanced compounds inhibit PLK1 in the enzymatic assay at the nM level and exhibit good activity in cell proliferation on A2780 cells. Furthermore, these compounds showed high levels of selectivity on a panel of unrelated kinases, as well as against PLK2 and PLK3 isoforms. Additionally, the compounds show acceptable oral bioavailability in mice making these inhibitors suitable candidates for further in vivo activity studies., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

As part of our drug discovery effort, we identified and developed 4,5-dihydro-1H-pyrazolo[4,3-h]quinazoline derivatives as PLK1 inhibitors. We now report the optimization of this class that led to the identification of NMS-P937, a potent, selective and orally available PLK1 inhibitor. Also, in order to understand the source of PLK1 selectivity, we determined the crystal structure of PLK1 with NMS-P937. The compound was active in vivo in HCT116 xenograft model after oral administration and is presently in Phase I clinical trials evaluation., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The purpose of this study was to evaluate the effects of the speed, agility, quickness (SAQ) training method on power performance in soccer players. Soccer players were assigned randomly to 2 groups: experimental group (EG; n = 50) and control group (n = 50). Power performance was assessed by a test of quickness--the 5-m sprint, a test of acceleration--the 10-m sprint, tests of maximal speed--the 20- and the 30-m sprint along with Bosco jump tests--squat jump, countermovement jump (CMJ), maximal CMJ, and continuous jumps performed with legs extended. The initial testing procedure took place at the beginning of the in-season period. The 8-week specific SAQ training program was implemented after which final testing took place. The results of the 2-way analysis of variance indicated that the EG improved significantly (p < 0.05) in 5-m (1.43 vs. 1.39 seconds) and in 10-m (2.15 vs. 2.07 seconds) sprints, and they also improved their jumping performance in countermovement (44.04 vs. 4.48 cm) and continuous jumps (41.08 vs. 41.39 cm) performed with legs extended (p < 0.05). The SAQ training program appears to be an effective way of improving some segments of power performance in young soccer players during the in-season period. Soccer coaches could use this information in the process of planning in-season training. Without proper planning of the SAQ training, soccer players will most likely be confronted with decrease in power performance during in-season period.

A series of 4,5-dihydro-1H-pyrazolo[4,3-h]quinazoline derivatives was optimized as Polo-like kinase 1 inhibitors. Extensive SAR afforded a highly potent and selective PLK1 compound. The compound showed good antiproliferative activity when tested in a panel of tumor cell lines with PLK1 related mechanism of action and with good in vivo antitumor efficacy in two xenograft models after i.v. administration., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Cdc7 serine/threonine kinase is a key regulator of DNA synthesis in eukaryotic organisms. Cdc7 inhibition through siRNA or prototype small molecules causes p53 independent apoptosis in tumor cells while reversibly arresting cell cycle progression in primary fibroblasts. This implies that Cdc7 kinase could be considered a potential target for anticancer therapy. We previously reported that pyrrolopyridinones (e.g., 1) are potent and selective inhibitors of Cdc7 kinase, with good cellular potency and in vitro ADME properties but with suboptimal pharmacokinetic profiles. Here we report on a new chemical class of 5-heteroaryl-3-carboxamido-2-substituted pyrroles (1A) that offers advantages of chemistry diversification and synthetic simplification. This work led to the identification of compound 18, with biochemical data and ADME profile similar to those of compound 1 but characterized by superior efficacy in an in vivo model. Derivative 18 represents a new lead compound worthy of further investigation toward the ultimate goal of identifying a clinical candidate.

Altered expression and activity of cyclin-dependent kinase (CDK) and tropomyosin receptor kinase (TRK) families are observed in a wide variety of tumors. In those malignancies with aberrant CDK activation, the retinoblastoma protein (pRb) pathway is deregulated, leading to uncontrolled cell proliferation. Constitutive activation of TRKs is instead linked to cancer cell survival and dissemination. Here, we show that the novel small-molecule PHA-848125, a potent dual inhibitor of CDKs and TRKs, possesses significant antitumor activity. The compound inhibits cell proliferation of a wide panel of tumoral cell lines with submicromolar IC(50). PHA-848125-treated cells show cell cycle arrest in G(1) and reduced DNA synthesis, accompanied by inhibition of pRb phosphorylation and modulation of other CDK-dependent markers. The compound additionally inhibits phosphorylation of TRKA and its substrates in cells, which functionally express this receptor. Following oral administration, PHA-848125 has significant antitumor activity in various human xenografts and carcinogen-induced tumors as well as in disseminated primary leukemia models, with plasma concentrations in rodents in the same range as those found active in inhibiting cancer cell proliferation. Mechanism of action was also confirmed in vivo as assessed in tumor biopsies from treated mice. These results show that the dual CDK-TRK inhibitor PHA-848125 has the potential for being a novel and efficacious targeted drug for cancer treatment., ((c) 2010 AACR.)

A transcriptional signature of the pan-cyclin-dependent kinase (Cdk) inhibitor PHA-793887 was evaluated as a potential pharmacodynamic and/or response biomarker in tumor and skin biopsies from patients treated in a phase I clinical study. We first analyzed the expression of a number of known E2F-dependent genes that were predicted to be modulated after Cdk2 and Cdk4 inhibition in xenograft tumor and skin samples of mice treated with the compound. This panel of 58 selected genes was then analyzed in biopsies from seven patients treated with PHA-793887 in a phase I dose escalation clinical trial in solid tumors. Quantitative real-time PCR or microarray analyses were done in paired skin and tumor biopsies obtained at baseline and at cycle 1. Analysis by quantitative real-time PCR of the signature in skin biopsies of patients treated at three different doses showed significant transcriptional downregulation with a dose-response correlation. These data show that PHA-793887 modulates genes involved in cell cycle regulation and proliferation in a clinical setting. The observed changes are consistent with its mechanism of action and correlate with target modulation in skin and with clinical benefit in tumors.

Abnormal proliferation mediated by disruption of the mechanisms that keep the cell cycle under control is a hallmark of virtually all cancer cells. Compounds targeting complexes between cyclin-dependent kinases (CDKs) and cyclins (Cy) and inhibiting their activity are regarded as promising antitumor agents to complement the existing therapies. An expansion of pyrazolo[4,3-h]quinazoline chemical class oriented to the development of three points of variability was undertaken leading to a series of compounds able to inhibit CDKs both in vitro and in vivo. Starting from the CDK selective but poorly soluble hit compound 1, we succeeded in obtaining several compounds showing enhanced inhibitory activity both on CDKs and on tumor cells and displaying improved physical properties and pharmacokinetic behavior. Our study led to the identification of compound 59 as a highly potent, orally bioavailable CDK inhibitor that exhibited significant in vivo efficacy on the A2780 ovarian carcinoma xenograft model. The demonstrated mechanisms of action of compound 59 on cancer cell lines and its ability to inhibit tumor growth in vivo render this compound very interesting as potential antineoplastic agent.

We have recently reported CDK inhibitors based on the 6-substituted pyrrolo[3,4-c]pyrazole core structure. Improvement of inhibitory potency against multiple CDKs, antiproliferative activity against cancer cell lines and optimization of the physico-chemical properties led to the identification of highly potent compounds. Compound 31 (PHA-793887) showed good efficacy in the human ovarian A2780, colon HCT-116 and pancreatic BX-PC3 carcinoma xenograft models and was well tolerated upon daily treatments by iv administration. It was identified as a drug candidate for clinical evaluation in patients with solid tumors., (Copyright 2010 Elsevier Ltd. All rights reserved.)

The discovery of a novel class of inhibitors of cyclin dependent kinases (CDKs) is described. Starting from compound 1, showing good potency as inhibitor of CDKs but being poorly selective against a panel of serine-threonine and tyrosine kinases, new analogues were synthesized. Enhancement in selectivity, antiproliferative activity against A2780 human ovarian carcinoma cells, and optimization of the physical properties and pharmacokinetic profile led to the identification of highly potent and orally available compounds. Compound 28 (PHA-848125), which in the preclinical xenograft A2780 human ovarian carcinoma model showed good efficacy and was well tolerated upon repeated daily treatments, was identified as a drug candidate for further development. Compound 28 is currently undergoing phase I and phase II clinical trials.

Cdc7 kinase is a key regulator of the S-phase of the cell cycle, known to promote the activation of DNA replication origins in eukaryotic organisms. Cdc7 inhibition can cause tumor-cell death in a p53-independent manner, supporting the rationale for developing Cdc7 inhibitors for the treatment of cancer. In this paper, we conclude the structure-activity relationships study of the 2-heteroaryl-pyrrolopyridinone class of compounds that display potent inhibitory activity against Cdc7 kinase. Furthermore, we also describe the discovery of 89S, [(S)-2-(2-aminopyrimidin-4-yl)-7-(2-fluoro-ethyl)-1,5,6,7-tetrahydropyrrolo[3,2-c]pyridin-4-one], as a potent ATP mimetic inhibitor of Cdc7. Compound 89S has a Ki value of 0.5 nM, inhibits cell proliferation of different tumor cell lines with an IC50 in the submicromolar range, and exhibits in vivo tumor growth inhibition of 68% in the A2780 xenograft model.

Somatosensory evoked potentials (SSEP) of the n. medianus, with clinically significant neurological findings helps in diagnosing a number of different cervical spine lesions. SSEP of the n. medianus and neurological observations also aid in diagnosticis of the disc-radicular conflict. A final diagnosis is set by the computerised tomography (CT). The tested positive predictive value of assessment of disc-radicular conflict of the cervical spine by means of SSEP is 87%. The 95% confidence interval (CI) is 0.87 (0.81-0.93) (Tab. 3, Fig. 4, Ref. 23). Full Text (Free, PDF) www.bmj.sk.

Cdc7 is an essential kinase that promotes DNA replication by activating origins of replication. Here, we characterized the potent Cdc7 inhibitor PHA-767491 (1) in biochemical and cell-based assays, and we tested its antitumor activity in rodents. We found that the compound blocks DNA synthesis and affects the phosphorylation of the replicative DNA helicase at Cdc7-dependent phosphorylation sites. Unlike current DNA synthesis inhibitors, PHA-767491 prevents the activation of replication origins but does not impede replication fork progression, and it does not trigger a sustained DNA damage response. Treatment with PHA-767491 results in apoptotic cell death in multiple cancer cell types and tumor growth inhibition in preclinical cancer models. To our knowledge, PHA-767491 is the first molecule that directly affects the mechanisms controlling initiation as opposed to elongation in DNA replication, and its activities suggest that Cdc7 kinase inhibition could be a new strategy for the development of anticancer therapeutics.

We have recently reported a new class of CDK2/cyclin A inhibitors based on a bicyclic tetrahydropyrrolo[3,4-c]pyrazole scaffold. The introduction of small alkyl or cycloalkyl groups in position 6 of this scaffold allowed variation at the other two diversity points. Conventional and polymer-assisted solution phase chemistry provided a way of generating compounds with improved biochemical and cellular activity. Optimization of the physical properties and pharmacokinetic profile led to a compound which exhibited good efficacy in vivo on A2780 human ovarian carcinoma.

Living-cell microarrays are powerful tools for functional genomics and drug discovery. However, despite several attempts to improve this technology, it is still a challenge to obtain microarrays of cells efficiently overexpressing or downregulating specific genes to address complex phenotypes. Here, we present a cell-based microarray for phenotype screening on primary and cancer cells based on the localized reverse infection by retroviruses. Viral vectors are immobilized on a nanostructured titanium dioxide (ns-TiO2) film obtained by depositing a supersonic beam of titania clusters on a glass substrate. We validated the retroviral cell array by overexpression of GFP reporter genes in primary and cancer cells, and by RNA interference of p53 in primary cells by analyzing effects in cell growth. We demonstrate that ns-TiO2 retroviral arrays are an enabling tool for the study of gene function of families of genes for complex phenotypes and for the identification of novel drug targets.

The synthesis of the major metabolite of a potent 3-aminopyrazole CDK2/cyclin A inhibitor is presented. A stereoconservative approach starting from malic acid was employed to construct the hydroxy-substituted pyrrolidinone moiety. In the key step of the synthesis the use of cyanoborohydride immobilized on Amberlyst 26 in trifluoroethanol represented a valid alternative to conventional solution-phase reducing agents.

Purpose: To determine whether a regimen of cyclosporine (CSA) and methotrexate (MTX), or CSA and hydroxychloroquine (HCQ) introduced in early rheumatoid arthritis (RA) can produce a significant improvement in clinical outcome and/or retard radiographic damage in comparison with standard monotherapy with CSA alone., Methods: One hundred five patients with active RA of less than 36 months duration, who had never previously been treated with immunosuppressive agents, were included in a 12-month, multi-center, open, randomized trial. Patients who fulfilled the criteria for early severe RA were randomized to receive either combination therapy (CSA + MTX n = 34, CSA + HCQ n = 35) or CSA alone (n = 36)., Results: CSA + MTX was more effective than the other two treatment groups in controlling RA symptoms. CSA+MTX did not show a significant radiographic progression according to Larsen-Dale (0.90 +/- 3.89 compared to baseline values, P > 0.05); moreover, patients treated with CSA alone or CSA+HCQ showed a significant worsening of Larsen-Dale score (2.91 +/- 5.99 and 2.97 +/- 4.28 respectively vs baseline values, P < 0.05), although not significant when compared with the CSA + HCQ group (P = 0.56 and 0.39, respectively)., Conclusions: This trial indicated that CSA+MTX was more effective than the other two treatments in improving clinical data and inhibiting radiographic progression, although the differences were not significant in this relatively small study. However, the difference was significant in favor of CSA + MTX regarding ACR 50% response.