Correlates of protection and determinants of SARS-CoV-2 breakthrough infections 1 year after third dose vaccination.

BACKGROUND: The emergence of new SARS-CoV-2 variants and the waning of immunity raise concerns about vaccine effectiveness and protection against COVID-19. While antibody response has been shown to correlate with the risk of infection with the original variant and earlier variants of concern, the effectiveness of antibody-mediated protection against Omicron and the factors associated with protection remain uncertain. METHODS: We evaluated antibody responses to SARS-CoV-2 spike (S) and nucleocapsid (N) antigens from Wuhan and variants of concern by Luminex and their role in preventing breakthrough infections 1 year after a third dose of mRNA vaccination, in a cohort of health care workers followed since the pandemic onset in Spain (N = 393). Data were analyzed in relation to COVID-19 history, demographic factors, comorbidities, vaccine doses, brand, and adverse events. RESULTS: Higher levels of anti-S IgG and IgA to Wuhan, Delta, and Omicron were associated with protection against vaccine breakthroughs (IgG against Omicron S antigen HR, 0.06, 95%CI, 0.26-0.01). Previous SARS-CoV-2 infection was positively associated with antibody levels and protection against breakthroughs, and a longer time since last infection was associated with lower protection. In addition, priming with BNT162b2 followed by mRNA-1273 booster was associated with higher antibody responses than homologous mRNA-1273 vaccination. CONCLUSIONS: Data show that IgG and IgA induced by vaccines against the original strain or by hybrid immunization are valid correlates of protection against Omicron BA.1 despite immune escape and support the benefits of heterologous vaccination regimens to enhance antibodies and the prioritization of booster vaccination in individuals without recent infections.

An allopurinol adherence tool using plasma oxypurinol concentrations.

AIMS: This study aimed to develop and evaluate an allopurinol adherence tool based on steady-state oxypurinol plasma concentrations, allopurinol's active metabolite. METHODS: Plasma oxypurinol concentrations were simulated stochastically from an oxypurinol pharmacokinetic model for allopurinol doses of 100-800 mg daily, accounting for differences in renal function, diuretic use and ethnicity. For each scenario, the 20th percentile for peak and trough concentrations defined the adherence threshold, below which imperfect adherence was assumed. Predictive performance was evaluated using both simulated low adherence and against data from 146 individuals with paired oxypurinol plasma concentrations and adherence measures. Sensitivity and specificity (S&S), negative and positive predictive values (NPV, PPV) and receiver operating characteristic (ROC) area under the curve (AUC) were determined. The predictive performance of the tool was evaluated using adherence data from an external study (CKD-FIX). RESULTS: The allopurinol adherence tool produced S&S values for trough thresholds of 89-98% and 76-84%, respectively, and 90%-98% and 76-83% for peak thresholds. PPV and NPV were 79-84% and 88-94%, respectively, for trough and 80-85% and 89-98%, respectively, for peak concentrations. The ROC AUC values ranged from 0.84 to 0.88 and from 0.86 to 0.89 for trough and peak concentrations, respectively. S&S values for the external evaluation were found to be 75.8% and 86.5%, respectively, producing an ROC AUC of 0.8113. CONCLUSION: A tool to identify people with gout who require additional support to maintain adherence using plasma oxypurinol concentrations was developed and evaluated. The predictive performance of the tool is suitable for adherence screening in clinical trials and may have utility in some clinical practice settings.

Atoms in molecules in real space: a fertile field for chemical bonding.

In this perspective, we review some recent advances in the concept of atoms-in-molecules from a real space perspective. We first introduce the general formalism of atomic weight factors that allows unifying the treatment of fuzzy and non-fuzzy decompositions under a common algebraic umbrella. We then show how the use of reduced density matrices and their cumulants allows partitioning any quantum mechanical observable into atomic or group contributions. This circumstance provides access to electron counting as well as energy partitioning, on the same footing. We focus on how the fluctuations of atomic populations, as measured by the statistical cumulants of the electron distribution functions, are related to general multi-center bonding descriptors. Then we turn our attention to the interacting quantum atom energy partitioning, which is briefly reviewed since several general accounts on it have already appeared in the literature. More attention is paid to recent applications to large systems. Finally, we consider how a common formalism to extract electron counts and energies can be used to establish an algebraic justification for the extensively used bond order-bond energy relationships. We also briefly review a path to recover one-electron functions from real space partitions. Although most of the applications considered will be restricted to real space atoms taken from the quantum theory of atoms in molecules, arguably the most successful of all the atomic partitions devised so far, all the take-home messages from this perspective are generalizable to any real space decompositions.

Metadherin Regulates Inflammatory Breast Cancer Invasion and Metastasis.

Inflammatory breast cancer (IBC) is one of the most lethal subtypes of breast cancer (BC), accounting for approximately 1-5% of all cases of BC. Challenges in IBC include accurate and early diagnosis and the development of effective targeted therapies. Our previous studies identified the overexpression of metadherin (MTDH) in the plasma membrane of IBC cells, further confirmed in patient tissues. MTDH has been found to play a role in signaling pathways related to cancer. However, its mechanism of action in the progression of IBC remains unknown. To evaluate the function of MTDH, SUM-149 and SUM-190 IBC cells were edited with CRISPR/Cas9 vectors for in vitro characterization studies and used in mouse IBC xenografts. Our results demonstrate that the absence of MTDH significantly reduces IBC cell migration, proliferation, tumor spheroid formation, and the expression of NF-κB and STAT3 signaling molecules, which are crucial oncogenic pathways in IBC. Furthermore, IBC xenografts showed significant differences in tumor growth patterns, and lung tissue revealed epithelial-like cells in 43% of wild-type (WT) compared to 29% of CRISPR xenografts. Our study emphasizes the role of MTDH as a potential therapeutic target for the progression of IBC.

QM/MM Energy Decomposition Using the Interacting Quantum Atoms Approach.

The interacting quantum atoms (IQA) method decomposes the quantum mechanical (QM) energy of a molecular system in terms of one- and two-center (atomic) contributions within the context of the quantum theory of atoms in molecules. Here, we demonstrate that IQA, enhanced with molecular mechanics (MM) and Poisson-Boltzmann surface-area (PBSA) solvation methods, is naturally extended to the realm of hybrid QM/MM methodologies, yielding intra- and inter-residue energy terms that characterize all kinds of covalent and noncovalent bonding interactions. To test the robustness of this approach, both metal-water interactions and QM/MM boundary artifacts are characterized in terms of the IQA descriptors derived from QM regions of varying size in Zn(II)- and Mg(II)-water clusters. In addition, we analyze a homologous series of inhibitors in complex with a matrix metalloproteinase (MMP-12) by carrying out QM/MM-PBSA calculations on their crystallographic structures followed by IQA energy decomposition. Overall, these applications not only show the advantages of the IQA QM/MM approach but also address some of the challenges lying ahead for expanding the QM/MM methodology.

Understanding the Conformational Properties of Fluorinated Polypeptides: Molecular Modelling of Unguisin A.

In this work, we investigate the conformational properties of unguisin A, a natural macrocyclic heptapeptide that incorporates a γ-aminobutyric acid (Gaba), and four of its difluorinated stereoisomers at the Gaba residue. According to nuclear magnetic resonance (NMR) experiments, their secondary structure depends dramatically on the stereochemistry of the fluorinated carbon atoms. However, many molecular details of the structure and flexibility of these systems remain unknown, so that a rationale of the conformational changes induced by the fluorine atoms in the macrocycle is still missing. To fill this gap, we apply enhanced molecular dynamics (MD) techniques to explore the peptide conformational space in dimethyl sulfoxide solution followed by 4-8 µs of conventional MD simulations that provide extensive equilibrium sampling. The simulations, which compare reasonably well with the NMR-based observations, show that the secondary structure of the macrocycle is altered substantially upon fluorination, except for the (S,S) diastereomer. It also turns out that the conformations of the fluorinated peptides are visited during the enhanced MD simulation of natural unguisin A, suggesting thus that conformations accessible to the unsubstituted macrocyclic peptide may be selected by fluorination. Therefore, computational characterization of the macrocyclic peptides could be helpful in the rational design of stereoselective fluorinated peptides with fine-tuned conformation and activity.

Ivermectin to prevent hospitalizations in patients with COVID-19 (IVERCOR-COVID19) a randomized, double-blind, placebo-controlled trial.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) has changed our lives. The scientific community has been investigating re-purposed treatments to prevent disease progression in coronavirus disease (COVID-19) patients. OBJECTIVE: To determine whether ivermectin treatment can prevent hospitalization in individuals with early COVID-19. DESIGN, SETTING AND PARTICIPANTS: A randomized, double-blind, placebo-controlled study was conducted in non-hospitalized individuals with COVID-19 in Corrientes, Argentina. Patients with SARS-CoV-2 positive nasal swabs were contacted within 48 h by telephone to invite them to participate. The trial randomized 501 patients between August 19th 2020 and February 22nd 2021. INTERVENTION: Patients were randomized to ivermectin (N = 250) or placebo (N = 251) arms in a staggered dose, according to the patient's weight, for 2 days. MAIN OUTCOMES AND MEASURES: The efficacy of ivermectin to prevent hospitalizations was evaluated as primary outcome. We evaluated secondary outcomes in relationship to safety and other efficacy end points. RESULTS: The mean age was 42 years (SD ± 15.5) and the median time since symptom onset to the inclusion was 4 days [interquartile range 3-6]. The primary outcome of hospitalization was met in 14/250 (5.6%) individuals in ivermectin group and 21/251 (8.4%) in placebo group (odds ratio 0.65; 95% confidence interval, 0.32-1.31; p = 0.227). Time to hospitalization was not statistically different between groups. The mean time from study enrollment to invasive mechanical ventilatory support (MVS) was 5.25 days (SD ± 1.71) in ivermectin group and 10 days (SD ± 2) in placebo group, (p = 0.019). There were no statistically significant differences in the other secondary outcomes including polymerase chain reaction test negativity and safety outcomes. LIMITATIONS: Low percentage of hospitalization events, dose of ivermectin and not including only high-risk population. CONCLUSION: Ivermectin had no significant effect on preventing hospitalization of patients with COVID-19. Patients who received ivermectin required invasive MVS earlier in their treatment. No significant differences were observed in any of the other secondary outcomes. TRIAL REGISTRATION: ClinicalTrials.gov NCT04529525 .

[Design of a real and perceived academic workload measurement instrument for health care undergraduate students]. / Diseño y validación de un instrumento de medición de la carga académica de estudiantes de carreras de la salud.

BACKGROUND: A high academic workload may have adverse consequences among university students. AIM: To design and validate an instrument to measure both real and perceived academic workload for health care students. MATERIAL AND METHODS: The questionnaire was designed based on a bibliographic revision and the conduction of two focus groups conformed by undergraduate students from a Faculty of Medicine. Afterwards, it was submitted to qualitative pre-tests. The final instrument consists of a self-applied questionnaire with both a characterization section (10 questions) and one concerning academic workload by subject and semester (five and two questions, respectively). A national and international panel of 14 experts evaluated the survey content's validity. The analysis was performed according to the Content Validity Ratio and the Content Validity Index. RESULTS: The complete instrument was validated with an 84% consensus between the judges. Each section of the instrument was approved separately with a 77% and a 94% agreement, respectively. After being individually analyzed by the judges, each question was validated. The wording of questions was improved taking the experts comments into consideration. CONCLUSIONS: The proposed instrument constitutes a contribution for the measurement of real and perceived academic workload for students.

Alkali and Alkaline-Earth Cations in Complexes with Small Bioorganic Ligands: Ab†Initio Benchmark Calculations and Bond Energy Decomposition.

Herein, we report a computational database for the complexes of alkali [Li(I), Na(I), K(I)] and alkaline-earth [Be(II), Mg(II) and Ca(II)] cations with 25 small ligands with varying charge and donor atoms ("O", "N", and "S") that provides geometries and accurate bond energies useful to analyze metal-ligand interactions in proteins and nucleic acids. The role of the ligand→metal charge transfer, the equilibrium bond distance, the electronegativity of the donor atom, the ligand polarizability, and the relative stability of the complexes are discussed in detail. The interacting quantum atoms (IQA) method is used to decompose the binding energy into electrostatic and quantum mechanical contributions. In addition, bond energies are also estimated by means of multipolar electrostatic calculations. No simple correlation exists between bond energies and structural/electronic descriptors unless the data are segregated by the type of ligand or metal. The electrostatic attraction of some molecules (H2 O, NH3 , CH3 OH) towards the metal cations is well reproduced using their (unrelaxed) atomic multipoles, but the same comparison is much less satisfactory for other ligands (e. g. benzene, thiol/thiolate groups, etc.). Besides providing reference structures and bond energies, the database can contribute to validate molecular mechanics potentials capable of yielding a balanced description of alkali and alkaline-earth metals binding to biomolecules.

Chronic Systemic Inflammation Is Associated With Symptoms of Late-Life Depression: The ARIC Study.

OBJECTIVE: The current study examined how the pattern of systemic inflammation in the decades leading up to late-life relates to depression symptoms in older adults. METHODS: Within the Atherosclerosis Risk in Communities Study, we measured high-sensitivity C-reactive protein (CRP), a nonspecific marker of systemic inflammation, at three visits: 21 years and 14 years before, and concurrent with the assessment of depression symptoms, defined using the 11-item Center for Epidemiologic Studies Depression (CESD) scale. We categorized participants into one of four groups based on their 21-year longitudinal pattern of elevated (≥3 mg/L) versus low (<3 mg/L) CRP (stable low; unstable low; unstable elevated; stable elevated). Analyses excluded participants with suspected depression during midlife. RESULTS: A total of 4,614 participants were included (age at CESD assessment: 75.5 [SD: 5.1]; 59% female; follow-up time: 20.7 years [SD: 1.0]). Compared to participants who maintained low CRP levels (stable low), participants who had elevated CRP at two of three visits (unstable elevated; ß = 0.09; 95% confidence interval [CI]: 0.02, 0.17) and participants who maintained elevated CRP at all three visits (stable elevated; ß = 0.13; 95% CI: 0.05, 0.21) had greater depression symptoms as older adults, after adjusting for confounders. After excluding participants with late-life cognitive impairment, only participants with stable elevated CRP demonstrated significantly greater late-life depression symptoms. In a secondary analysis, stable elevated CRP was associated with increased risk for clinically significant late-life depression symptoms. CONCLUSION: Chronic or repeated inflammation in the decades leading up to older adulthood is associated with late-life depression, even in the context of normal cognition.

Onychomycosis caused by the environmental mold Neoscytalidium dimidiatum in Colombia, and in vitro antifungal susceptibility evaluation.

Neoscytalidium dimidiatum is a plant pathogen, but can also cause onychomycosis. We compared clinical and epidemiological data of cases of onychomycosis caused by N. dimidiatum and Trichophyton rubrum. We also evaluated the in vitro antifungal susceptibility of N. dimidiatum clinical isolates. It was not possible to establish any statistical differences between groups, except the place of residence and the number of affected nails. The results suggest that onychomycosis caused by N. dimidiatum is clinically similar to that caused by T. rubrum; besides, N. dimidiatum has been shown to have low sensitivity to itraconazole, but high to terbinafine. LAY SUMMARY: Cases of onychomycosis caused by Neoscytalidium dimidiatum were studied and compared to cases of onychomycosis caused by T. rubrum. The individuals affected were adults, and the clinical characteristics were not different between groups; accordingly, mycological diagnosis is mandatory.

SARS-CoV-2 Main Protease: A Molecular Dynamics Study.

Herein, we investigate the structure and flexibility of the hydrated SARS-CoV-2 main protease by means of 2.0 µs molecular dynamics (MD) simulations in explicit solvent. After having performed electrostatic pKa calculations on several X-ray structures, we consider both the native (unbound) configuration of the enzyme and its noncovalent complex with a model peptide, Ace-Ala-Val-Leu-Gln∼Ser-Nme, which mimics the polyprotein sequence recognized at the active site. For each configuration, we also study their monomeric and homodimeric forms. The simulations of the unbound systems show that the relative orientation of domain III is not stable in the monomeric form and provide further details about interdomain motions, protomer-protomer interactions, inter-residue contacts, accessibility at the catalytic site, etc. In the presence of the peptide substrate, the monomeric protease exhibits a stable interdomain arrangement, but the relative orientation between the scissile peptide bond and the catalytic dyad is not favorable for catalysis. By means of comparative analysis, we further assess the catalytic impact of the enzyme dimerization, the actual flexibility of the active site region, and other structural effects induced by substrate binding. Overall, our computational results complement previous crystallographic studies on the SARS-CoV-2 enzyme and, together with other simulation studies, should contribute to outline useful structure-activity relationships.

Affinity Calculations of Cyclodextrin Host-Guest Complexes: Assessment of Strengths and Weaknesses of End-Point Free Energy Methods.

The end-point methods like MM/PBSA or MM/GBSA estimate the free energy of a biomolecule by combining its molecular mechanics energy with solvation free energy and entropy terms. On the one hand, their performance largely depends on the particular system of interest, and despite numerous attempts to improve their reliability that have resulted in many variants, there is still no clear alternative to improve their accuracy. On the other hand, the relatively small cyclodextrin host-guest complexes, for which high-quality binding calorimetric data are usually available, are becoming reference models for testing the accuracy of free energy methods. In this work, we further assess the performance of various MM/PBSA-like approaches as applied to cyclodextrin complexes. To this end, we select a set of complexes between ß-cyclodextrin and 57 small organic molecules that has been previously studied with the binding energy distribution analysis method in combination with an implicit solvent model ( Wickstrom, L.; He, P.; Gallicchio, E.; Levy, R. M. J. Chem. Theory Comput. 2013 , 9 , 3136 - 3150 ). For each complex, a conventional 1.0 µs molecular dynamics simulation in explicit solvent is performed. Then we employ semiempirical quantum chemical calculations, several variants of the MM-PB(GB)SA methods, entropy estimations, etc., to assess the reliability of the end-point affinity calculations. The best end-point protocol in this study, which combines DFTB3 energies with entropy corrections, yields estimations of the binding free energies that still have substantial errors (RMSE = 2.2 kcal/mol), but it exhibits a good prediction capacity in terms of ligand ranking ( R2 = 0.66) that is close to or even better than that of rigorous free energy methodologies. Our results can be helpful to discriminate between the intrinsic limitations of the end-point methods and other sources of error, such as the underlying energy and continuum solvation methods.

Fluorine conformational effects characterized by energy decomposition analysis.

Electrostatic and stereoelectronic effects associated with fluorine atoms can be exploited as conformational tools for the design of shape-controlled functional molecules. To gain further insight into the nature and strength of these effects, we use the Interacting Quantum Atoms (IQA) method augmented with the semiclassical pairwise dispersion potential to decompose the conformational energies of fluoro-substituted molecules into fragment-based energy contributions, which include deformation/distortion terms and the electrostatic, exchange-correlation and dispersion interactions. The studied molecules comprise various F-CH2-CH2-X and F-CH2-CO-X systems, as well as selected conformers of an α,ß-difluoro-γ-amino-acid derivative that is potentially useful for the design of shape-controlled bioactive amino acids and peptides. We identify the most relevant exchange-correlation and/or electrostatic interaction terms contributing to the stability of the various conformers, and we show that IQA can be used to assess the gauche/anti or trans/cis preferences in molecules with two or more rotatable bonds as well as to study the roles played by other concomitant effects (e.g., CH/OH/NHF contacts). For the α,ß-difluoro-γ-amino acid derivatives, our theoretical analysis indicates that the gauche/anti and trans/cis effects associated with fluorine bonds can be significantly attenuated by other specific intra-molecular contacts.

Application of the Interacting Quantum Atoms Approach to the S66 and Ionic-Hydrogen-Bond Datasets for Noncovalent Interactions.

The interacting quantum atoms (IQA) method can assess, systematically and in great detail, the strength and physics of both covalent and noncovalent interactions. The lack of a pair density in density functional theory (DFT), which precludes the direct IQA decomposition of the characteristic exchange-correlation energy, has been recently overcome by means of a scaling technique, which can largely expand the applicability of the method. To better assess the utility of the augmented IQA methodology to derive quantum chemical decompositions at the atomic and molecular levels, we report the results of Hartree-Fock (HF) and DFT calculations on the complexes included in the S66 and the ionic H-bond databases of benchmark geometry and binding energies. For all structures, we perform single-point and geometry optimizations using HF and selected DFT methods with triple-ζ basis sets followed by full IQA calculations. Pairwise dispersion energies are accounted for by the D3 method. We analyze the goodness of the HF-D3 and DFT-D3 binding energies, the magnitude of numerical errors, the fragment and atomic distribution of formation energies, etc. It is shown that fragment-based IQA decomposes the formation energies in comparable terms to those of perturbative approaches and that the atomic IQA energies hold the promise of rigorously quantifying atomic and group energy contributions in larger biomolecular systems.

Interacting Quantum Atoms Approach and Electrostatic Solvation Energy: Assessing Atomic and Group Solvation Contributions.

The interacting quantum atoms (IQA) method decomposes the total energy of a molecular system in terms of one- and two-center (atomic) contributions within the context of the quantum theory of atoms in molecules. Here we incorporate electrostatic continuum solvent effects into the IQA energy decomposition. To this end, the interaction between the solute electrostatic potential and the solvent screening charges as defined within the COSMO solvation model is now included in a new version of the PROMOLDEN code, allowing thus to apply IQA in combination with COSMO-quantum chemical methods as well as to partition the electrostatic solvation energy into effective atomic and group contributions. To test the robustness of this approach, we carry out COSMO-HF/aug-cc-pVTZ calculations followed by IQA calculations on more than 400 neutral and ionic solutes extracted from the MNSol database. The computational results reveal a detailed atomic mapping of the electrostatic solvation energy that is useful to assess to what extent the solvation energy can be decomposed into atomic and group contributions of various parts of a solute molecule, as generally assumed by empirical methodologies that estimate solvation energy and/or logP values.

A Cross-Sectional Survey of the Zika Virus and its Prevention: The Knowledge, Thoughts, and Beliefs of a Community of Residents in Caguas, Puerto Rico.

OBJECTIVE: The objective of this study was to explore the knowledge, thoughts, and beliefs regarding the Zika virus and its prevention in a community of residents in the municipality of Caguas, Puerto Rico, and elicit their concerns and perceptions of risk. METHODS: A quantitative, non experimental, descriptive, cross-sectional correlational study was conducted in a community in Caguas, Puerto Rico. A structured questionnaire was administered to a sample of 158 residents, aged 21 and older, who participated voluntarily. The data were analyzed using SPSS version 17 via univariate and bivariate analysis. RESULTS: Of 158 surveyed, 64.6% were women; with a population average of 53.85 years. Of the respondents who believed that they would be affected in some way if they were infected by the Zika virus, over half (52.3%) felt that the virus represented a significant threat to their emotional stability. Of those who perceived emotional threat, 39.5% (n=32) continued to study after completing high school (X2=9.217, p=0.027), 57.9% (n=55) had private health insurance (X2=6.325; p=0.042), and 67.9% (n=55) reported it was little or unlikely to become infected (X2= 6.783; p=0.034). Out of those concerned, 57.4% (n=54) considered Zika very or extremely severe (X2=22.827, p<0.001) and 98.9% (n=93) clean the house surroundings as a preventive measure (X2 = 4.951, p=0.026). Lack of interest was the most common reason identified for not complying with preventive actions by the residents (89.2%). CONCLUSION: The underestimation both of the risk concerning the Zika virus and of its consequences was evident. This study reaffirms the need to develop a network that effectively and constantly communicates risk estimates, doing so while addressing the specific needs within the communities served by that network. Community interventions aimed at improving the benefits of and reducing the risks associated with and the perceived barriers to preventive behaviors are needed.

Ligand Strain and Entropic Effects on the Binding of Macrocyclic and Linear Inhibitors: Molecular Modeling of Penicillopepsin Complexes.

Using extensive molecular dynamics simulations, we investigate the structure and dynamics of the complexes formed between penicillopepsin and two peptidomimetic inhibitors: a linear compound, isovaleryl(P4)-valine(P3)-asparagine(P2)-leucine(P1)-phosphonate-phenylalanine(P1'), and its macrocylic analog that includes a methylene bridge between the Asn(P2) and Phe(P1') side chains. The macrocyclic inhibitor, which has a 420-fold stronger affinity than that of the acyclic one, has been considered to lower the entropic penalty for binding. To better understand this binding preference, the solution structure of the inhibitors is studied by molecular dynamics simulations. Subsequently, we assess the influence of the enzyme/inhibitor contacts, the enzyme-induced inhibitor strain, the variation of the ligand configurational entropy and the enzyme reorganization by combining molecular-mechanics Poisson-Boltzmann surface area and normal mode calculations with conformational entropy calculations. We find that there is no relevant entropic stabilization on the binding of the cyclic inhibitor with respect to the acyclic analog because the methylene bridge does not reduce appreciably the conformational flexibility of the free inhibitor. The most important factors explaining the stronger affinity of the macrocyclic inhibitor are the conformational filtering and the lower ligand strain induced by the methylene bridge.

Conformational and entropy analyses of extended molecular dynamics simulations of α-, ß- and γ-cyclodextrins and of the ß-cyclodextrin/nabumetone complex.

Herein, we report the results of 5.0 µs molecular dynamics simulations of native α-, ß- and γ-cyclodextrins (CDs) in explicit water solvent that are useful to describe, in a comparative manner, the distorted geometry of the CD molecules in aqueous solution, the width and fluctuations of their cavities, and the number of cavity waters. By discretizing the time evolution of the dihedral angles, the rate of conformational change of the torsional motions and the conformational entropy are calculated for the three CDs, thus allowing the analysis of the extent of the MD sampling and the entropic significance of the CD flexibility. To obtain a first estimation of the conformational and entropy changes in the host molecule upon ligand binding, the inclusion complex formed between ß-CD and nabumetone is also studied. Overall, the simulations complement previous experimental results on the structure and dynamics of native CDs, and together with the results obtained for the inclusion complex, provide insight into the entropic effects at work on the binding equilibria between CDs and guest ligands.

Rab5 is required in metastatic cancer cells for Caveolin-1-enhanced Rac1 activation, migration and invasion.

Rab5 is a small GTPase that regulates early endosome trafficking and other cellular processes, including cell adhesion and migration. Specifically, Rab5 promotes Rac1 activation and cancer cell migration, but little is known about the upstream regulators of Rab5. We have previously shown that the scaffolding protein Caveolin-1 (CAV1) promotes Rac1 activation and migration of cancer cells. Here, we hypothesized that CAV1 stimulates Rab5 activation, leading to increased Rac1 activity and cell migration. Expression of CAV1 in B16-F10 mouse melanoma and HT-29(US) human colon adenocarcinoma cells increased the GTP loading of Rab5, whereas shRNA-mediated targeting of endogenous CAV1 in MDA-MB-231 breast cancer cells decreased Rab5-GTP levels. Accordingly, shRNA-mediated downregulation of Rab5 decreased CAV1-mediated Rac1 activation, cell migration and invasion in B16-F10 and HT-29(US) cells. Expression of CAV1 was accompanied by increased recruitment of Tiam1, a Rac1 guanine nucleotide exchange factor (GEF), to Rab5-positive early endosomes. Using the inhibitor NSC23766, Tiam1 was shown to be required for Rac1 activation and cell migration induced by CAV1 and Rab5. Mechanistically, we provide evidence implicating p85α (also known as PIK3R1), a Rab5 GTPase-activating protein (GAP), in CAV1-dependent effects, by showing that CAV1 recruits p85α, precluding p85α-mediated Rab5 inactivation and increasing cell migration. In summary, these studies identify a novel CAV1-Rab5-Rac1 signaling axis, whereby CAV1 prevents Rab5 inactivation, leading to increased Rac1 activity and enhanced tumor cell migration and invasion.