Development of machine learning models based on molecular fingerprints for selection of small molecule inhibitors against JAK2 protein.

Janus kinase 2 (JAK2) is emerging as a potential therapeutic target for many inflammatory diseases such as myeloproliferative disorders (MPD), cancer and rheumatoid arthritis (RA). In this study, we have collected experimental data of JAK2 protein containing 6021 unique inhibitors. We then characterized them based on Morgan (ECFP6) fingerprints followed by clustering into training and test set based on their molecular scaffolds. These data were used to build the classification models with various supervised machine learning (ML) algorithms that could prioritize novel inhibitors for future drug development against JAK2 protein. The best model built by Random Forest (RF) and Morgan fingerprints achieved the G-mean value of 0.84 on the external test set. As an application of our classification model, virtual screening was performed against Drugbank molecules in order to identify the potential inhibitors based on the confidence score by RF model. Nine potential molecules were identified, which were further subject to molecular docking studies to evaluate the virtual screening results of the best RF model. This proposed method can prove useful for developing novel target-specific JAK2 inhibitors.

Few-body hydrodynamic interactions probed by optical trap pulling experiment.

We study the hydrodynamic coupling of neighboring micro-beads placed in a multiple optical trap setup allowing us to precisely control the degree of coupling and directly measure time-dependent trajectories of entrained beads. We performed measurements on configurations with increasing complexity starting with a pair of entrained beads moving in one dimension, then in two dimensions, and finally a triplet of beads moving in two dimensions. The average experimental trajectories of a probe bead compare well with the theoretical computation, illustrating the role of viscous coupling and setting timescales for probe bead relaxation. The findings also provide direct experimental corroborations of hydrodynamic coupling at large, micrometer spatial scales and long, millisecond timescales, of relevance to, e.g., microfluidic device design and hydrodynamic-assisted colloidal assembly, improving the capability of optical tweezers, and understanding the coupling between micrometer-scale objects within a living cell.

In Silico Screening of Quorum Sensing Inhibitor Candidates Obtained by Chemical Similarity Search.

Quorum sensing (QS) is a bacterial communication using signal molecules, by which they sense population density of their own species, leading to group behavior such as biofilm formation and virulence. Autoinducer-2 (AI2) is a QS signal molecule universally used by both gram-positive and gram-negative bacteria. Inhibition of QS mediated by AI2 is important for various practical applications, including prevention of gum-disease caused by biofilm formation of oral bacteria. In this research, molecular docking and molecular dynamics (MD) simulations were performed for molecules that are chemically similar to known AI2 inhibitors that might have a potential to be quorum sensing inhibitors. The molecules that form stable complexes with the AI2 receptor protein were found, suggesting that they could be developed as a novel AI2 inhibitors after further in vitro validation. The result suggests that combination of ligand-based drug design and computational methods such as MD simulation, and experimental verification, may lead to development of novel AI inhibitor, with a broad range of practical applications.

Hydrodynamic interaction facilitates the unsteady transport of two neighboring vesicles.

We obtain a numerical solution of the equation for the synchronous unsteady motion of two spherical vesicles in incompressible viscous fluid in the presence of both Stokes drag and hydrodynamics memory. We find that for a given amount of work performed, the final distance traveled by each vesicle is increased by the presence of the other vesicle moving in the same direction. The result suggests that the unsteady transport of the vesicles by molecular motors in vivo may be facilitated due to an effective hydrodynamic interaction between the neighboring vesicles.

Bifidobacterium pseudolongum in the Ceca of Rats Fed Hi-Maize Starch Has Characteristics of a Keystone Species in Bifidobacterial Blooms.

Starches resistant to mammalian digestion are present in foods and pass to the large bowel, where they may be degraded and fermented by the microbiota. Increases in relative abundances of bifidobacteria (blooms) have been reported in rats whose diet was supplemented with Hi-Maize resistant starch. We determined that the bifidobacterial species present in the rat cecum under these circumstances mostly belonged to Bifidobacterium animalis However, cultures of B. animalis isolated from the rats failed to degrade Hi-Maize starch to any extent. In contrast, Bifidobacterium pseudolongum also detected in the rat microbiota had high starch-degrading ability. Transcriptional comparisons showed increased expression of a type 1 pullulanase, alpha-amylase, and glycogen debranching enzyme by B. pseudolongum when cultured in medium containing Hi-Maize starch. Maltose was released into the culture medium, and B. animalis cultures had shorter doubling times in maltose medium than did B. pseudolongum Thus, B. pseudolongum, which was present at a consistently low abundance in the microbiota, but which has extensive enzymatic capacity to degrade resistant starch, showed the attributes of a keystone species associated with the bifidobacterial bloom.IMPORTANCE This study addresses the microbiology and function of a natural ecosystem (the rat gut) using DNA-based observations and in vitro experimentation. The microbial community of the large bowel of animals, including humans, has been studied extensively through the use of high-throughput DNA sequencing methods and advanced bioinformatics analysis. These studies reveal the compositions and genetic capacities of microbiotas but not the intricacies of how microbial communities function. Our work, combining DNA sequence analysis and laboratory experiments with cultured strains of bacteria, revealed that the increased abundance of bifidobacteria in the rat gut, induced by feeding indigestible starch, involved a species that cannot itself degrade the starch (Bifidobacterium animalis) but cohabits with a species that can (Bifidobacterium pseudolongum). B. pseudolongum has the characteristics of a keystone species in the community because it had low abundance but high ability to perform a critical function, the hydrolysis of resistant starch.

Method for identification of rigid domains and hinge residues in proteins based on exhaustive enumeration.

Many proteins undergo large-scale motions where relatively rigid domains move against each other. The identification of rigid domains, as well as the hinge residues important for their relative movements, is important for various applications including flexible docking simulations. In this work, we develop a method for protein rigid domain identification based on an exhaustive enumeration of maximal rigid domains, the rigid domains not fully contained within other domains. The computation is performed by mapping the problem to that of finding maximal cliques in a graph. A minimal set of rigid domains are then selected, which cover most of the protein with minimal overlap. In contrast to the results of existing methods that partition a protein into non-overlapping domains using approximate algorithms, the rigid domains obtained from exact enumeration naturally contain overlapping regions, which correspond to the hinges of the inter-domain bending motion. The performance of the algorithm is demonstrated on several proteins.

RNA-stable-isotope probing shows utilization of carbon from inulin by specific bacterial populations in the rat large bowel.

Knowledge of the trophisms that underpin bowel microbiota composition is required in order to understand its complex phylogeny and function. Stable-isotope ((13)C)-labeled inulin was added to the diet of rats on a single occasion in order to detect utilization of inulin-derived substrates by particular members of the cecal microbiota. Cecal digesta from Fibruline-inulin-fed rats was collected prior to (0 h) and at 6, 12, 18 and 24 h following provision of the [(13)C]inulin diet. RNA was extracted from these cecal specimens and fractionated in isopycnic buoyant density gradients in order to detect (13)C-labeled nucleic acid originating in bacterial cells that had metabolized the labeled dietary constituent. RNA extracted from specimens collected after provision of the labeled diet was more dense than 0-h RNA. Sequencing of 16S rRNA genes amplified from cDNA obtained from these fractions showed that Bacteroides uniformis, Blautia glucerasea, Clostridium indolis, and Bifidobacterium animalis were the main users of the (13)C-labeled substrate. Culture-based studies of strains of these bacterial species enabled trophisms associated with inulin and its hydrolysis products to be identified. B. uniformis utilized Fibruline-inulin for growth, whereas the other species used fructo-oligosaccharide and monosaccharides. Thus, RNA-stable-isotope probing (RNA-SIP) provided new information about the use of carbon from inulin in microbiota metabolism.

Integrated epigenomics identifies BMP4 as a modulator of cisplatin sensitivity in gastric cancer.

OBJECTIVE: Cisplatin is a widely used gastric cancer (GC) chemotherapy; however, genetic factors regulating GC responses to cisplatin remain obscure. Identifying genes regulating cisplatin resistance could aid clinicians in tailoring treatments, by distinguishing cisplatin sensitive patients from those who might benefit from alternative platinum therapies, and highlight novel targeted strategies for overcoming cisplatin resistance. Here integrated epigenomics is applied to identify genes associated with GC cisplatin resistance. DESIGN: 20 GC cell lines were subjected to gene expression profiling, DNA methylation profiling and drug response assays. The molecular data were integrated to identify genes highly expressed and unmethylated specifically in cisplatin-resistant lines. Candidate genes were functionally tested by several in vitro and in vivo assays. Clinical impact of candidate genes was also assessed in a cohort of 197 GC patients. RESULTS: Epigenomic analysis identified bone morphogenetic protein 4 (BMP4) as an epigenetically regulated gene highly expressed in cisplatin-resistant lines. Functional assays confirmed that BMP4 is necessary and sufficient for the expression of several prooncogenic traits, likely mediated through stimulation of the epithelial-mesenchymal transition. In primary tumours, BMP4 promoter methylation levels were inversely correlated with BMP4 expression, and patients with high BMP4-expressing tumours exhibited significantly worse prognosis. Therapeutically, targeted genetic inhibition of BMP4 caused significant sensitisation of GC cells to cisplatin. Notably, BMP4-expressing GCs also did not exhibit cross resistance to oxaliplatin. CONCLUSIONS: BMP4 epigenetic and expression status may represent promising biomarkers for GC cisplatin resistance. Targeting BMP4 may sensitise GC cells to cisplatin. Oxaliplatin, a clinically acceptable cisplatin alternative, may represent a potential therapeutic option for BMP4-positive GCs.

Comprehensive genomic meta-analysis identifies intra-tumoural stroma as a predictor of survival in patients with gastric cancer.

OBJECTIVE: Gastric adenocarcinoma (gastric cancer, GC) is a major cause of global cancer mortality. Identifying molecular programmes contributing to GC patient survival may improve our understanding of GC pathogenesis, highlight new prognostic factors and reveal novel therapeutic targets. The authors aimed to produce a comprehensive inventory of gene expression programmes expressed in primary GCs, and to identify those expression programmes significantly associated with patient survival. DESIGN: Using a network-modelling approach, the authors performed a large-scale meta-analysis of GC transcriptome data integrating 940 gastric transcriptomes from multiple independent patient cohorts. The authors analysed a training set of 428 GCs and 163 non-malignant gastric samples, and a validation set of 288 GCs and 61 non-malignant gastric samples. RESULTS: The authors identified 178 gene expression programmes ('modules') expressed in primary GCs, which were associated with distinct biological processes, chromosomal location patterns, cis-regulatory motifs and clinicopathological parameters. Expression of a transforming growth factor ß (TGF-ß) signalling associated 'super-module' of stroma-related genes consistently predicted patient survival in multiple GC validation cohorts. The proportion of intra-tumoural stroma, quantified by morphometry in tissue sections from gastrectomy specimens, was also significantly associated with stromal super-module expression and GC patient survival. CONCLUSION: Stromal gene expression predicts GC patient survival in multiple independent cohorts, and may be closely related to the intra-tumoural stroma proportion, a specific morphological GC phenotype. These findings suggest that therapeutic approaches targeting the GC stroma may merit evaluation.

Further Validity Evidence for Patient-Specific Virtual Reality Temporal Bone Surgical Simulation.

OBJECTIVE: Patient-specific virtual reality (VR) simulation of cochlear implant (CI) surgery potentially enables preoperative rehearsal and planning. We aim to gather supporting validity evidence for patient-specific simulation through the analysis of virtual performance and comparison with postoperative imaging. METHODS: Prospective, multi-institutional study. Pre- and postoperative cone-beam CT scans of CI surgical patients were obtained and processed for patient-specific VR simulation. The virtual performances of five trainees and four attendings were recorded and (1) compared with volumes removed during actual surgery as determined in postoperative imaging, and (2) assessed using the Copenhagen Cochlear Implant Surgery Assessment Tool (CISAT) by two blinded raters. The volumes compared were cortical mastoidectomy, facial recess, and round window (RW) cochleostomy as well as violation of the facial nerve and chorda. RESULTS: Trainees drilled more volume in the cortical mastoidectomy and facial recess, whereas attendings drilled more volume for the RW cochleostomy and made more violations. Except for the cochleostomy, attendings removed volumes closer to that determined in postoperative imaging. Trainees achieved a higher CISAT performance score compared with attendings (22.0 vs. 18.4 points) most likely due to lack of certain visual cues. CONCLUSION: We found that there were differences in performance of trainees and attendings in patient-specific VR simulation of CI surgery as assessed by raters and in comparison with actual drilled volumes. The presented approach of volume comparison is novel and might be used for further validation of patient-specific VR simulation before clinical implementation for preoperative rehearsal in temporal bone surgery. LEVEL OF EVIDENCE: n/a Laryngoscope, 134:1403-1409, 2024.

Variability in Manual Segmentation of Temporal Bone Structures in Cone Beam CT Images.

PURPOSE: Manual segmentation of anatomical structures is the accepted "gold standard" for labeling structures in clinical images. However, the variability in manual segmentation of temporal bone structures in CBCT images of the temporal bone has not been systematically evaluated using multiple reviewers. Therefore, we evaluated the intravariability and intervariability of manual segmentation of inner ear structures in CBCT images of the temporal bone. METHODS: Preoperative CBCTs scans of the inner ear were obtained from 10 patients who had undergone cochlear implant surgery. The cochlea, facial nerve, chorda tympani, mid-modiolar (MM) axis, and round window (RW) were manually segmented by five reviewers in two separate sessions that were at least 1 month apart. Interreviewer and intrareviewer variabilities were assessed using the Dice coefficient (DICE), volume similarity, mean Hausdorff Distance metrics, and visual review. RESULTS: Manual segmentation of the cochlea was the most consistent within and across reviewers with a mean DICE of 0.91 (SD = 0.02) and 0.89 (SD = 0.01) respectively, followed by the facial nerve with a mean DICE of 0.83 (SD = 0.02) and 0.80 (SD = 0.03), respectively. The chorda tympani had the greatest amount of reviewer variability due to its thin size, and the location of the centroid of the RW and the MM axis were also quite variable between and within reviewers. CONCLUSIONS: We observed significant variability in manual segmentation of some of the temporal bone structures across reviewers. This variability needs to be considered when interpreting the results in studies using one manual reviewer.

Exact partition function zeros of the Wako-Saitô-Muñoz-Eaton protein model.

I compute exact partition function zeros of the Wako-Saitô-Muñoz-Eaton model for various secondary structural elements and for two proteins, 1BBL and 1I6C, by using both analytic and numerical methods. Two-state and barrierless downhill folding transitions can be distinguished by a gap in the distribution of zeros at the positive real axis.

Nonadditive entropies yield probability distributions with biases not warranted by the data.

Different quantities that go by the name of entropy are used in variational principles to infer probability distributions from limited data. Shore and Johnson showed that maximizing the Boltzmann-Gibbs form of the entropy ensures that probability distributions inferred satisfy the multiplication rule of probability for independent events in the absence of data coupling such events. Other types of entropies that violate the Shore and Johnson axioms, including nonadditive entropies such as the Tsallis entropy, violate this basic consistency requirement. Here we use the axiomatic framework of Shore and Johnson to show how such nonadditive entropy functions generate biases in probability distributions that are not warranted by the underlying data.

Bowel microbiota moderate host physiological responses to dietary konjac in weanling rats.

Diets rich in complex carbohydrates that resist digestion in the small bowel can alter large bowel ecology and microbiota biochemistry because the carbohydrates become substrates for bacterial growth and metabolism. Conventional or germ-free weanling rats were fed a control diet or diets containing 1.25, 2.5, or 5% konjac (KJ), a commonly used ingredient in Asian foods, for 28 d. In the absence of bowel microbiota, 5% KJ elicited a significant increase in colonic goblet cell numbers and increased expression of mast cell protease genes and of genes that were overrepresented in the KEGG pathway "Metabolism of xenobiotics by cytochrome P450" relative to the control diet. In contrast, feeding 5% KJ caused few changes in mucosal gene expression in conventional rats. Analysis of the colonic microbiota of conventional rats fed KJ showed modest increases in the proportions of Actinobacteria and Bacteroidetes relative to rats fed the control diet, with a concomitant reduction in Firmicutes, which included a 50% reduction in Lactobacillus abundance. Colonic concentrations of short-chain fatty acids and colonic crypt lengths were increased by feeding KJ. Goblet cell numbers were greater in conventional rats fed KJ relative to the control diet but were lower compared with germ-free animals. Serum metabolite profiles were different in germ-free and conventional rats. Metabolites that differed in concentration included several phospholipids, a bile acid metabolite, and an intermediate product of tryptophan metabolism. Overall, KJ in the diet was potentially damaging to the bowel mucosa and produced a protective response from the host. This response was reduced by the presence of the bowel microbiota, which therefore ameliorated potentially detrimental effects of dietary KJ.

The FALC-Loop web server for protein loop modeling.

The FALC-Loop web server provides an online interface for protein loop modeling by employing an ab initio loop modeling method called FALC (fragment assembly and analytical loop closure). The server may be used to construct loop regions in homology modeling, to refine unreliable loop regions in experimental structures or to model segments of designed sequences. The FALC method is computationally less expensive than typical ab initio methods because the conformational search space is effectively reduced by the use of fragments derived from a structure database. The analytical loop closure algorithm allows efficient search for loop conformations that fit into the protein framework starting from the fragment-assembled structures. The FALC method shows prediction accuracy comparable to other state-of-the-art loop modeling methods. Top-ranked model structures can be visualized on the web server, and an ensemble of loop structures can be downloaded for further analysis. The web server can be freely accessed at http://falc-loop.seoklab.org/.

Characterization of the common bed bug's eggshell and egg glue proteins.

An insect egg is one of the most vulnerable stages of insect life, and the evolutionary success of a species depends on the eggshell protecting the embryo and the egg glue securing the attachment. The common bed bug (Cimex lectularius), notorious for its painful and itchy bites, infests human dwellings to feed on blood. They are easier to find these days as they adapt to develop resistance against commonly used insecticides. In this study, we identify and characterize the eggshell protein and the probable egg glue protein (i.e. keratin associated protein 5-10 like protein) of the bed bug by using mass spectrometry and bioinformatics analysis. Furthermore, by using transcription profiling and in vivo RNA interference, we show evidences that the keratin associated protein 5-10 like protein functions as the glue protein. Finally, structural characterizations on the two proteins are performed using recombinant proteins. Amino acid sequences of various insect eggshell and egg glue proteins support their independent evolution among different insect groups. Hence, inhibiting the function of these proteins related to the earliest stage of life can achieve species-specific population control. In this respect, our results would be a starting point in developing new ways to control bed bug population.

Mental health-related presentations to a tertiary emergency department during the COVID-19 pandemic.

Introduction: The coronavirus disease 2019 (COVID-19) pandemic has caused significant mental distress in populations globally. At the frontline of the pandemic, emergency departments (EDs) are the prime setting to observe the effects of the pandemic on the mental health of the population. We aimed to describe the trend of mental health-related ED attendances at an acute hospital in Singapore before and during the various stages of the COVID-19 pandemic. Methods: This is a retrospective, descriptive study of patients who presented to the ED between 1 January 2019 and 31 December 2020. Patients diagnosed with mental health-related systematised nomenclature of medicine who visited the ED during this period were identified and were placed into mental health diagnosis categories for analysis. A comparison was made between patients who presented before the pandemic (2019) and during the pandemic (2020). Results: During the study periods, we identified 1,421 patients, of whom 27 were excluded due to non-mental health-related diagnoses, leaving 1,394 patients for analysis. There was a 36.7% increase in mental health-related ED presentations from 2019 to 2020. The proportion of higher-acuity mental health-related ED attendances and number of suicide attempts also increased. Conclusion: Our study described an increase in the proportion of high-acuity mental health-related ED attendances during the COVID-19 pandemic. Emergency physicians must be cognisant of the effects of the pandemic on mental health. Further research should be conducted to better equip the healthcare system for handling all aspects of the pandemic.

Intrinsic subtypes of gastric cancer, based on gene expression pattern, predict survival and respond differently to chemotherapy.

BACKGROUND & AIMS: Gastric cancer (GC) is a heterogeneous disease comprising multiple subtypes that have distinct biological properties and effects in patients. We sought to identify new, intrinsic subtypes of GC by gene expression analysis of a large panel of GC cell lines. We tested if these subtypes might be associated with differences in patient survival times and responses to various standard-of-care cytotoxic drugs. METHODS: We analyzed gene expression profiles for 37 GC cell lines to identify intrinsic GC subtypes. These subtypes were validated in primary tumors from 521 patients in 4 independent cohorts, where the subtypes were determined by either expression profiling or subtype-specific immunohistochemical markers (LGALS4, CDH17). In vitro sensitivity to 3 chemotherapy drugs (5-fluorouracil, cisplatin, oxaliplatin) was also assessed. RESULTS: Unsupervised cell line analysis identified 2 major intrinsic genomic subtypes (G-INT and G-DIF) that had distinct patterns of gene expression. The intrinsic subtypes, but not subtypes based on Lauren's histopathologic classification, were prognostic of survival, based on univariate and multivariate analysis in multiple patient cohorts. The G-INT cell lines were significantly more sensitive to 5-fluorouracil and oxaliplatin, but more resistant to cisplatin, than the G-DIF cell lines. In patients, intrinsic subtypes were associated with survival time following adjuvant, 5-fluorouracil-based therapy. CONCLUSIONS: Intrinsic subtypes of GC, based on distinct patterns of expression, are associated with patient survival and response to chemotherapy. Classification of GC based on intrinsic subtypes might be used to determine prognosis and customize therapy.

Changes in bowel microbiota induced by feeding weanlings resistant starch stimulate transcriptomic and physiological responses.

The ability to predictably engineer the composition of bowel microbial communities (microbiota) using dietary components is important because of the reported associations of altered microbiota composition with medical conditions. In a synecological study, weanling conventional Sprague-Dawley rats (21 days old) were fed a basal diet (BD) or a diet supplemented with resistant starch (RS) at 5%, 2.5%, or 1.25% for 28 days. Pyrosequencing of 16S rRNA genes and temporal temperature gradient electrophoresis (TTGE) profiles in the colonic digesta showed that rats fed RS had altered microbiota compositions due to blooms of Bacteroidetes and Actinobacteria. The altered microbiota was associated with changes in colonic short-chain fatty acid (SCFA) concentrations, colonic-tissue gene expression (Gsta2 and Ela1), and host physiology (serum metabolite profiles and colonic goblet cell numbers). Comparisons between germ-free and conventional rats showed that transcriptional and serum metabolite differences were mediated by the microbiota and were not the direct result of diet composition. Altered transcriptomic and physiological responses may reflect the young host's attempts to maintain homeostasis as a consequence of exposure to a new collection of bacteria and their associated biochemistry.

A derivation of the master equation from path entropy maximization.

The master equation and, more generally, Markov processes are routinely used as models for stochastic processes. They are often justified on the basis of randomization and coarse-graining assumptions. Here instead, we derive nth-order Markov processes and the master equation as unique solutions to an inverse problem. We find that when constraints are not enough to uniquely determine the stochastic model, an nth-order Markov process emerges as the unique maximum entropy solution to this otherwise underdetermined problem. This gives a rigorous alternative for justifying such models while providing a systematic recipe for generalizing widely accepted stochastic models usually assumed to follow from the first principles.