A Small Molecule RAS-Mimetic Disrupts RAS Association with Effector Proteins to Block Signaling.

Oncogenic activation of RAS genes via point mutations occurs in 20%-30% of human cancers. The development of effective RAS inhibitors has been challenging, necessitating new approaches to inhibit this oncogenic protein. Functional studies have shown that the switch region of RAS interacts with a large number of effector proteins containing a common RAS-binding domain (RBD). Because RBD-mediated interactions are essential for RAS signaling, blocking RBD association with small molecules constitutes an attractive therapeutic approach. Here, we present evidence that rigosertib, a styryl-benzyl sulfone, acts as a RAS-mimetic and interacts with the RBDs of RAF kinases, resulting in their inability to bind to RAS, disruption of RAF activation, and inhibition of the RAS-RAF-MEK pathway. We also find that ribosertib binds to the RBDs of Ral-GDS and PI3Ks. These results suggest that targeting of RBDs across multiple signaling pathways by rigosertib may represent an effective strategy for inactivation of RAS signaling.

Identification of a Novel Keto Sugar Component in Streptococcus pneumoniae Serotype 12F Capsular Polysaccharide and Impact on Vaccine Immunogenicity.

Implementation of conjugate vaccine technology revolutionized the ability to effectively elicit long-lasting immune responses to bacterial capsular polysaccharides. Although expansion of conjugate vaccine serotype coverage is designed to target residual disease burden to pneumococcal serotypes not contained in earlier vaccine versions, details of polysaccharide Ag structure, heterogeneity, and epitope structure components contributing to vaccine-mediated immunity are not always clear. Analysis of Streptococcus pneumoniae serotype 12F polysaccharide by two-dimensional nuclear magnetic resonance spectroscopy and mass spectrometry revealed a partial substitution of N-acetyl-galactosamine by the keto sugar 2-acetamido-2,6-dideoxy-xylo-hexos-4-ulose (Sug) in up to 25% of the repeat units. This substitution was not described in previous published structures for 12F. Screening a series of contemporary 12F strains isolated from humans (n = 17) identified Sug incorporation at varying levels in all strains examined. Thus, partial Sug substitution in S. pneumoniae serotype 12F may have always been present but is now detectable by state-of-the-art analytical techniques. During the steps of conjugation, the serotype 12F Sug epitope is modified by reduction, and both polysaccharide PPSV23 and conjugate PCV20 vaccines contain 12F Ags with little to no Sug epitope. Both PCV20 and PPSV23 vaccines were evaluated for protection against circulating 12F strains with varying amounts of Sug in their repeat unit based on an opsonophagocytic killing assay involving HL-60 cells and rabbit complement. Both vaccines elicited human-derived neutralizing Abs against serotype 12F, independent of Sug level between ∼2 and 25 mol%. These findings suggest that the newly identified serotype 12F Sug epitope is likely not an essential epitope for vaccine-elicited protection.

Usability of Health Care Price Transparency Data in the United States: Mixed Methods Study.

BACKGROUND: Increasing health care expenditure in the United States has put policy makers under enormous pressure to find ways to curtail costs. Starting January 1, 2021, hospitals operating in the United States were mandated to publish transparent, accessible pricing information online about the items and services in a consumer-friendly format within comprehensive machine-readable files on their websites. OBJECTIVE: The aims of this study are to analyze the available files on hospitals' websites, answering the question-is price transparency (PT) information as provided usable for patients or for machines?-and to provide a solution. METHODS: We analyzed 39 main hospitals in Florida that have published machine-readable files on their website, including commercial carriers. We created an Excel (Microsoft) file that included those 39 hospitals along with the 4 most popular services-Current Procedural Terminology (CPT) 45380, 29827, and 70553 and Diagnosis-Related Group (DRG) 807-for the 4 most popular commercial carriers (Health Maintenance Organization [HMO] or Preferred Provider Organization [PPO] plans)-Aetna, Florida Blue, Cigna, and UnitedHealthcare. We conducted an A/B test using 67 MTurkers (randomly selected from US residents), investigating the level of awareness about PT legislation and the usability of available files. We also suggested format standardization, such as master field names using schema integration, to make machine-readable files consistent and usable for machines. RESULTS: The poor usability and inconsistent formats of the current PT information yielded no evidence of its usefulness for patients or its quality for machines. This indicates that the information does not meet the requirements for being consumer-friendly or machine readable as mandated by legislation. Based on the responses to the first part of the experiment (PT awareness), it was evident that participants need to be made aware of the PT legislation. However, they believe it is important to know the service price before receiving it. Based on the responses to the second part of the experiment (human usability of PT information), the average number of correct responses was not equal between the 2 groups, that is, the treatment group (mean 1.23, SD 1.30) found more correct answers than the control group (mean 2.76, SD 0.58; t65=6.46; P<.001; d=1.52). CONCLUSIONS: Consistent machine-readable files across all health systems facilitate the development of tools for estimating customer out-of-pocket costs, aligning with the PT rule's main objective-providing patients with valuable information and reducing health care expenditures.

The Effect of Monetary Incentives on Health Care Social Media Content: Study Based on Topic Modeling and Sentiment Analysis.

BACKGROUND: While there is high-quality online health information, a lot of recent work has unfortunately highlighted significant issues with the health content on social media platforms (eg, fake news and misinformation), the consequences of which are severe in health care. One solution is to investigate methods that encourage users to post high-quality content. OBJECTIVE: Incentives have been shown to work in many domains, but until recently, there was no method to provide financial incentives easily on social media for users to generate high-quality content. This study investigates the following question: What effect does the provision of incentives have on the creation of social media health care content? METHODS: We analyzed 8328 health-related posts from an incentive-based platform (Steemit) and 1682 health-related posts from a traditional platform (Reddit). Using topic modeling and sentiment analysis-based methods in machine learning, we analyzed these posts across the following 3 dimensions: (1) emotion and language style using the IBM Watson Tone Analyzer service, (2) topic similarity and difference from contrastive topic modeling, and (3) the extent to which posts resemble clickbait. We also conducted a survey using 276 Amazon Mechanical Turk (MTurk) users and asked them to score the quality of Steemit and Reddit posts. RESULTS: Using the Watson Tone Analyzer in a sample of 2000 posts from Steemit and Reddit, we found that more than double the number of Steemit posts had a confident language style compared with Reddit posts (77 vs 30). Moreover, 50% more Steemit posts had analytical content and 33% less Steemit posts had a tentative language style compared with Reddit posts (619 vs 430 and 416 vs 627, respectively). Furthermore, more than double the number of Steemit posts were considered joyful compared with Reddit posts (435 vs 200), whereas negative posts (eg, sadness, fear, and anger) were 33% less on Steemit than on Reddit (384 vs 569). Contrastive topic discovery showed that only 20% (2/10) of topics were common, and Steemit had more unique topics than Reddit (5 vs 3). Qualitatively, Steemit topics were more informational, while Reddit topics involved discussions, which may explain some of the quantitative differences. Manual labeling marked more Steemit headlines as clickbait than Reddit headlines (66 vs 26), and machine learning model labeling consistently identified a higher percentage of Steemit headlines as clickbait than Reddit headlines. In the survey, MTurk users said that at least 57% of Steemit posts had better quality than Reddit posts, and they were at least 52% more likely to like and comment on Steemit posts than Reddit posts. CONCLUSIONS: It is becoming increasingly important to ensure high-quality health content on social media; therefore, incentive-based social media could be important in the design of next-generation social platforms for health information.

Preclinical Immunogenicity and Efficacy of Optimized O25b O-Antigen Glycoconjugates To Prevent MDR ST131 E. coli Infections.

Multivalent O-antigen polysaccharide glycoconjugate vaccines are under development to prevent invasive infections caused by pathogenic Enterobacteriaceae. Sequence type 131 (ST131) Escherichia coli of serotype O25b has emerged as the predominant lineage causing invasive multidrug-resistant extraintestinal pathogenic E. coli (ExPEC) infections. We observed the prevalence of E. coli O25b ST131 among a contemporary collection of isolates from U.S. bloodstream infections from 2013 to 2016 (n = 444) and global urinary tract infections from 2014 to 2017 (n = 102) to be 25% and 24%, respectively. To maximize immunogenicity of the serotype O25b O antigen, we investigated glycoconjugate properties, including CRM197 carrier protein cross-linking (single-end versus cross-linked "lattice") and conjugation chemistry (reductive amination chemistry in dimethyl sulfoxide [RAC/DMSO] versus ((2-((2-oxoethyl)thio)ethyl)carbamate [eTEC] linker). Using opsonophagocytic assays (OPAs) to measure serum functional antibody responses to vaccination, we observed that higher-molecular-mass O25b long-chain lattice conjugates showed improved immunogenicity in mice compared with long- or short-chain O antigens conjugated via single-end attachment. The lattice conjugates protected mice from lethal challenge with acapsular O25b ST131 strains as well as against hypervirulent O25b isolates expressing K5 or K100 capsular polysaccharides. A single 1-µg dose of long-chain O25b lattice conjugate constructed with both chemistries also elicited robust serum IgG and OPA responses in cynomolgus macaques. Our findings show that key properties of the O-antigen carrier protein conjugate such as saccharide epitope density and degree of intermolecular cross-linking can significantly enhance functional immunogenicity.

Impact of Drug Conjugation and Loading on Target Antigen Binding and Cytotoxicity in Cysteine Antibody-Drug Conjugates.

Antibody-drug conjugates (ADCs) consist of a target-specific antibody that is covalently conjugated to a drug via a linker. ADCs are designed to deliver cytotoxic drugs (payloads), specifically to cancer cells, while minimizing systemic toxicity. Conventional cysteine conjugation typically results in the formation of ADC molecules containing a heterogeneous mixture of 2, 4, 6, and 8 drug-loaded species. The drug-to-antibody ratio (DAR) of the mixture represents the weighted average of these species. In this report, we have investigated the impact of the hydrophobicity of payloads and the overall drug loading on the in vitro binding and cytotoxicity of ADC species. Several ADCs were prepared by conventional cysteine conjugation using different payloads. ADC species with different DAR values were purified from the ADC mixture and characterized by standard analytical techniques. These ADC species were evaluated for target antigen binding using an immunoassay, enzyme-linked immunosorbent assay (ELISA). The potency was assessed using a cell-based cytotoxicity assay. These structure-function studies lead to a better understanding of factors that impact the in vitro target binding and cytotoxicity of ADC species. ADC species containing hydrophobic payloads with high DAR were found to have lower target binding by ELISA compared to that of the unconjugated antibody or the heterogeneous reference ADC with DAR ∼4. Under similar assay conditions, the ADCs conjugated to hydrophilic payloads did not show a significant impact on the target binding. The cytotoxic potency of ADC species increased with increasing level of drug loading in the cell-based cytotoxicity assay.

Thermodynamic characterization of the multivalent interactions underlying rapid and selective translocation through the nuclear pore complex.

Intrinsically disordered proteins (IDPs) play important roles in many biological systems. Given the vast conformational space that IDPs can explore, the thermodynamics of the interactions with their partners is closely linked to their biological functions. Intrinsically disordered regions of Phe-Gly nucleoporins (FG Nups) that contain multiple phenylalanine-glycine repeats are of particular interest, as their interactions with transport factors (TFs) underlie the paradoxically rapid yet also highly selective transport of macromolecules mediated by the nuclear pore complex. Here, we used NMR and isothermal titration calorimetry to thermodynamically characterize these multivalent interactions. These analyses revealed that a combination of low per-FG motif affinity and the enthalpy-entropy balance prevents high-avidity interaction between FG Nups and TFs, whereas the large number of FG motifs promotes frequent FG-TF contacts, resulting in enhanced selectivity. Our thermodynamic model underlines the importance of functional disorder of FG Nups. It helps explain the rapid and selective translocation of TFs through the nuclear pore complex and further expands our understanding of the mechanisms of "fuzzy" interactions involving IDPs.

Slide-and-exchange mechanism for rapid and selective transport through the nuclear pore complex.

Nucleocytoplasmic transport is mediated by the interaction of transport factors (TFs) with disordered phenylalanine-glycine (FG) repeats that fill the central channel of the nuclear pore complex (NPC). However, the mechanism by which TFs rapidly diffuse through multiple FG repeats without compromising NPC selectivity is not yet fully understood. In this study, we build on our recent NMR investigations showing that FG repeats are highly dynamic, flexible, and rapidly exchanging among TF interaction sites. We use unbiased long timescale all-atom simulations on the Anton supercomputer, combined with extensive enhanced sampling simulations and NMR experiments, to characterize the thermodynamic and kinetic properties of FG repeats and their interaction with a model transport factor. Both the simulations and experimental data indicate that FG repeats are highly dynamic random coils, lack intrachain interactions, and exhibit significant entropically driven resistance to spatial confinement. We show that the FG motifs reversibly slide in and out of multiple TF interaction sites, transitioning rapidly between a strongly interacting state and a weakly interacting state, rather than undergoing a much slower transition between strongly interacting and completely noninteracting (unbound) states. In the weakly interacting state, FG motifs can be more easily displaced by other competing FG motifs, providing a simple mechanism for rapid exchange of TF/FG motif contacts during transport. This slide-and-exchange mechanism highlights the direct role of the disorder within FG repeats in nucleocytoplasmic transport, and resolves the apparent conflict between the selectivity and speed of transport.

A study on the accuracy of horizontal condylar guidance values in edentulous patients using preprosthetic diagnostic radiographs.

CONTEXT: Determination of horizontal condylar guidance (HCG) by various clinical and radiographic methods was performed by several investigators. If a correlation between HCG values using lateral radiographic tracing and protrusive interocclusal records can be established, the necessity of performing elaborate recording procedures can be eliminated. AIMS: The aim of this study is to evaluate and to compare the correlation between HCG values in edentulous people using the protrusive interocclusal records mounted on a semi-adjustable articulator with the manual tracing of panoramic radiograph and lateral cephalogram. MATERIALS AND METHODS: A total of 20 completely edentulous individuals of either sex from 45 to 75 years (mean age 63.15 years) fulfilling the inclusion criteria were included in this in vivo study. In all the participants, HCG angles were determined clinically using protrusive interocclusal records and semi-adjustable articulator after intraoral gothic arch tracing. Radiographically, it was obtained by cephalometric tracing of panoramic radiograph and lateral cephalogram. RESULTS: The present study shows mean HCG ± standard deviation (SD) of 28.17° ± 5.99° for interocclusal protrusive record while cephalometric tracing method yielded HCG ± SD of 38.95° ± 4.77° and 35.2° ± 4.94° for lateral cephalogram and orthopantomogram, respectively. A statistically significant positive correlation (P < 0.0001) was found among these three methods. CONCLUSION: HCG can be successfully determined in edentulous participants by using three aforementioned methods. HCG values from cephalometric tracing of diagnostic radiographs can be used as an adjunct to the clinical method but cannot be used independently for programming a semi-adjustable articulator.

A comparative study to correlate between clinically and radiographically determined sagittal condylar guidance in participants with different skeletal relationships.

CONTEXT: Many authors have conducted studies that determine horizontal condylar guidance (HCG) using various methods, articulator systems, and recording materials. However, there is a dearth of literature on variability existing in HCG in individuals with different skeletal relationships. This study is an attempt to verify whether such a difference exists or not. AIMS: The aim of this study is to determine and correlate the HCG in individuals with Angle's Class I, Class II, and Class III malocclusion using radiographic and clinical methods. SETTINGS AND DESIGN: HCG was recorded for thirty individuals, ten of each class. For each individual, HCG was recorded clinically as well as radiographically. SUBJECTS AND METHODS: Clinically, HCG was recorded using protrusive check bites and a semi-adjustable articulator. Radiographically, two methods were employed. First, a "tangent method" wherein the angle made by a tangent to the posterior slope of articular eminence with the Frankfurt horizontal (FH) plane was considered as the HCG, and second, a "protrusive method" where the position of the condyle at maximum intercuspation and 6 mm protrusion were traced, and the angle this path made with the FH plane was recorded as the HCG. STATISTICAL ANALYSIS: Descriptive statistical analysis along with Tukey's test and analysis of variance was used to calculate and compare the mean values. Pearson correlation coefficient was used to establish correlation between various means. RESULTS: A significant difference in the HCG of three skeletal relationships was seen, with Class II having a steeper angle than the other two. Among the various methods used, a correlation was found between the clinical and the protrusive method; however, the tangent method yielded greater values of HCG. CONCLUSIONS: The average value of HCG should not be used as it differs according to the skeletal relationship. Radiographic method can be used to yield consistent HCG; however, the protrusive method should be employed.

Mechanisms mediating enhanced neutralization efficacy of staphylococcal enterotoxin B by combinations of monoclonal antibodies.

Staphylococcal enterotoxin B (SEB) is a superantigen that cross-links the major histocompatibility complex class II and specific V-ß chains of the T-cell receptor, thus forming a ternary complex. Developing neutralizing mAb to disrupt the ternary complex and abrogate the resulting toxicity is a major therapeutic challenge because SEB is effective at very low concentrations. We show that combining two SEB-specific mAbs enhances their efficacy, even though one of the two mAbs by itself has no effect on neutralization. Crystallography was employed for fine-mapping conformational epitopes in binary and ternary complexes between SEB and Fab fragments. NMR spectroscopy was used to validate and identify subtle allosteric changes induced by mAbs binding to SEB. The mapping of epitopes established that a combination of different mAbs can enhance efficacy of mAb-mediated protection from SEB induced lethal shock by two different mechanisms: one mAb mixture promoted clearance of the toxin both in vitro and in vivo by FcR-mediated cross-linking and clearance, whereas the other mAb mixture induced subtle allosteric conformational changes in SEB that perturbed formation of the SEB·T-cell receptor·major histocompatibility complex class II trimer. Finally structural information accurately predicted mAb binding to other superantigens that share conformational epitopes with SEB. Fine mapping of conformational epitopes is a powerful tool to establish the mechanism and optimize the action of synergistic mAb combinations.

Use of digital panoramic radiology in presurgical implant treatment planning to accurately assess bone density.

STATEMENT OF PROBLEM: No cost-effective method of ascertaining bone density from 2-dimensional radiographic images is currently available for dental implants before surgery. PURPOSE: The purpose of this in vivo study was to use digital panoramic radiology and dental computed tomography (CT) to evaluate the bone density of specific points in the jaw near the tooth-bearing areas. The objective was to determine whether digital panoramic radiology can be used in assessing bone density as an alternative to a more expensive and complex dental CT. MATERIAL AND METHODS: This study involved determining bone densities at predetermined anatomic landmarks near tooth-bearing areas of the jaws of 20 participants, using digital panoramic radiology in gray-level scale with a lead step wedge. Subsequently, the bone densities of the same points were determined in Hounsfield units (Hu) with dental CT. The data collected after interpretation of the panoramic radiograph and CT were tabulated and analyzed statistically. RESULTS: Bone density measured using CT correlated with the first 3 steps of (A, B, and C) the digital scale of gray. Further analysis conducted using the Mann-Whitney U test showed a significant association between step A to detect D4 bone, step B to detect D3 bone, and step C to detect D2 type bone. CONCLUSIONS: The digital scale of gray obtained from a lead step wedge can be effectively used with digital orthopanoramic radiology to assess bone density before the placement of implants, but with certain restrictions.

Interactions of amelogenin with phospholipids.

Amelogenin protein has the potential to interact with other enamel matrix proteins, mineral, and cell surfaces. We investigated the interactions of recombinant amelogenin rP172 with small unilamellar vesicles as model membranes, toward the goal of understanding the mechanisms of amelogenin-cell interactions during amelogenesis. Dynamic light scattering (DLS), fluorescence spectroscopy, circular dichroism (CD), and nuclear magnetic resonance (NMR) were used. In the presence of phospholipid vesicles, a blue shift in the Trp fluorescence emission maxima of rP172 was observed (∼334 nm) and the Trp residues of rP172 were inaccessible to the aqueous quencher acrylamide. DLS studies indicated complexation of rP172 and phospholipids, although the possibility of fusion of phospholipids following amelogenin addition cannot be ruled out. NMR and CD studies revealed a disorder-order transition of rP172 in a model membrane environment. Strong fluorescence resonance energy transfer from Trp in rP172 to DNS-bound-phospholipid was observed, and fluorescence polarization studies indicated that rP172 interacted with the hydrophobic core region of model membranes. Our data suggest that amelogenin has ability to interact with phospholipids and that such interactions may play key roles in enamel biomineralization as well as reported amelogenin signaling activities.

Identification of Hydrophobic Interfaces in Protein-Ligand Complexes by Selective Saturation Transfer NMR Spectroscopy.

The proper characterization of protein-ligand interfaces is essential for structural biology, with implications ranging from the fundamental understanding of biological processes to pharmacology. Nuclear magnetic resonance is a powerful technique for such studies. We propose a novel approach to the direct determination of the likely pose of a peptide ligand onto a protein partner, by using frequency-selective cross-saturation with a low stringency isotopic labeling methods. Our method illustrates a complex of the Src homology 3 domain of C-terminal Src kinase with a peptide from the proline-enriched tyrosine phosphatase.

Regulatory interactions between a bacterial tyrosine kinase and its cognate phosphatase.

The cyclic process of autophosphorylation of the C-terminal tyrosine cluster (YC) of a bacterial tyrosine kinase and its subsequent dephosphorylation following interactions with a counteracting tyrosine phosphatase regulates diverse physiological processes, including the biosynthesis and export of polysaccharides responsible for the formation of biofilms or virulence-determining capsules. We provide here the first detailed insight into this hitherto uncharacterized regulatory interaction at residue-specific resolution using Escherichia coli Wzc, a canonical bacterial tyrosine kinase, and its opposing tyrosine phosphatase, Wzb. The phosphatase Wzb utilizes a surface distal to the catalytic elements of the kinase, Wzc, to dock onto its catalytic domain (WzcCD). WzcCD binds in a largely YC-independent fashion near the Wzb catalytic site, inducing allosteric changes therein. YC dephosphorylation is proximity-mediated and reliant on the elevated concentration of phosphorylated YC near the Wzb active site resulting from WzcCD docking. Wzb principally recognizes the phosphate of its phosphotyrosine substrate and further stabilizes the tyrosine moiety through ring stacking interactions with a conserved active site tyrosine.

Structural adaptation of tooth enamel protein amelogenin in the presence of SDS micelles.

Amelogenin, the major extracellular matrix protein of developing tooth enamel is intrinsically disordered. Through its interaction with other proteins and mineral, amelogenin assists enamel biomineralization by controlling the formation of highly organized enamel crystal arrays. We used circular dichroism (CD), dynamic light scattering (DLS), fluorescence, and NMR spectroscopy to investigate the folding propensity of recombinant porcine amelogenin rP172 following its interaction with SDS, at levels above critical micelle concentration. The rP172-SDS complex formation was confirmed by DLS, while an increase in the structure moiety of rP172 was noted through CD and fluorescence experiments. Fluorescence quenching analyses performed on several rP172 mutants where all but one Trp was replaced by Tyr at different sequence regions confirmed that the interaction of amelogenin with SDS micelles occurs via the N-terminal region close to Trp25 where helical segments can be detected by NMR. NMR spectroscopy and structural refinement calculations using CS-Rosetta modeling confirm that the highly conserved N-terminal domain is prone to form helical structure when bound to SDS micelles. Our findings reported here reveal interactions leading to significant changes in the secondary structure of rP172 upon treatment with SDS. These interactions may reflect the physiological relevance of the flexible nature of amelogenin and its sequence specific helical propensity that might enable it to structurally adapt with charged and potential targets such as cell surface, mineral, and other proteins during enamel biomineralization.

Solution structure and DNA-binding properties of the phosphoesterase domain of DNA ligase D.

The phosphoesterase (PE) domain of the bacterial DNA repair enzyme LigD possesses distinctive manganese-dependent 3'-phosphomonoesterase and 3'-phosphodiesterase activities. PE exemplifies a new family of DNA end-healing enzymes found in all phylogenetic domains. Here, we determined the structure of the PE domain of Pseudomonas aeruginosa LigD (PaePE) using solution NMR methodology. PaePE has a disordered N-terminus and a well-folded core that differs in instructive ways from the crystal structure of a PaePE•Mn(2+)• sulfate complex, especially at the active site that is found to be conformationally dynamic. Chemical shift perturbations in the presence of primer-template duplexes with 3'-deoxynucleotide, 3'-deoxynucleotide 3'-phosphate, or 3' ribonucleotide termini reveal the surface used by PaePE to bind substrate DNA and suggest a more efficient engagement in the presence of a 3'-ribonucleotide. Spectral perturbations measured in the presence of weakly catalytic (Cd(2+)) and inhibitory (Zn(2+)) metals provide evidence for significant conformational changes at and near the active site, compared to the relatively modest changes elicited by Mn(2+).

Deep learning generation of preclinical positron emission tomography (PET) images from low-count PET with task-based performance assessment.

BACKGROUND: Preclinical low-count positron emission tomography (LC-PET) imaging offers numerous advantages such as facilitating imaging logistics, enabling longitudinal studies of long- and short-lived isotopes as well as increasing scanner throughput. However, LC-PET is characterized by reduced photon-count levels resulting in low signal-to-noise ratio (SNR), segmentation difficulties, and quantification uncertainties. PURPOSE: We developed and evaluated a novel deep-learning (DL) architecture-Attention based Residual-Dilated Net (ARD-Net)-to generate standard-count PET (SC-PET) images from LC-PET images. The performance of the ARD-Net framework was evaluated for numerous low count realizations using fidelity-based qualitative metrics, task-based segmentation, and quantitative metrics. METHOD: Patient Derived tumor Xenograft (PDX) with tumors implanted in the mammary fat-pad were subjected to preclinical [18F]-Fluorodeoxyglucose (FDG)-PET/CT imaging. SC-PET images were derived from a 10 min static FDG-PET acquisition, 50 min post administration of FDG, and were resampled to generate four distinct LC-PET realizations corresponding to 10%, 5%, 1.6%, and 0.8% of SC-PET count-level. ARD-Net was trained and optimized using 48 preclinical FDG-PET datasets, while 16 datasets were utilized to assess performance. Further, the performance of ARD-Net was benchmarked against two leading DL-based methods (Residual UNet, RU-Net; and Dilated Network, D-Net) and non-DL methods (Non-Local Means, NLM; and Block Matching 3D Filtering, BM3D). The performance of the framework was evaluated using traditional fidelity-based image quality metrics such as Structural Similarity Index Metric (SSIM) and Normalized Root Mean Square Error (NRMSE), as well as human observer-based tumor segmentation performance (Dice Score and volume bias) and quantitative analysis of Standardized Uptake Value (SUV) measurements. Additionally, radiomics-derived features were utilized as a measure of quality assurance (QA) in comparison to true SC-PET. Finally, a performance ensemble score (EPS) was developed by integrating fidelity-based and task-based metrics. Concordance Correlation Coefficient (CCC) was utilized to determine concordance between measures. The non-parametric Friedman Test with Bonferroni correction was used to compare the performance of ARD-Net against benchmarked methods with significance at adjusted p-value ≤0.01. RESULTS: ARD-Net-generated SC-PET images exhibited significantly better (p ≤ 0.01 post Bonferroni correction) overall image fidelity scores in terms of SSIM and NRMSE at majority of photon-count levels compared to benchmarked DL and non-DL methods. In terms of task-based quantitative accuracy evaluated by SUVMean and SUVPeak, ARD-Net exhibited less than 5% median absolute bias for SUVMean compared to true SC-PET and lower degree of variability compared to benchmarked DL and non-DL based methods in generating SC-PET. Additionally, ARD-Net-generated SC-PET images displayed higher degree of concordance to SC-PET images in terms of radiomics features compared to non-DL and other DL approaches. Finally, the ensemble score suggested that ARD-Net exhibited significantly superior performance compared to benchmarked algorithms (p ≤ 0.01 post Bonferroni correction). CONCLUSION: ARD-Net provides a robust framework to generate SC-PET from LC-PET images. ARD-Net generated SC-PET images exhibited superior performance compared other DL and non-DL approaches in terms of image-fidelity based metrics, task-based segmentation metrics, and minimal bias in terms of task-based quantification performance for preclinical PET imaging.

A Mobile Health Behavior Change Intervention for Women With Coronary Heart Disease: A RANDOMIZED CONTROLLED PILOT STUDY.

PURPOSE: The aim of this study was to evaluate the effects of a mobile health (mHealth) intervention, HerBeat, compared with educational usual care (E-UC) for improving exercise capacity (EC) and other patient-reported outcomes at 3 mo among women with coronary heart disease. METHODS: Women were randomized to the HerBeat group (n = 23), a behavior change mHealth intervention with a smartphone, smartwatch, and health coach or to the E-UC group (n = 24) who received a standardized cardiac rehabilitation workbook. The primary endpoint was EC measured with the 6-min walk test (6MWT). Secondary outcomes included cardiovascular disease risk factors and psychosocial well-being. RESULTS: A total of 47 women (age 61.2 ± 9.1 yr) underwent randomization. The HerBeat group significantly improved on the 6MWT from baseline to 3 mo ( P = .016, d = .558) while the E-UC group did not ( P = .894, d =-0.030). The between-group difference of 38 m at 3 mo was not statistically significant. From baseline to 3 mo, the HerBeat group improved in anxiety ( P = .021), eating habits confidence ( P = .028), self-efficacy for managing chronic disease ( P = .001), diastolic blood pressure ( P = .03), general health perceptions ( P = .047), perceived bodily pain ( P = .02), and waist circumference ( P = .008) while the E-UC group showed no improvement on any outcomes. CONCLUSIONS: The mHealth intervention led to improvements in EC and several secondary outcomes from baseline to 3 mo while the E-UC intervention did not. A larger study is required to detect small differences between groups. The implementation and outcomes evaluation of the HerBeat intervention was feasible and acceptable with minimal attrition.

Preclinical validation of an Escherichia coli O-antigen glycoconjugate for the prevention of serotype O1 invasive disease.

A US collection of invasive Escherichia coli serotype O1 bloodstream infection (BSI) isolates were assessed for genotypic and phenotypic diversity as the basis for designing a broadly protective O-antigen vaccine. Eighty percent of the BSI isolate serotype O1 strains were genotypically ST95 O1:K1:H7. The carbohydrate repeat unit structure of the O1a subtype was conserved in the three strains tested representing core genome multi-locus sequence types (MLST) sequence types ST95, ST38, and ST59. A long-chain O1a CRM197 lattice glycoconjugate antigen was generated using oxidized polysaccharide and reductive amination chemistry. Two ST95 strains were investigated for use in opsonophagocytic assays (OPA) with immune sera from vaccinated animals and in murine lethal challenge models. Both strains were susceptible to OPA killing with O1a glycoconjugate post-immune sera. One of these, a neonatal sepsis strain, was found to be highly lethal in the murine challenge model for which virulence was shown to be dependent on the presence of the K1 capsule. Mice immunized with the O1a glycoconjugate were protected from challenges with this strain or a second, genotypically related, and similarly virulent neonatal isolate. This long-chain O1a CRM197 lattice glycoconjugate shows promise as a component of a multi-valent vaccine to prevent invasive E. coli infections. IMPORTANCE: The Escherichia coli serotype O1 O-antigen serogroup is a common cause of invasive bloodstream infections (BSI) in populations at risk such as newborns and the elderly. Sequencing of US BSI isolates and structural analysis of O polysaccharide antigens purified from strains that are representative of genotypic sub-groups confirmed the relevance of the O1a subtype as a vaccine antigen. O polysaccharide was purified from a strain engineered to produce long-chain O1a O-antigen and was chemically conjugated to CRM197 carrier protein. The resulting glycoconjugate elicited functional antibodies and was protective in mice against lethal challenges with virulent K1-encapsulated O1a isolates.