Characterization and Sequence Mapping of Large RNA and mRNA Therapeutics Using Mass Spectrometry.

Large RNA including mRNA (mRNA) has emerged as an important new class of therapeutics. Recently, this has been demonstrated by two highly efficacious vaccines based on mRNA sequences encoding for a modified version of the SARS-CoV-2 spike protein. There is currently significant demand for the development of new and improved analytical methods for the characterization of large RNA including mRNA therapeutics. In this study, we have developed an automated, high-throughput workflow for the rapid characterization and direct sequence mapping of large RNA and mRNA therapeutics. Partial RNase digestions using RNase T1 immobilized on magnetic particles were performed in conjunction with high-resolution liquid chromatography-mass spectrometry analysis. Sequence mapping was performed using automated oligoribonucleotide annotation and identifications based on MS/MS spectra. Using this approach, a >80% sequence of coverage of a range of large RNAs and mRNA therapeutics including the SARS-CoV-2 spike protein was obtained in a single analysis. The analytical workflow, including automated sample preparation, can be completed within 90 min. The ability to rapidly identify, characterize, and sequence map large mRNA therapeutics with high sequence coverage provides important information for identity testing, sequence validation, and impurity analysis.

Plain language summary of the HERO study comparing relugolix with leuprolide for men with advanced prostate cancer.

WHAT IS THIS SUMMARY ABOUT?: This is a summary of a research study (known as a clinical trial) called HERO. The HERO study compared how well relugolix and leuprolide worked in lowering blood testosterone to sustained castration levels in men with advanced prostate cancer. Sustained castration is a blood testosterone level below 50 ng/dl from Day 29 through 48 weeks of treatment. WHAT WERE THE RESULTS?: Researchers looked at 930 adult men with advanced prostate cancer: 622 of these men took relugolix (by mouth once daily) and 308 received leuprolide (injected every 3 months). The HERO study showed that more men taking relugolix (97%) achieved sustained castration through 48 weeks than men receiving leuprolide (89%). This decrease in testosterone also happened more quickly in men taking relugolix. In 184 men who were followed up for 90 days after completing treatment, blood levels of testosterone returned to normal in more men who took relugolix than men who received leuprolide. Side effects were similar among men taking relugolix or receiving leuprolide, and most were identified as mild or moderate in terms of how bad they were. WHAT DO THE RESULTS OF THE STUDY MEAN?: In men with advanced prostate cancer and compared with those receiving leuprolide, more men taking relugolix had lower levels of blood testosterone. ClinicalTrials.gov NCT number: NCT03085095.

Assessing frailty indicators in the context of psychiatric disorder: A Delphi consensus study.

OBJECTIVES: Substantial construct overlap exists between indicators of frailty and symptoms of some psychiatric disorders. This study aimed to gain consensus of expert academic opinion on the potential impact of psychiatric illness on frailty assessment and how best to conceptualise and measure frailty indicators in the context of psychiatric symptoms. DESIGN: A classic Delphi approach was employed across two studies to achieve consensus: The first-round questionnaire consisted of open-ended questions, analysed through content analysis. The results informed the development of statements for participants to rate their agreement with in subsequent Delphi rounds. Statements with ≥66% agreement were accepted. Delphi Study 1 recruited experts in frailty assessment (n = 13) and Delphi Study 2 recruited experts in frailty and psychiatric disorder (n = 8). Experts were recruited globally. RESULTS: Overall, 40% of Delphi Study 1 statements and 43% of Delphi Study 2 statements were accepted. Primarily, consensus was reached for statements concerning the influence of depression/anxiety on frailty assessment and potential methods of conceptualising and measuring frailty indicators in the context of psychiatric symptoms. Little consensus was reached concerning the ease and importance of differentiating between frailty indicators and psychiatric assessment criteria with substantial overlap. CONCLUSIONS: The Delphi studies provide a novel exploration and consensus of expert academic opinions concerning the assessment of frailty indicators in the context of psychiatric symptoms. The results will inform future research into the adaptation or development of a frailty assessment tool specifically for use in older adult psychiatric populations.

Histological, electrophysiological and clinical effects of thermal radiofrequency therapy of the saphenous nerve and pulsed radiofrequency therapy of the sciatic nerve in dogs.

OBJECTIVE: Thermal radiofrequency (TRF) of the saphenous nerve (a sensory nerve) combined with pulsed radiofrequency (PRF) of the sciatic nerve (a sensory and motor nerve) might relieve intractable stifle osteoarthritis (OA) pain in dogs. The objective was to determine if saphenous nerve TRF induces Wallerian degeneration and if sciatic nerve PRF induces degeneration or dysfunction. STUDY DESIGN: Blinded, controlled, randomized, preclinical study. ANIMALS: A group of six intact, female Beagle dogs aged 14-16 months. METHODS: In each dog, one pelvic limb was assigned randomly to the control group and the other to the treatment group. Dogs were anesthetized and, using ultrasonography, radiofrequency electrodes were positioned adjacent to saphenous and sciatic nerves bilaterally; TRF and PRF were performed only in the treatment limb. Motor nerve conduction velocity (MNCV) was measured in both sciatic nerves 2 weeks later, and the dogs were euthanized. Hematoxylin and eosin-stained sections of saphenous and sciatic nerves were examined using light microscopy. Degeneration and inflammation were scored 0 (none) to 3 (severe). A one-tailed, paired Wilcoxon signed-rank test was used to test for differences in scores and MNCV between control and treatment nerves. RESULTS: Degeneration and inflammation scores were higher in treatment saphenous nerves in 5/6 dogs [83%; 95% confidence interval (CI), 36%, 99%]; however, after Bonferroni correction only degeneration score was higher (p = 0.0313). Degeneration, inflammation or decreased MNCV were not observed in sciatic nerves (each outcome: 0/6 nerves, 0%; 95% CI, 0%, 48%). No dogs experienced postprocedural pain or neurological deficits. CONCLUSIONS AND CLINICAL RELEVANCE: The degeneration in TRF-treated saphenous nerves appears sufficient to impair transmission. Sciatic nerve PRF did not cause degeneration with attendant motor deficits, consistent with a proposed neuromodulatory mechanism. A clinical trial is needed to confirm the combined techniques produce analgesia without motor deficits in dogs with stifle OA.

Size and characteristics of the biomedical research workforce associated with U.S. National Institutes of Health extramural grants.

The U.S. National Institutes of Health (NIH) annually invests approximately $22 billion in biomedical research through its extramural grant programs. Since fiscal year (FY) 2010, all persons involved in research during the previous project year have been required to be listed on the annual grant progress report. These new data have enabled the production of the first-ever census of the NIH-funded extramural research workforce. Data were extracted from All Personnel Reports submitted for NIH grants funded in FY 2009, including position title, months of effort, academic degrees obtained, and personal identifiers. Data were de-duplicated to determine a unique person count. Person-years of effort (PYE) on NIH grants were computed. In FY 2009, NIH funded 50,885 grant projects, which created 313,049 full- and part-time positions spanning all job functions involved in biomedical research. These positions were staffed by 247,457 people at 2,604 institutions. These persons devoted 121,465 PYE to NIH grant-supported research. Research project grants each supported 6 full- or part-time positions, on average. Over 20% of positions were occupied by postdoctoral researchers and graduate and undergraduate students. These baseline data were used to project workforce estimates for FYs 2010-2014 and will serve as a foundation for future research.

Psychometric properties of multicomponent tools designed to assess frailty in older adults: A systematic review.

BACKGROUND: Frailty is widely recognised as a distinct multifactorial clinical syndrome that implies vulnerability. The links between frailty and adverse outcomes such as death and institutionalisation have been widely evidenced. There is currently no gold standard frailty assessment tool; optimizing the assessment of frailty in older people therefore remains a research priority. The objective of this systematic review is to identify existing multi-component frailty assessment tools that were specifically developed to assess frailty in adults aged ≥60 years old and to systematically and critically evaluate the reliability and validity of these tools. METHODS: A systematic literature review was conducted using the standardised COnsensus-based Standards for the selection of health Measurement INstruments (COSMIN) checklist to assess the methodological quality of included studies. RESULTS: Five thousand sixty-three studies were identified in total: 73 of which were included for review. 38 multi-component frailty assessment tools were identified: Reliability and validity data were available for 21 % (8/38) of tools. Only 5 % (2/38) of the frailty assessment tools had evidence of reliability and validity that was within statistically significant parameters and of fair-excellent methodological quality (the Frailty Index-Comprehensive Geriatric Assessment [FI-CGA] and the Tilburg Frailty Indicator [TFI]). CONCLUSIONS: The TFI has the most robust evidence of reliability and validity and has been the most extensively examined in terms of psychometric properties. However, there is insufficient evidence at present to determine the best tool for use in research and clinical practice. Further in-depth evaluation of the psychometric properties of these tools is required before they can fulfil the criteria for a gold standard assessment tool.

Practical Guidance on [177Lu]Lu-PSMA-617 Treatment, Including Radiation Safety, Adverse Event Monitoring, and Patient Counseling.

BACKGROUND: The approval of the prostate-specific membrane antigen (PSMA)-targeted radio-ligand therapy [177Lu]Lu-PSMA-617 represents a shift toward precision medicine-based treatments for metastatic castration-resistant prostate cancer. OBJECTIVES: This review aims to provide practical advice and clinical considerations for working with [177Lu]Lu-PSMA-617; particularly, regarding the role of nurses in managing radiation safety, monitoring and reporting of adverse events, and counseling patients receiving this therapy. METHODS: Clinical data, protocols, and guidelines are summarized alongside real-world insights to provide practical recommendations for nurses caring for patients treated with [177Lu]Lu-PSMA-617. FINDINGS: Nurses coordinate safe care for patients receiving [177Lu]Lu-PSMA-617 by facilitating communication among physicians, managing logistic concerns, monitoring for adverse events related to treatment, providing education, and counseling patients and caregivers throughout treatment.

Exploration patterns shape cognitive map learning.

Spontaneous, volitional spatial exploration is crucial for building up a cognitive map of the environment. However, decades of research have primarily measured the fidelity of cognitive maps after discrete, controlled learning episodes. We know little about how cognitive maps are formed during naturalistic free exploration. Here, we investigated whether exploration trajectories predicted cognitive map accuracy, and how these patterns were shaped by environmental structure. In two experiments, participants freely explored a previously unfamiliar virtual environment. We related their exploration trajectories to a measure of how long they spent in areas with high global environmental connectivity (integration, as assessed by space syntax). In both experiments, we found that participants who spent more time on paths that offered opportunities for integration formed more accurate cognitive maps. Interestingly, we found no support for our pre-registered hypothesis that self-reported trait differences in navigation ability would mediate this relationship. Our findings suggest that exploration patterns predict cognitive map accuracy, even for people who self-report low ability, and highlight the importance of considering both environmental structure and individual variability in formal theory- and model-building.

Oligoether-strapped calix[4]pyrrole: an ion-pair receptor displaying cation-dependent chloride anion transport.

A ditopic ion-pair receptor (1), which has tunable cation- and anion-binding sites, has been synthesized and characterized. Spectroscopic analyses provide support for the conclusion that receptor 1 binds fluoride and chloride anions strongly and forms stable 1:1 complexes ([1·F](-) and [1·Cl](-)) with appropriately chosen salts of these anions in acetonitrile. When the anion complexes of 1 were treated with alkali metal ions (Li(+), Na(+), K(+), Cs(+), as their perchlorate salts), ion-dependent interactions were observed that were found to depend on both the choice of added cation and the initially complexed anion. In the case of [1·F](-), no appreciable interaction with the K(+) ion was seen. On the other hand, when this complex was treated with Li(+) or Na(+) ions, decomplexation of the bound fluoride anion was observed. In contrast to what was seen with Li(+), Na(+), K(+), treating [1·F](-) with Cs(+) ions gave rise to a stable, host-separated ion-pair complex, [F·1·Cs], which contains the Cs(+) ion bound in the cup-like portion of the calix[4]pyrrole. Different complexation behavior was seen in the case of the chloride complex, [1·Cl](-). Here, no appreciable interaction was observed with Na(+) or K(+). In contrast, treating with Li(+) produces a tight ion-pair complex, [1·Li·Cl], in which the cation is bound to the crown moiety. In analogy to what was seen for [1·F](-), treatment of [1·Cl](-) with Cs(+) ions gives rise to a host-separated ion-pair complex, [Cl·1·Cs], in which the cation is bound to the cup of the calix[4]pyrrole. As inferred from liposomal model membrane transport studies, system 1 can act as an effective carrier for several chloride anion salts of Group 1 cations, operating through both symport (chloride+cation co-transport) and antiport (nitrate-for-chloride exchange) mechanisms. This transport behavior stands in contrast to what is seen for simple octamethylcalix[4]pyrrole, which acts as an effective carrier for cesium chloride but does not operates through a nitrate-for-chloride anion exchange mechanism.

Individual differences in the allocation of visual attention during navigation.

Individual differences exist in the ability to create an accurate mental survey representation (i.e., a cognitive map) of a novel environment, yet the mechanisms underlying differences in cognitive map accuracy are still under investigation. To determine whether differences in overt attention allocation contribute to these individual differences, the current study examined whether looking times to landmarks and other objects while navigating in a dynamic virtual environment were related to cognitive map accuracy. Participants completed a battery of spatial tests; some tests assessed spatial skills prior to the navigation task (the Santa Barbara Sense of Direction Scale and the Spatial Orientation Test), and others tested memory of the virtual environment Silcton after an exploration period (a landmark recognition task, a direction estimation task, a map-building task, and a route construction task). Individuals with inaccurate cognitive maps of Silcton, as measured by the direction estimation and map-building tasks, showed equivalent eye fixations to buildings and objects when exploring Silcton as those with accurate maps. Despite similar looking times, the inaccurate mappers were significantly worse at judgments of relative direction between landmarks in Silcton and showed poorer memory for landmarks in Silcton than accurate mappers. These findings suggest that cognitive mechanisms, such as mental perspective-taking, occurring after attention allocation underlie differences in cognitive map accuracy. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Mass spectrometric discovery and selective reaction monitoring (SRM) of putative protein biomarker candidates in first trimester Trisomy 21 maternal serum.

The accurate diagnosis of Trisomy 21 requires invasive procedures that carry a risk of miscarriage. The current state-of-the-art maternal serum screening tests measure levels of PAPP-A, free bhCG, AFP, and uE3 in various combinations with a maximum sensitivity of 60-75% and a false positive rate of 5%. There is currently an unmet need for noninvasive screening tests with high selectivity that can detect pregnancies at risk, preferably within the first trimester. The aim of this study was to apply proteomics and mass spectrometry techniques for the discovery of new putative biomarkers for Trisomy 21 in first trimester maternal serum coupled with the immediate development of quantitative selective reaction monitoring (SRM) assays. The results of the novel workflow were 2-fold: (1) we identified a list of differentially expressed proteins in Trisomy 21 vs Normal samples, including PAPP-A, and (2) we developed a multiplexed, high-throughput SRM assay for verification of 12 new putative markers identified in the discovery experiments. To narrow down the initial large list of differentially expressed candidates resulting from the discovery experiments, we incorporated receiver operating characteristic (ROC) curve algorithms early in the data analysis process. We believe this approach provides a substantial advantage in sifting through the large and complex data typically obtained from discovery experiments. The workflow efficiently mined information derived from high-resolution LC-MS/MS discovery data for the seamless construction of rapid, targeted assays that were performed on unfractionated serum digests. The SRM assay lower limit of detection (LLOD) for the target peptides in a background of digested serum matrix was approximately 250-500 attomoles on column and the limit of accurate quantitation (LOQ) was approximately 1-5 femtomoles on column. The assay error as determined by coefficient of variation at LOQ and above ranged from 0 to 16%. The workflow developed in this study bridges the gap between proteomic biomarker discovery and translation into a clinical research environment. Specifically, for Trisomy 21, the described multiplexed SRM assay provides a vehicle for high-throughput verification of these, and potentially other, peptide candidates on larger sample cohorts.

Optimisation of the use of sliding window deconvolution for comprehensive characterisation of trastuzumab and adalimumab charge variants by native high resolution mass spectrometry.

The biopharmaceutical industry continues to develop mAb-based biotherapeutics in increasing numbers. Due to their complexity, there are several critical quality attributes (CQAs) that need to be measured and controlled to guarantee product safety and efficacy. Charge variant analysis is a widely used method to monitor changes in product quality during the manufacturing process of monoclonal antibodies (mAbs) and, together with a bottom-up peptide centred approach, acts as a key analytical platform to fulfil regulatory requirements. Native MS measures biomolecules under conditions that preserve most aspects of protein tertiary and quaternary structure, enabling direct characterization of large intact proteins such as mAbs. The resulting native mass spectrum of a mAb is characterized by a narrower charge-state envelope that simplifies the spectra and also condenses the ion signals into fewer peaks, increasing the signal-to-noise ratio. Algorithmic spectral deconvolution is needed for routine accurate and rapid molecular weight determination, and consequently, multiple deconvolution algorithms have evolved over the past decade. Here, we demonstrate the utility of the sliding window algorithm as a robust and powerful deconvolution tool for comprehensive characterisation of charge variant analysis data for mAbs. Optimum performance is evaluated by studying the impact of critical software parameters on detection, identification and relative quantitation of protein isoforms. By combining molecular mass and retention time information, it was possible to identify multiple modifications on adalimumab and trastuzumab, both IgG1 mAbs, including lysine truncation, deamidation and succinimide formation, along with the N-glycan distribution of each of the identified charge variants. Sliding window deconvolution also provides a key benefit of low abundant variant detection in a single analysis and the ability to detect co-eluting components with different relative abundances. The studied mAbs demonstrate the algoritms applicability for efficient data processing of both simple and complex mAbs analysed using pH gradient cation exchange chromatography coupled to native mass spectrometry.

Competition between two MHC binding registers in a single peptide processed from myelin basic protein influences tolerance and susceptibility to autoimmunity.

Experimental allergic encephalomyelitis (EAE) is an animal model for multiple sclerosis induced by stimulating myelin basic protein (MBP)-specific T cells. The MBP-specific repertoire in B10.PL mice is shaped by tolerance mechanisms that eliminate MBP121-150-specific T cells. In contrast, MBPAc1-11-specific T cells escape tolerance and constitute the encephalitogenic repertoire. To determine if this differential tolerance is caused by differences in the abundance of MBP epitopes generated by processing, MBP peptides were eluted from I-Au complexes and analyzed by mass spectrometry. Peptides were identified from both the NH2-terminal and MBP121-150 regions. Unexpectedly, MBPAc1-18 and Ac1-17, which contain the MBPAc1-11 epitope, were much more abundant than MBP121-150 peptides. The results demonstrate that competition between two I-Au binding registers, a low affinity register defined by MBPAc1-11 and a high affinity register defined by MBP5-16, prevents most of the NH2-terminal naturally processed peptides from binding in the MBPAc1-11 register. The small fraction of MBPAc1-18 bound in the MBPAc1-11 register is not sufficient to induce tolerance but provides a ligand for MBPAc1-11-specific T cells during disease. These results provide a basis for both the lack of tolerance to MBPAc1-11 and the ability of this epitope to become a target during autoimmunity.

Dynamic host energetics and cytoskeletal proteomes in human immunodeficiency virus type 1-infected human primary CD4 cells: analysis by multiplexed label-free mass spectrometry.

We report on a proteomic analysis of ex vivo human immunodeficiency virus (HIV) type 1 infection in human primary CD4 cells by shotgun liquid chromatography-tandem mass spectrometry analysis, revealing two distinct proteomic profiles at two phases of virus replication. Relative to mock-infected cells, 168 signature proteins exhibited abundance changes at the first sign of Gag p24 production (8 h postinfection [p.i.]) or the peak of virus replication (24 h p.i.); interestingly, most of the changes were exclusive to only one phase of virus replication. Based on characterization by functional ontology and known human-HIV protein interactions, we observed the enrichment for protein abundance increases pertaining to protein synthesis and nucleasomal reorganization amid an otherwise placid cellular proteome at the first sign of HIV replication. In contrast, we observed indications of decreased protein turnover, concomitant with heightened DNA repair activities and preludes to apoptosis, in the presence of robust virus replication. We also observed hints of disruptions in protein and small molecule trafficking. Our label-free proteomic strategy allowed us to perform multiplexed comparisons-we buttressed our detection specificity with the use of a reverse transcriptase inhibitor as a counterscreen, enabling highlighting of cellular protein abundance changes unique to robust virus replication as opposed to viral entry. In conjunction with complementary high-throughput screens for cellular partners of HIV, we put forth a model pinpointing specific rerouting of cellular biosynthetic, energetic, and trafficking pathways as HIV replication accelerates in human primary CD4 cells.

Outcomes of a randomized controlled trial assessing a smartphone Application to reduce unmet needs among people diagnosed with CancEr (ACE).

BACKGROUND: Smartphone technology represents an opportunity to deliver practical solutions for people affected by cancer at a scale that was previously unimaginable, such as information, appointment monitoring, and improved access to cancer support services. This study aimed to determine whether a smartphone application (app) reduced the unmet needs among people newly diagnosed with cancer. METHODS: A single blind, multisite randomized controlled trial to determine the impact of an app-based, 4-month intervention. Newly diagnosed cancer patients were approached at three health service treatment clinics. RESULTS: Eighty-two people were randomized (intervention; n = 43 and control; n = 39), average age was 59.5 years (SD: 12.9); 71% female; 67% married or in a de facto relationship. At baseline, there were no differences in participants' characteristics between the groups. No significant effects, in reducing unmet needs, were demonstrated at the end of intervention (4-month) or 12-month follow-up. Overall, 94% used the app in weeks 1-4, which decreased to 41% in weeks 13-16. Mean app use time per participant: Cancer Information, 6.9 (SD: 18.9) minutes; Appointment Schedule, 5.1 (SD: 9.6) minutes; Cancer Services 1.5 minutes (SD: 6.8); Hospital Navigation, 1.4 (SD: 2.8) minutes. CONCLUSIONS: Despite consumer involvement in the design of this smartphone technology, the app did not reduce unmet needs. This may have been due to the study being underpowered. To contribute to a meaningful understanding and improved implementation of smartphone technology to support people affected by cancer, practical considerations, such as recruitment issues and access to, and confidence with, apps, need to be considered. Australian New Zealand Clinical Trials Registration (ACTRN) Trial Registration: 12616001251415; WEF 7/9/2016.

Individual differences in cognitive map accuracy: Investigating the role of landmark familiarity.

Broad individual differences exist in the ability to create a cognitive map of a new environment. The current studies investigated whether familiarizing participants with to-be-learned target landmarks (Experiment 1) or target landmarks plus the order they would be encountered along routes (Experiment 2) before exploring the Silcton virtual environment would increase performance on tasks assaying spatial memory of Silcton. Participants in both experiments were randomly assigned to be pre-exposed either to information about target landmarks in Silcton or control landmarks on the university campus. In both experiments, participants explored Silcton via four prescribed routes and then performed a direction estimation task and a map building task based on memory for the locations of the target landmarks. In addition, participants completed the Spatial Orientation Test of perspective-taking. Pre-exposure to Silcton landmarks versus control landmarks did not affect scores on Silcton-based tasks in either experiment. Some sex differences in direction estimation were observed in Experiment 1 but not Experiment 2. While facilitating familiarity with landmarks did not improve cognitive map accuracy, both sex and perspective taking ability were found to contribute to individual differences in the ability to create a cognitive map. (PsycINFO Database Record (c) 2019 APA, all rights reserved).

Multicomponent Frailty Assessment Tools for Older People with Psychiatric Disorders: A Systematic Review.

OBJECTIVE: To review evidence evaluating the use of multicomponent frailty assessment tools in assessing frailty in older adults with psychiatric disorders. METHODS: A systematic literature review was conducted to identify all multicomponent frailty assessment tools (ie, a tool that assesses two or more indicators of frailty). The items of each frailty assessment tool were compared with Diagnostic and Statistical Manual of Mental Disorders, 5th edition (DSM-5) diagnostic criteria for psychiatric disorders to assess construct overlap. Studies conducted in community, inpatient, and outpatient clinical settings were considered for inclusion. PARTICIPANTS: Adults aged 60 years or older. RESULTS: A total of 5639 records were identified following the removal of duplicates, from which 95 studies were included for review. Of the 48 multicomponent frailty assessment tools identified, no tool had been developed for, or validated in, older adult populations with a psychiatric disorder. Overall, 20 of 48 frailty assessment tools contained a psychological assessment domain, with 17 of 48 tools citing the presence of depressed mood and/or anxiety as a frailty indicator. Common areas of construct overlap in frailty assessment tools and DSM-5 diagnostic criteria included weight loss (29 of 48) and fatigue (21 of 48). CONCLUSIONS: Significant construct overlap exists between the indicators of frailty as conceptualized in existing frailty assessment tools and DSM-5 diagnostic criteria for common psychiatric disorders including major depressive episode and generalized anxiety disorder that has the potential to confound frailty assessment results. Further research is necessary to establish a reliable and valid tool to assess frailty in this population. J Am Geriatr Soc 67:1085-1095, 2019.

Memory without awareness: pigeons do not show metamemory in delayed matching to sample.

Metamemory, the ability to report on memory strength, is clearly established in rhesus macaques (Macaca mulatta) by converging evidence from several paradigms. In contrast, A. Inman and S. J. Shettleworth (1999) found no conclusive evidence of metamemory in pigeons. The authors studied pigeons further in 3 paradigms, with multiple tests of metamemory in each. Pigeons encountered a safe alternative to a matching-to-sample test either before (Experiment 2) or concurrently with (Experiment 3) the matching test. Choices of the safe option did not vary consistently with matching accuracy or change in trials with omitted samples in the way predicted for an animal with metamemory. In Experiment 4, confidence ratings following completion of the matching test also did not vary consistently as predicted by metamemory.

Charge variant native mass spectrometry benefits mass precision and dynamic range of monoclonal antibody intact mass analysis.

The preponderance and diversity of charge variants in therapeutic monoclonal antibodies has implications for antibody efficacy and degradation. Understanding the extent and impact of minor antibody variants is of great interest, and it is also a critical regulatory requirement. Traditionally, a combination of approaches is used to characterize antibody charge heterogeneity, including ion exchange chromatography and independent mass spectrometric variant site mapping after proteolytic digestion. Here, we describe charge variant native mass spectrometry (CVMS), an integrated native ion exchange mass spectrometry-based charge variant analytical approach that delivers detailed molecular information in a single, semi-automated analysis. We utilized pure volatile salt mobile phases over a pH gradient that effectively separated variants based on minimal differences in isoelectric point. Characterization of variants such as deamidation, which are traditionally unattainable by intact mass due to their minimal molecular weight differences, were measured unambiguously by mass and retention time to allow confident MS1 identification. We demonstrate that efficient chromatographic separation allows introduction of the purified forms of the charge variant isoforms into the Orbitrap mass spectrometer. Our CVMS method allows confident assignment of intact monoclonal antibody isoforms of similar mass and relative abundance measurements across three orders of magnitude dynamic range.