How are early-career dementia researchers considered and supported on a national level by dementia plans and organizations? An overview of global policy approaches.

INTRODUCTION: Despite representing an essential workforce, it is unclear how global policy efforts target early-career dementia researchers (ECDRs). Thus, this study aimed to provide an overview of policies through which ECDRs are considered and supported by dementia plans and organizations. METHODS: G20 member states were evaluated for their national dementia plan alongside policies of leading dementia organizations. Data targeting support for ECDRs were extracted and subject to content analysis using inductive coding. Findings were categorized and narratively synthesized. RESULTS: Only China, Denmark, England, Greece, Northern Ireland, Scotland, Spain, and the United States mentioned ECDRs in their national plan. Additionally, 17 countries formalized ECDR support via dementia organizations. Support efforts included research funding, dissemination and networking, career development, and research advice. DISCUSSION: Few nations formally recognized ECDRs in dementia plans or through dementia organizations. To facilitate equal prospects for ECDRs, top-down approaches are urged to enhance and align their efforts. HIGHLIGHTS: Few G20 countries (8/46) had national dementia plans for early-career researchers. Targeted support comes from government and nongovernmental dementia organizations. Support includes funding, training, advice, research dissemination, and networking. Inconsistent definitions and eligibility criteria are barriers to accessing support. Global coordination and top-down policy will aid early-career dementia researchers.

Improved determination of Neisseria gonorrhoeae gyrase A genotype results in clinical specimens.

BACKGROUND: The emergence of drug-resistant Neisseria gonorrhoeae has prompted the development of rapid molecular assays designed to determine antimicrobial susceptibility. One common assay uses high-resolution melt analysis to target codon 91 of the gyrase A gene (gyrA) to predict N. gonorrhoeae susceptibility to ciprofloxacin. METHODS: We extracted DNA from remnant clinical specimens that had previously tested positive for N. gonorrhoeae using the Aptima Combo 2 for CT/NG assay (Hologic, San Diego, CA, USA). We selected DNA extracts from specimens with indeterminate, WT and mutant gyrA genotype results from a previous study using high-resolution melt analysis to detect the gyrA codon 91 mutation. We re-tested those specimens using the recently CE-marked ResistancePlus GC (beta) assay (SpeeDx, Sydney, Australia). RESULTS: Of 86 specimens with indeterminate gyrA genotypes on high-resolution melt analysis, the ResistancePlus GC (beta) assay (SpeeDx) identified 30 (35%) WT, 22 (26%) mutant and 34 (40%) indeterminate gyrA genotypes. CONCLUSIONS: The ResistancePlus GC (beta) assay showed improved N. gonorrhoeae gyrA genotype determination compared with a prior gyrA genotypic high-resolution melt assay.

Effect of a Same-Day-Appointment Initiative on Access-Related Benchmarks in Radiation Oncology.

PURPOSE: The timely delivery of health care is an important quality indicator that has been shown to correlate with outcomes for cancer patients. We present our single-institution experience with the implementation of a same-day-access scheduling initiative in outpatient radiation oncology. METHODS AND MATERIALS: From March 2021 to March 2023, a total of 4301 consecutive new patients referred for radiation oncology consultation were offered same-day appointments as part of a prospective pilot initiative conducted at a tertiary-based academic medical center. Descriptive statistics were used to study the effect of this initiative on access-related benchmarks compared with historical control patients referred during a preceding 24-month period. RESULTS: Among the 3414 patients scheduled, 477 (14%) opted for same-day appointments. Black, Latino, and Asian patients were significantly more likely to use same-day access versus Caucasian patients (P < .001). The same-day-access initiative increased the proportion of patients seen within 5 days from referral from 22% to 61% (P < .001). The median time from referral to consult was 12 days (range, 0-149 days) before the implementation of the same-day-access initiative compared with 3 days (range, 0-101 days) after (P < .001). The no-show rate was reduced from 15% to 7% with the initiative (P < .001). All patients who requested a same-day appointment were successfully accommodated. CONCLUSIONS: The implementation of this same-day-access initiative enhanced operational efficiency and reduced barriers to care in the outpatient setting.

Timing of chemotherapy and radiation delivery for patients receiving chemoradiation for head and neck cancer: When does "concurrent" mean concurrent?

PURPOSE: To analyze practice patterns focusing on variations in the timing of chemotherapy relative to radiation in patients treated with concurrent chemoradiation for head and neck cancer. METHODS AND MATERIALS: The medical records of 302 consecutive adult patients treated with concurrent chemoradiation for head and neck cancer between April 2014 and February 2022 were reviewed. After excluding 38 patients who received non-platinum-based regimens, induction chemotherapy, and/or had non-squamous cell histology, a total of 264 patients formed the primary population. To study the variability in which concurrent chemoradiation was delivered, descriptive statistics were used to determine the percentage of patients who deviated from starting chemotherapy and radiation on the same day. The chi-square statistic was used to compare differences in proportion among various subsets. A Cox proportional hazards model was then used to perform a multi-variate analysis to identify factors which independently influenced the likelihood for non-adeherence. RESULTS: Among the 264 patients, a total of 187 patients (70.8%) had chemotherapy and radiation started on the same day with 171 of these (91.4%) receiving chemotherapy prior to radiation delivery. On multivariate analysis, both non-Caucasian ethnicity (OR: 1.13, 95% C.I. 1.01-1.20) and being non-English speaking (OR: 1.39; 95% C.I. 1.18--1.51) was significantly associated with greater likelihood of the receipt of radiation and chemotherapy on different days. CONCLUSION: Significant variation exists in the timing of chemotherapy relative to radiation for concurrent chemoradiation in the clinical setting. The potential repercussions on outcome warrante further invesigtation and are discussed.

Posttreatment Surveillance Imaging After Radiation for Head and Neck Cancer.

Importance: The role of surveillance imaging after treatment for head and neck cancer is controversial and evidence to support decision-making is limited. Objective: To determine the use of surveillance imaging in asymptomatic patients with head and neck cancer in remission after completion of chemoradiation. Design, Setting, and Participants: This was a retrospective, comparative effectiveness research review of adult patients who had achieved a complete metabolic response to initial treatment for head and neck cancer as defined by having an unequivocally negative positron emission tomography (PET) scan using the PET response criteria in solid tumors (PERCIST) scale within the first 6 months of completing therapy. The medical records of 501 consecutive patients who completed definitive radiation therapy (with or without chemotherapy) for newly diagnosed squamous cell carcinoma of the head and neck between January 2014 and June 2022 were reviewed. Exposure: Surveillance imaging was defined as the acquisition of a PET with computed tomography (CT), magnetic resonance imaging (MRI), or CT of the head and neck region in the absence of any clinically suspicious symptoms and/or examination findings. For remaining patients, subsequent surveillance after the achievement of a complete metabolic response to initial therapy was performed on an observational basis in the setting of routine follow-up using history-taking and physical examination, including endoscopy. This expectant approach led to imaging only in the presence of clinically suspicious symptoms and/or physical examination findings. Main Outcome and Measures: Local-regional control, overall survival, and progression-free survival based on assignment to either the surveillance imaging or expectant management cohort. Results: This study included 340 patients (mean [SD] age, 59 [10] years; 201 males [59%]; 88 Latino patients [26%]; 145 White patients [43%]) who achieved a complete metabolic response during this period. There was no difference in 3-year local-regional control, overall survival, progression-free survival, or freedom from distant metastasis between patients treated with surveillance imaging vs those treated expectantly. Conclusions and Relevance: In this comparative effectiveness research, imaging-based surveillance failed to improve outcomes compared with expectant management for patients who were seemingly in remission after completion of primary radiation therapy for head and neck cancer.

Functional analysis of a common BAG3 allele associated with protection from heart failure.

Multiple genetic association studies have correlated a common allelic block linked to the BAG3 gene with a decreased incidence of heart failure, but the molecular mechanism remains elusive. In this study, we used induced pluripotent stem cells to test if the only coding variant in this allele block, BAG3C151R, alters protein and cellular function in human cardiomyocytes. Quantitative protein interaction analysis identified changes in BAG3C151R protein partners specific to cardiomyocytes. Knockdown of genes encoding for BAG3-interacting factors in cardiomyocytes followed by myofibrillar analysis revealed that BAG3C151R associates more strongly with proteins involved in the maintenance of myofibrillar integrity. Finally, we demonstrate that cardiomyocytes expressing the BAG3C151R variant have improved response to proteotoxic stress in a dose-dependent manner. This study suggests that BAG3C151R could be responsible for the cardioprotective effect of the haplotype block, by increasing cardiomyocyte protection from stress. Preferential binding partners of BAG3C151R may reveal potential targets for cardioprotective therapies.

Gain-of-function cardiomyopathic mutations in RBM20 rewire splicing regulation and re-distribute ribonucleoprotein granules within processing bodies.

Mutations in the cardiac splicing factor RBM20 lead to malignant dilated cardiomyopathy (DCM). To understand the mechanism of RBM20-associated DCM, we engineered isogenic iPSCs with DCM-associated missense mutations in RBM20 as well as RBM20 knockout (KO) iPSCs. iPSC-derived engineered heart tissues made from these cell lines recapitulate contractile dysfunction of RBM20-associated DCM and reveal greater dysfunction with missense mutations than KO. Analysis of RBM20 RNA binding by eCLIP reveals a gain-of-function preference of mutant RBM20 for 3' UTR sequences that are shared with amyotrophic lateral sclerosis (ALS) and processing-body associated RNA binding proteins (FUS, DDX6). Deep RNA sequencing reveals that the RBM20 R636S mutant has unique gene, splicing, polyadenylation and circular RNA defects that differ from RBM20 KO. Super-resolution microscopy verifies that mutant RBM20 maintains very limited nuclear localization potential; rather, the mutant protein associates with cytoplasmic processing bodies (DDX6) under basal conditions, and with stress granules (G3BP1) following acute stress. Taken together, our results highlight a pathogenic mechanism in cardiac disease through splicing-dependent and -independent pathways.

A BAG3 chaperone complex maintains cardiomyocyte function during proteotoxic stress.

Molecular chaperones regulate quality control in the human proteome, pathways that have been implicated in many diseases, including heart failure. Mutations in the BAG3 gene, which encodes a co-chaperone protein, have been associated with heart failure due to both inherited and sporadic dilated cardiomyopathy. Familial BAG3 mutations are autosomal dominant and frequently cause truncation of the coding sequence, suggesting a heterozygous loss-of-function mechanism. However, heterozygous knockout of the murine BAG3 gene did not cause a detectable phenotype. To model BAG3 cardiomyopathy in a human system, we generated an isogenic series of human induced pluripotent stem cells (iPSCs) with loss-of-function mutations in BAG3. Heterozygous BAG3 mutations reduced protein expression, disrupted myofibril structure, and compromised contractile function in iPSC-derived cardiomyocytes (iPS-CMs). BAG3-deficient iPS-CMs were particularly sensitive to further myofibril disruption and contractile dysfunction upon exposure to proteasome inhibitors known to cause cardiotoxicity. We performed affinity tagging of the endogenous BAG3 protein and mass spectrometry proteomics to further define the cardioprotective chaperone complex that BAG3 coordinates in the human heart. Our results establish a model for evaluating protein quality control pathways in human cardiomyocytes and their potential as therapeutic targets and susceptibility factors for cardiac drug toxicity.

Miniaturized iPS-Cell-Derived Cardiac Muscles for Physiologically Relevant Drug Response Analyses.

Tissue engineering approaches have the potential to increase the physiologic relevance of human iPS-derived cells, such as cardiomyocytes (iPS-CM). However, forming Engineered Heart Muscle (EHM) typically requires >1 million cells per tissue. Existing miniaturization strategies involve complex approaches not amenable to mass production, limiting the ability to use EHM for iPS-based disease modeling and drug screening. Micro-scale cardiospheres are easily produced, but do not facilitate assembly of elongated muscle or direct force measurements. Here we describe an approach that combines features of EHM and cardiospheres: Micro-Heart Muscle (µHM) arrays, in which elongated muscle fibers are formed in an easily fabricated template, with as few as 2,000 iPS-CM per individual tissue. Within µHM, iPS-CM exhibit uniaxial contractility and alignment, robust sarcomere assembly, and reduced variability and hypersensitivity in drug responsiveness, compared to monolayers with the same cellular composition. µHM mounted onto standard force measurement apparatus exhibited a robust Frank-Starling response to external stretch, and a dose-dependent inotropic response to the ß-adrenergic agonist isoproterenol. Based on the ease of fabrication, the potential for mass production and the small number of cells required to form µHM, this system provides a potentially powerful tool to study cardiomyocyte maturation, disease and cardiotoxicology in vitro.

CRISPR Interference Efficiently Induces Specific and Reversible Gene Silencing in Human iPSCs.

Developing technologies for efficient and scalable disruption of gene expression will provide powerful tools for studying gene function, developmental pathways, and disease mechanisms. Here, we develop clustered regularly interspaced short palindromic repeat interference (CRISPRi) to repress gene expression in human induced pluripotent stem cells (iPSCs). CRISPRi, in which a doxycycline-inducible deactivated Cas9 is fused to a KRAB repression domain, can specifically and reversibly inhibit gene expression in iPSCs and iPSC-derived cardiac progenitors, cardiomyocytes, and T lymphocytes. This gene repression system is tunable and has the potential to silence single alleles. Compared with CRISPR nuclease (CRISPRn), CRISPRi gene repression is more efficient and homogenous across cell populations. The CRISPRi system in iPSCs provides a powerful platform to perform genome-scale screens in a wide range of iPSC-derived cell types, dissect developmental pathways, and model disease.