Somatic CpG hypermutation is associated with mismatch repair deficiency in cancer.

Somatic hypermutation in cancer has gained momentum with the increased use of tumour mutation burden as a biomarker for immune checkpoint inhibitors. Spontaneous deamination of 5-methylcytosine to thymine at CpG dinucleotides is one of the most ubiquitous endogenous mutational processes in normal and cancer cells. Here, we performed a systematic investigation of somatic CpG hypermutation at a pan-cancer level. We studied 30,191 cancer patients and 103 cancer types and developed an algorithm to identify somatic CpG hypermutation. Across cancer types, we observed the highest prevalence in paediatric leukaemia (3.5%), paediatric high-grade glioma (1.7%), and colorectal cancer (1%). We discovered germline variants and somatic mutations in the mismatch repair complex MutSα (MSH2-MSH6) as genetic drivers of somatic CpG hypermutation in cancer, which frequently converged on CpG sites and TP53 driver mutations. We further observe an association between somatic CpG hypermutation and response to immune checkpoint inhibitors. Overall, our study identified novel cancer types that display somatic CpG hypermutation, strong association with MutSα-deficiency, and potential utility in cancer immunotherapy.

Multi-omic analysis of SDHB-deficient pheochromocytomas and paragangliomas identifies metastasis and treatment-related molecular profiles.

Hereditary SDHB-mutant pheochromocytomas (PC) and paragangliomas (PG) are rare tumours with a high propensity to metastasize although their clinical behaviour is unpredictable. To characterize the genomic landscape of these tumours and identify metastasis biomarkers, we performed multi-omic analysis on 94 tumours from 79 patients using seven molecular methods. Sympathetic (chromaffin cell) and parasympathetic (non-chromaffin cell) PCPG had distinct molecular profiles reflecting their cell-of-origin and biochemical profile. TERT and ATRX-alterations were associated with metastatic PCPG and these tumours had an increased mutation load, and distinct transcriptional and telomeric features. Most PCPG had quiet genomes with some rare co-operative driver events observed, including EPAS1/HIF-2α mutations. Two mechanisms of acquired resistance to DNA alkylating chemotherapies were also detected - MGMT overexpression and mismatch repair-deficiency causing hypermutation. Our comprehensive multi-omic analysis of SDHB-mutant PCPG therefore identified features of metastatic disease and treatment response, expanding our understanding of these rare neuroendocrine tumours.

Insulin dependence negatively impacts outcomes in anterior cervical discectomy with fusions: a 10-year retrospective analysis.

BACKGROUND CONTEXT: Although anterior cervical discectomy and fusion (ACDF) procedures for cervical spine disease have been increasing amid a growing diabetic patient population, there is a paucity of literature focusing on insulin-dependence as a risk-factor for postoperative ACDF complications. PURPOSE: To evaluate the differential impact of insulin dependence on perioperative outcomes including total length of stay, surgical, and medical complications within thirty days following ACDF. STUDY DESIGN/SETTING: A retrospective cohort, large multicenter database study. PATIENT SAMPLE: The American College of Surgeons National Surgical Quality Improvement Program database was queried to retrospectively identify patients who had undergone ACDF between 2011 and 2021 using the Current Procedural Terminology code 22551. OUTCOME MEASURES: Perioperative surgical and medical complications. METHODS: The study population was divided into 3 groups 1) insulin-dependent diabetes mellitus (IDDM), 2) noninsulin-dependent diabetes mellitus (NIDDM), and 3) no diabetes mellitus (non-DM). One-way analysis of variance for continuous variables and chi-square tests for categorical variables were used to identify differences in perioperative variables between the 3 groups. Multivariable logistic regression analysis assessed the effect of diabetes mellitus status on postoperative medical and surgical outcomes. RESULTS: A total of 85,758 ACDF procedures were identified between 2011 and 2021, of which 5,178 were IDDM, 9,652 were NIDDM, and 70,982 were non-DM. The rates of surgical and medical complication varied between the 3 groups. IDDM patients had the highest rates of at least one medical complication (6.1%). Only IDDM increased the risk for medical complications (OR: 1.320, 95% CI [1.144-1.518]) and extended hospital length of stay (LOS) (OR: 1.244, 95% CI [1.071-1.441]) following a multivariate logistic regression analysis. CONCLUSION: Patients with IDDM were at an increased risk for postoperative medical complications and extended hospital LOS. Personalized postoperative management, guided by risk assessment is indicated for this population. These findings can be used to improve risk stratification and informed consent for DM patients who are insulin dependent.

Mechanosensitive ion channels MSL8, MSL9, and MSL10 have environmentally sensitive intrinsically disordered regions with distinct biophysical characteristics in vitro.

Intrinsically disordered protein regions (IDRs) are highly dynamic sequences that rapidly sample a collection of conformations over time. In the past several decades, IDRs have emerged as a major component of many proteomes, comprising ~30% of all eukaryotic protein sequences. Proteins with IDRs function in a wide range of biological pathways and are notably enriched in signaling cascades that respond to environmental stresses. Here, we identify and characterize intrinsic disorder in the soluble cytoplasmic N-terminal domains of MSL8, MSL9, and MSL10, three members of the MscS-like (MSL) family of mechanosensitive ion channels. In plants, MSL channels are proposed to mediate cell and organelle osmotic homeostasis. Bioinformatic tools unanimously predicted that the cytosolic N-termini of MSL channels are intrinsically disordered. We examined the N-terminus of MSL10 (MSL10N) as an exemplar of these IDRs and circular dichroism spectroscopy confirms its disorder. MSL10N adopted a predominately helical structure when exposed to the helix-inducing compound trifluoroethanol (TFE). Furthermore, in the presence of molecular crowding agents, MSL10N underwent structural changes and exhibited alterations to its homotypic interaction favorability. Lastly, interrogations of collective behavior via in vitro imaging of condensates indicated that MSL8N, MSL9N, and MSL10N have sharply differing propensities for self-assembly into condensates, both inherently and in response to salt, temperature, and molecular crowding. Taken together, these data establish the N-termini of MSL channels as intrinsically disordered regions with distinct biophysical properties and the potential to respond uniquely to changes in their physiochemical environment.

SHEPHARD: a modular and extensible software architecture for analyzing and annotating large protein datasets.

MOTIVATION: The emergence of high-throughput experiments and high-resolution computational predictions has led to an explosion in the quality and volume of protein sequence annotations at proteomic scales. Unfortunately, sanity checking, integrating, and analyzing complex sequence annotations remains logistically challenging and introduces a major barrier to entry for even superficial integrative bioinformatics. RESULTS: To address this technical burden, we have developed SHEPHARD, a Python framework that trivializes large-scale integrative protein bioinformatics. SHEPHARD combines an object-oriented hierarchical data structure with database-like features, enabling programmatic annotation, integration, and analysis of complex datatypes. Importantly SHEPHARD is easy to use and enables a Pythonic interrogation of largescale protein datasets with millions of unique annotations. We use SHEPHARD to examine three orthogonal proteome-wide questions relating protein sequence to molecular function, illustrating its ability to uncover novel biology. AVAILABILITY AND IMPLEMENTATION: We provided SHEPHARD as both a stand-alone software package (https://github.com/holehouse-lab/shephard), and as a Google Colab notebook with a collection of precomputed proteome-wide annotations (https://github.com/holehouse-lab/shephard-colab).

Noticing Acute Changes in Health in Long-Term Care Residents.

PURPOSE: Early signs of acute conditions and increased fall risk often go unrecognized in patients in long-term care facilities. The aim of this study was to examine how healthcare staff identify and act on changes in health status in this patient population. DESIGN: A qualitative study design was used for this study. METHODS: Six focus groups across two Department of Veterans Affairs long-term care facilities were conducted with 26 interdisciplinary healthcare staff members. Using thematic content analysis, the team preliminarily coded based on interview questions, reviewed and discussed emerging themes, and agreed on the resultant coding scheme for each category with additional independent scientist review. RESULTS: Themes included describing and explaining how "normal" or expected behavior is identified by staff, noticing changes in a resident, determining the significance of the change, hypothesizing reasons for an observed change, response to an observed change, and resolution of the clinical change. CONCLUSIONS: Despite limited training in formal assessment methods, long-term care staff have developed methods to conduct ongoing assessments of the residents. This technique, individual phenotyping, often identifies acute changes; however, the lack of formal methods, language, or tools to communicate the changes means that these assessments are not often formalized in a manner that informs the residents' changing care needs. CLINICAL RELEVANCE TO THE PRACTICE OF REHABILITATION NURSING: More formal objective measures of health change are needed to assist long-term care staff in expressing and interpreting the subjective phenotype changes into objective, easily communicated health status changes. This is particularly important for acute health changes and impending falls, both of which are associated with acute hospitalization.

Using spreading activation to understand repetitive negative thinking.

Repetitive negative thinking (RNT) describes a recursive, unproductive pattern of thought that is commonly observed in individuals who experience anxiety and depression. Past research on RNT has primarily relied on self-report, which fails to capture the potential mechanisms that underlie the persistence of maladaptive thought. We investigated whether RNT may be maintained by a negatively biased semantic network. The present study used a modified free association task to assess state RNT. Following the presentation of a valenced (positive, neutral, negative) cue word, participants generated a series of free associates, which allowed for the dynamic progression of responses. State RNT was conceptualised as the length of consecutive, negatively valenced free associates (i.e. chains). Participants also completed two self-report measures that assessed trait RNT and trait negative affect. Within a structural equation model, negative (but not positive or neutral) response chain length positively predicted trait RNT and negative affect, and this was only the case for positive (but not negative or neutral) cue words. These results suggest that RNT tendencies may be reflected in semantic retrieval and can be assessed without self-report.

Association Between Functional Tricuspid Regurgitation and Mortality Following Cardiac Surgery.

Background: Current guidelines recommend concomitant repair of certain non-severe cases of tricuspid regurgitation (TR) in patients undergoing cardiac surgery, but the prognostic relevance and postsurgical impact of the TR remain uncertain. Objectives: The purpose of this study was to determine the prognostic impact of functional TR in patients undergoing diverse cardiac surgeries and to examine the effect-modifying role of patient characteristics in patients in whom TR confers a greater risk of adverse outcomes. Methods: Patients undergoing coronary artery bypass, aortic, and mitral valve surgery were included. Patients with severe TR, organic tricuspid valve pathology, undergoing tricuspid valve surgery or without a recent preoperative echocardiogram were excluded. Clinical variables were extracted from the Society of Thoracic Surgeons Adult Cardiac Surgery Database. An independent cohort was used for external validation. Results: Of 2,119 patients (mean age 67.4 years; 29% females), TR severity was moderate in 185 (9%), mild in 636 (30%), trivial in 1,126 (53%), and absent in 172 (8%). There were 238 deaths during the median follow-up period of 2.6 years. After adjusting for relevant factors, moderate TR was found to be independently associated with mid-term mortality (HR: 2.58; 95% CI: 1.22-5.47) and with in-hospital mortality or major morbidity (OR: 3.18; 95% CI: 1.37-7.42). The association between TR and mortality was apparent when preoperative pulmonary artery systolic pressure was <40 mm Hg but not ≥40 mm Hg (P for interaction = 0.036). Conclusions: In this diverse cohort of contemporary cardiac surgery patients, moderate functional TR was associated with increased mortality and major morbidity, particularly in the absence of pulmonary hypertension.

Single-nuclei and bulk-tissue gene-expression analysis of pheochromocytoma and paraganglioma links disease subtypes with tumor microenvironment.

Pheochromocytomas (PC) and paragangliomas (PG) are rare neuroendocrine tumors associated with autonomic nerves. Here we use single-nuclei RNA-seq and bulk-tissue gene-expression data to characterize the cellular composition of PCPG and normal adrenal tissues, refine tumor gene-expression subtypes and make clinical and genotypic associations. We confirm seven PCPG gene-expression subtypes with significant genotype and clinical associations. Tumors with mutations in VHL, SDH-encoding genes (SDHx) or MAML3-fusions are characterized by hypoxia-inducible factor signaling and neoangiogenesis. PCPG have few infiltrating lymphocytes but abundant macrophages. While neoplastic cells transcriptionally resemble mature chromaffin cells, early chromaffin and neuroblast markers are also features of some PCPG subtypes. The gene-expression profile of metastatic SDHx-related PCPG indicates these tumors have elevated cellular proliferation and a lower number of non-neoplastic Schwann-cell-like cells, while GPR139 is a potential theranostic target. Our findings therefore clarify the diverse transcriptional programs and cellular composition of PCPG and identify biomarkers of potential clinical significance.

PTSD Avoidance Symptoms Associated With Alcohol Craving in Treatment-Seeking Veteran Population.

Objective: Posttraumatic stress disorder (PTSD) is one of the most frequently treated behavioral health conditions within the Department of Veterans Affairs and often co-occurs with alcohol or substance use. Past research suggests that alcohol and/or substance use may be used to cope with PTSD symptoms but there are inconsistent findings in how specific PTSD symptom clusters are associated with alcohol use disorder (AUD) or substance use disorder (SUD). Evaluating the relationship between PTSD symptom clusters and craving for individual drug of dependence may help explain these ambiguous results. Methods: Veterans (N = 167) recently engaged in mental health residential treatment were recruited to participate in a semi-structured diagnostic interview (Structured Clinical Interview for the DSM-5, Research Version [SCID-5-RV]) to assess for past 12-month history of AUD/SUD. Participants also completed the Posttraumatic Stress Disorder Checklist for DSM-5 (PCL-5) to assess PTSD symptoms. Results: Covarying for severity of alcohol use, avoidance symptoms were significantly associated with alcohol craving for veterans with alcohol as their drug of dependence. Covarying for severity of stimulant use, no PTSD symptom clusters were associated with stimulant craving for veterans with stimulants as their drug of dependence. Conclusions: Veterans with high levels of PTSD avoidance symptoms may experience alcohol craving symptoms because they believe that alcohol use will eliminate or alleviate thoughts, feelings, or external reminders of the trauma. These results have important clinical implications in the treatment of co-occurring PTSD and AUD.

Developing Acute Event Risk Profiles for Older Adults with Dementia in Long-Term Care Using Motor Behavior Clusters Derived from Deep Learning.

OBJECTIVES: This paper uses deep (machine) learning techniques to develop and test how motor behaviors, derived from location and movement sensor tracking data, may be associated with falls, delirium, and urinary tract infections (UTIs) in long-term care (LTC) residents. DESIGN: Longitudinal observational study. SETTING AND PARTICIPANTS: A total of 23 LTC residents (81,323 observations) with cognitive impairment or dementia in 2 northeast Department of Veterans Affairs LTC facilities. METHODS: More than 18 months of continuous (24/7) monitoring of motor behavior and activity levels used objective radiofrequency identification sensor data to track and record movement data. Occurrence of acute events was recorded each week. Unsupervised deep learning models were used to classify motor behaviors into 5 clusters; supervised decision tree algorithms used these clusters to predict acute health events (falls, delirium, and UTIs) the week before the week of the event. RESULTS: Motor behaviors were classified into 5 categories (Silhouette score = 0.67), and these were significantly different from each other. Motor behavior classifications were sensitive and specific to falls, delirium, and UTI predictions 1 week before the week of the event (sensitivity range = 0.88-0.91; specificity range = 0.71-0.88). CONCLUSION AND IMPLICATIONS: Intraindividual changes in motor behaviors predict some of the most common and detrimental acute events in LTC populations. Study findings suggest real-time locating system sensor data and machine learning techniques may be used in clinical applications to effectively prevent falls and lead to the earlier recognition of risk for delirium and UTIs in this vulnerable population.

Molecular dissection of RbpA-mediated regulation of fidaxomicin sensitivity in mycobacteria.

RNA polymerase (RNAP) binding protein A (RbpA) is essential for mycobacterial viability and regulates transcription initiation by increasing the stability of the RNAP-promoter open complex (RPo). RbpA consists of four domains: an N-terminal tail (NTT), a core domain (CD), a basic linker, and a sigma interaction domain. We have previously shown that truncation of the RbpA NTT and CD increases RPo stabilization by RbpA, implying that these domains inhibit this activity of RbpA. Previously published structural studies showed that the NTT and CD are positioned near multiple RNAP-σA holoenzyme functional domains and predict that the RbpA NTT contributes specific amino acids to the binding site of the antibiotic fidaxomicin (Fdx), which inhibits the formation of the RPo complex. Furthermore, deletion of the NTT results in decreased Mycobacterium smegmatis sensitivity to Fdx, but whether this is caused by a loss in Fdx binding is unknown. We generated a panel of rbpA mutants and found that the RbpA NTT residues predicted to directly interact with Fdx are partially responsible for RbpA-dependent Fdx activity in vitro, while multiple additional RbpA domains contribute to Fdx activity in vivo. Specifically, our results suggest that the RPo-stabilizing activity of RbpA decreases Fdx activity in vivo. In support of the association between RPo stability and Fdx activity, we find that another factor that promotes RPo stability in bacteria, CarD, also impacts to Fdx sensitivity. Our findings highlight how RbpA and other factors may influence RNAP dynamics to affect Fdx sensitivity.

Shame as a Mediator in the Association Between Emotion Dysregulation and Posttraumatic Stress Disorder Symptom Reductions Among Combat Veterans in a Residential Treatment Program.

Emotion dysregulation (ED) can be defined as one's inability to effectively respond to and manage internal experiences and the expression of emotion. ED has been linked to the development and maintenance of posttraumatic stress disorder (PTSD), with recent research suggesting that reductions in ED may predict improved treatment outcomes among both civilian and veteran populations. However, few studies have examined how changes in ED may predict treatment outcomes among veterans with PTSD and whether certain core features of PTSD, such as shame, may act as potential mediators in the association between ED and PTSD symptom reductions. The present study sought to explore facets of ED, feelings of shame, and PTSD symptoms among 43 combat veterans upon their admission and discharge to a residential PTSD program. The results demonstrated that all variables of interest significantly decreased from admission to discharge, ds = 0.75-1.84. Correlations indicated that reductions in ED, R2 = .184, and shame, R2 = .228, were associated with reductions in PTSD symptoms. However, the association between reductions in ED and PTSD was significantly mediated by reductions in shame. Overall, these results suggest that higher levels of emotion regulation may partially affect PTSD symptoms through reductions in shame. This may explain the efficacy of frontline PTSD treatments, as they explicitly focus on the processing of one's traumatic experience by reducing PTSD symptoms through regulation techniques that target emotional-behavioral cycles, which may include the shame-withdraw cycle.

Tumor mutational burden and purity adjustment before and after treatment with temozolomide in 27 paired samples of glioblastoma: a prospective study.

Treatment of glioblastoma (GBM) remains a challenging task, with limited treatment options, none offering a cure. Immune therapy has proven effective across different cancers with remarkable response rates. Tumor mutational burden (TMB) is a marker of response, but technical and methodological differences in TMB estimates have made a proper assessment and comparison challenging. Here, we analyzed a prospective collection of paired samples from 35 patients with newly diagnosed GBM, all of whom were wild-type (WT) for isocitrate dehydrogenase, before and after treatment with radiotherapy and temozolomide. Seven patients (20%) had O6-methylguanine-DNA methyltransferase-methylated tumors. Six patients (17%) had two relapse surgeries, and tissue from all three surgeries was collected. We found that accurate evaluation of TMB was confounded by high variability in the cancer cell fraction of relapse samples. To ameliorate this, we developed a model to adjust for tumor purity based on the relative density distribution of variant allele frequencies in each primary-relapse pair. Additionally, we examined the mutation spectra of shared and private mutations. After tumor purity adjustment, we found TMB comparison reliable in tumors with tumor purity between 15% and 40%, resulting in 27/35 patients (77.1%). TMB remained unchanged from 0.65 mutations per megabase (Mb) to 0.67/Mb before and after treatment, respectively. Examination of the mutation spectra revealed a dominance of C > T transitions at CpG sites in both shared and relapse-private mutations, consistent with cytosine deamination and the clock-like mutational signature 1. We present and apply a cellularity correction approach that enables more accurate assessment of TMB in paired tumor samples. We did not find a significant increase in TMB after correcting for cancer cell fraction. Our study raises significant concerns when determining TMB. Although a small sample size, corrected TMB can have a clinical significance when stratifying patients to experimental treatment, for example, immune checkpoint therapy.

Shame as a Moderator between Emotion Dysregulation and Posttraumatic Stress Disorder Severity among Combat Veterans Seeking Residential Treatment.

BACKGROUND: Growing evidence suggests that emotion dysregulation may be predictive of posttraumatic stress disorder (PTSD) severity, with emotional non-acceptance, lack of counter strategies, impulse control deficits, and lack of emotional awareness all being positive predictors. However, findings have been mixed. This may be due to no previous study examining the association between emotion dysregulation and PTSD severity with shame, a maintaining factor of PTSD, as a potential moderator. METHODS: The present study examined the relationship between emotion dysregulation, shame, and PTSD severity among 78 male combat veterans (mean age = 42.19) upon their admission to a residential combat PTSD program. RESULTS: Results demonstrated that shame and all facets of emotion dysregulation (except lack of emotional awareness & clarity) were positively associated with PTSD severity. Shame moderated the relationship between lack of emotional awareness and strategies. Among those at or below the sample mean on shame, lack of access to strategies was a positive predictor of PTSD severity. Comparatively, among those with high levels of shame, emotional awareness predicted greater PTSD severity, while among those with low levels of shame, emotional awareness predicted lower PTSD severity. LIMITATIONS: Limitations included reliance on self-report questionnaires and an all-male sample. CONCLUSIONS: Thus, emotion dysregulation may only predict PTSD severity among those reporting lower levels of shame, suggesting the importance of addressing shame as well as emotion dysregulation deficits among those with PTSD. Moreover, emotional awareness may be either a risk or protective factor depending on levels of shame.

TERT structural rearrangements in metastatic pheochromocytomas.

Pheochromocytomas (PC) and paragangliomas (PGL) are endocrine tumors for which the genetic and clinicopathological features of metastatic progression remain incompletely understood. As a result, the risk of metastasis from a primary tumor cannot be predicted. Early diagnosis of individuals at high risk of developing metastases is clinically important and the identification of new biomarkers that are predictive of metastatic potential is of high value. Activation of TERT has been associated with a number of malignant tumors, including PC/PGL. However, the mechanism of TERT activation in the majority of PC/PGL remains unclear. As TERT promoter mutations occur rarely in PC/PGL, we hypothesized that other mechanisms - such as structural variations - may underlie TERT activation in these tumors. From 35 PC and four PGL, we identified three primary PCs that developed metastases with elevated TERT expression, each of which lacked TERT promoter mutations and promoter DNA methylation. Using whole genome sequencing, we identified somatic structural alterations proximal to the TERT locus in two of these tumors. In both tumors, the genomic rearrangements led to the positioning of super-enhancers proximal to the TERT promoter, that are likely responsible for the activation of the normally tightly repressed TERT expression in chromaffin cells.