Randomized, controlled clinical trial of acoustic stimulation to reduce postconcussive symptoms.

OBJECTIVE: Effective interventions are needed to address postconcussive symptoms. We report the results of randomized, sham-controlled trial of Cereset Research™ Standard Operating Procedures (CR-SOP), a noninvasive, closed-loop, allostatic, acoustic stimulation neurotechnology previously shown to improve insomnia. METHODS: Military service members, veterans, or their spouses with persistent symptoms (Neurobehavioral Symptom Inventory [NSI] Score ≥23) after mTBI 3 months to 10 years ago, were randomized to receive 10 sessions of engineered tones linked to brainwaves (LB, intervention), or random engineered tones not linked to brainwaves (NL, sham control). The primary outcome was change in NSI, with secondary outcomes of heart rate variability and self-report measures of sleep, mood, and anxiety. RESULTS: Participants (n = 106, 22% female, mean age 37.1, 2.8 deployments, 3.8 TBIs) were randomized 1:1 to LB or NL, with no significant differences between groups at baseline. Among all study participants, the NSI declined from baseline 41.0 to 27.2 after (P < 0.0001), with gains largely sustained at 3 months (31.2) and 6 months (28.4). However, there were no significant differences between the LB (NSI declined from 39.9 at baseline to 28.2 post-intervention, 31.5 at 3 months, and 29.4 at 6 months) and NL (NSI declined from 41.5 at baseline to 26.2, 29.9, and 27.3, respectively. Similar patterns were observed for the PCL5 and PHQ-9 and there was no difference in HRV between groups. INTERPRETATION: Ten hours of acoustic stimulation while resting in a zero-gravity chair improves postconcussive symptoms. However, linking tones to brain electrical activity did not reduce symptoms more than random tones. REGISTRATION: ClinicalTrials.gov - NCT03649958.

Sequence differences between BAX and BAK core domains manifest as differences in their interactions with lipids.

The B-cell lymphoma 2 (BCL2) family members, BCL2-associated protein X (BAX) and BCL2 homologous antagonist killer (BAK), are required for programmed cell death via the mitochondrial pathway. When cells are stressed, damaged or redundant, the balance of power between the BCL2 family of proteins shifts towards BAX and BAK, allowing their transition from an inactive, monomeric state to a membrane-active oligomeric form that releases cytochrome c from the mitochondrial intermembrane space. That oligomeric state has an essential intermediate, a symmetric homodimer of BAX or BAK. Here we describe crystal structures of dimers of the core domain of BAX, comprising its helices α2-α5. These structures provide an atomic resolution description of the interactions that drive BAX homo-dimerisation and insights into potential interaction between core domain dimers and membrane lipids. The previously identified BAK lipid-interacting sites are not conserved with BAX and are likely to determine the differences between them in their interactions with lipids. We also describe structures of heterodimers of BAK/BAX core domains, yielding further insight into the differences in lipid binding between BAX and BAK.

Structural mapping of PEAK pseudokinase interactions identifies 14-3-3 as a molecular switch for PEAK3 signaling.

PEAK pseudokinases regulate cell migration, invasion and proliferation by recruiting key signaling proteins to the cytoskeleton. Despite lacking catalytic activity, alteration in their expression level is associated with several aggressive cancers. Here, we elucidate the molecular details of key PEAK signaling interactions with the adapter proteins CrkII and Grb2 and the scaffold protein 14-3-3. Our findings rationalize why the dimerization of PEAK proteins has a crucial function in signal transduction and provide biophysical and structural data to unravel binding specificity within the PEAK interactome. We identify a conserved high affinity 14-3-3 motif on PEAK3 and demonstrate its role as a molecular switch to regulate CrkII binding and signaling via Grb2. Together, our studies provide a detailed structural snapshot of PEAK interaction networks and further elucidate how PEAK proteins, especially PEAK3, act as dynamic scaffolds that exploit adapter proteins to control signal transduction in cell growth/motility and cancer.

Perceptions of Health and Wellbeing Among Employees in a Work Integration Social Enterprise in Sweden.

Background: Work Integration Social Enterprises (WISEs) constitute an important vehicle for providing employment opportunities for disadvantaged groups. Objective: The goal of this qualitative case study is to explore perceptions of health and wellbeing among employees working in a WISE located in the Gävleborg region, in east central Sweden. Methods: Data were gathered using 16 in-depth, semi-structured interviews with the social enterprise employees. Results: Findings were categorized into three main categories: the importance of financial independence and societal benefits; team spirit and a sense of belonging; and improved quality of life and wellbeing. Conclusion: The participants perceived that working in the WISE gave them a feeling of freedom and increased their self-esteem because of the possibility to earn an income. Also, they were satisfied with their job (e.g., with regard to work quality and flexibility) and believed that their work contributed to society. Moreover, through working in a WISE, the participants felt a sense of belonging and togetherness through interaction with co-workers and managers, and an improved quality of life for themselves and their families.

Common health assets protocol: a mixed-methods, realist evaluation and economic appraisal of how community-led organisations (CLOs) impact on the health and well-being of people living in deprived areas.

INTRODUCTION: This research investigates how community-led organisations' (CLOs') use of assets-based approaches improves health and well-being, and how that might be different in different contexts. Assets-based approaches involve 'doing with' rather than 'doing to' and bring people in communities together to achieve positive change using their own knowledge, skills and experience. Some studies have shown that such approaches can have a positive effect on health and well-being. However, research is limited, and we know little about which approaches lead to which outcomes and how different contexts might affect success. METHODS AND ANALYSIS: Using a realist approach, we will work with 15 CLOs based in disadvantaged communities in England, Scotland and Northern Ireland. A realist synthesis of review papers, and a policy analysis in different contexts, precedes qualitative interviews and workshops with stakeholders, to find out how CLOs' programmes work and identify existing data. We will explore participants' experiences through: a Q methodology study; participatory photography workshops; qualitative interviews and measure outcomes using a longitudinal survey, with 225 CLO participants, to assess impact for people who connect with the CLOs. An economic analysis will estimate costs and benefits to participants, for different contexts and mechanisms. A 'Lived Experience Panel' of people connected with our CLOs as participants or volunteers, will ensure the appropriateness of the research, interpretation and reporting of findings. ETHICS AND DISSEMINATION: This project, research tools and consent processes have been approved by the Glasgow Caledonian University School of Health and Life Sciences Ethics Committee, and affirmed by Ethics Committees at Bournemouth University, Queen's University Belfast and the University of East London. Common Health Assets does not involve any National Health Service sites, staff or patients.Findings will be presented through social media, project website, blogs, policy briefings, journal articles, conferences and visually in short digital stories, and photographic exhibitions.

Beliefs and Perceived Barriers Regarding Psychedelic-assisted Therapy in a Pilot Study of Service Members and Veterans With a History of Traumatic Brain Injury.

INTRODUCTION: Posttraumatic stress disorder (PTSD) and depression are common in service members and veterans, and the response to currently available treatments is often modest at best. Recent studies suggest potential benefit with psychedelic-assisted therapies (PATs), particularly 3,4-methylenedioxymethamphetamine-assisted therapy for PTSD and psilocybin-assisted therapy for depression. This study examined beliefs and perceived barriers regarding PAT among service members and veterans to inform the delivery of these treatments if they are approved by the FDA. MATERIALS AND METHODS: Twenty-one service members and veterans (67% male, 81% White, and 43% active duty) with a history of traumatic brain injury and co-occurring cognitive and psychological symptoms completed a measure assessing baseline knowledge and views of PAT, read a brief psychoeducation regarding PAT, and then responded to questions related to their beliefs and perceived barriers to PAT. RESULTS: Before psychoeducation, participants reported a neutral view of psychedelic drugs (M = 2.76; range: 1-5), PAT (M = 3.33), and interest in PAT (M = 3.10). After psychoeducation, participants reported a significantly more positive view of psychedelic drugs (M = 3.24, P = .014) and interest in PAT (M = 3.67, P = .016). Overall, participants indicated that they would support PAT availability in medical settings if proven beneficial (M = 4.52; 5 = "agree strongly") and they would support a loved one engaging in PAT (M = 4.29). The most frequently reported health concerns were concern of long-term effects (43%), fear of losing their mind (33%), fear of personality changes (33%), and fear of traumatic brain injury complications (24%). The most frequently endorsed barriers were time commitment, transportation, financial concerns, work, and childcare (33%-19%), with 48% reporting no barriers. CONCLUSIONS: This is the first study to explore beliefs and perceived barriers regarding PAT among service members and veterans. These results indicate that military populations may be interested in PAT, particularly if psychoeducation and outreach regarding these treatments occurred. If FDA approved, it will be important to facilitate command support and address logistical barriers to ensure appropriate access within military contexts.

Cytokine Profiles Differentiate Symptomatic from Asymptomatic PTSD in Service Members and Veterans with Chronic Traumatic Brain Injury.

Traumatic brain injuries (TBI) and posttraumatic stress disorder (PTSD) are commonly observed comorbid occurrences among military service members and veterans (SMVs). In this cross-sectional study, SMVs with a history of TBI were stratified into symptomatic and asymptomatic PTSD groups based on posttraumatic stress checklist-civilian (PCL-C) total scores. Blood-based biomarkers were assessed, and significant differential markers were associated with scores from multiple neurobehavioral self-report assessments. PCL-C cutoffs were total scores >50 (PTSD symptomatic) and <25 (asymptomatic). Cytokines IL6, IL8, TNFα, and IL10 were significantly elevated (p < 0.05−0.001) in the TBI+/PTSD symptomatic group compared to the TBI+/asymptomatic group. Cytokine levels of IL8, TNFα, and IL10 were strongly associated with PCL-C scores (0.356 < r > 0.624 for all, p < 0.01 for all), while TNFα and IL10 were additionally associated with NSI totals (r = 0.285 and r = 0.270, p < 0.05, respectively). This is the first study focused on PTSD symptom severity to report levels of circulating pro-inflammatory IL8, specifically in SMVs with TBI. These data suggest that within the military TBI population, there are unique cytokine profiles that relate to neurobehavioral outcomes associated with TBI and PTSD.

Methodology of the INVestigating traIning assoCiated blasT pAthology (INVICTA) study.

BACKGROUND: Subconcussive blast exposure during military training has been the subject of both anecdotal concerns and reports in the medical literature, but prior studies have often been small and have used inconsistent methods. METHODS: This paper presents the methodology employed in INVestigating traIning assoCiated blasT pAthology (INVICTA) to assess a wide range of aspects of brain function, including immediate and delayed recall, gait and balance, audiologic and oculomotor function, cerebral blood flow, brain electrical activity and neuroimaging and blood biomarkers. RESULTS: A number of the methods employed in INVICTA are relatively easy to reproducibly utilize, and can be completed efficiently, while other measures require greater technical expertise, take longer to complete, or may have logistical challenges. CONCLUSIONS: This presentation of methods used to assess the impact of blast exposure on the brain is intended to facilitate greater uniformity of data collection in this setting, which would enable comparison between different types of blast exposure and environmental circumstances, as well as to facilitate meta-analyses and syntheses across studies.

A selective and orally bioavailable VHL-recruiting PROTAC achieves SMARCA2 degradation in vivo.

Targeted protein degradation offers an alternative modality to classical inhibition and holds the promise of addressing previously undruggable targets to provide novel therapeutic options for patients. Heterobifunctional molecules co-recruit a target protein and an E3 ligase, resulting in ubiquitylation and proteosome-dependent degradation of the target. In the clinic, the oral route of administration is the option of choice but has only been achieved so far by CRBN- recruiting bifunctional degrader molecules. We aimed to achieve orally bioavailable molecules that selectively degrade the BAF Chromatin Remodelling complex ATPase SMARCA2 over its closely related paralogue SMARCA4, to allow in vivo evaluation of the synthetic lethality concept of SMARCA2 dependency in SMARCA4-deficient cancers. Here we outline structure- and property-guided approaches that led to orally bioavailable VHL-recruiting degraders. Our tool compound, ACBI2, shows selective degradation of SMARCA2 over SMARCA4 in ex vivo human whole blood assays and in vivo efficacy in SMARCA4-deficient cancer models. This study demonstrates the feasibility for broadening the E3 ligase and physicochemical space that can be utilised for achieving oral efficacy with bifunctional molecules.

The Redesign and Validation of Multimodal Motion-Assisted Memory Desensitization and Reconsolidation Hardware and Software: Mixed Methods, Modified Delphi-Based Validation Study.

BACKGROUND: In recent years, the delivery of evidence-based therapies targeting posttraumatic stress disorder (PTSD) has been the focus of the Departments of Defense in countries such as Canada, the Netherlands, and the United States. More than 66% of military members continue to experience symptoms of PTSD that significantly impact their daily functioning and quality of life after completing evidence-based treatments. Innovative, engaging, and effective treatments for PTSD are needed. Multimodal motion-assisted memory desensitization and reconsolidation (3MDR) is an exposure-based, virtual reality-supported therapy used to treat military members and veterans with treatment-resistant PTSD. Given the demonstrated efficacy of 3MDR in recently published randomized control trials, there is both an interest in and a need to adapt the intervention to other populations affected by trauma and to improve accessibility to the treatment. OBJECTIVE: We aimed to further innovate, develop, and validate new and existing hardware and software components of 3MDR to enhance its mobility, accessibility, feasibility, and applicability to other populations affected by trauma, including public safety personnel (PSP), via international collaboration. METHODS: This study used a modified Delphi expert consultation method and mixed methods quasi-experimental validation with the purpose of software validation among PSP (first responders, health care providers) participants (N=35). A team of international experts from the Netherlands, the United States, and Canada met on the web on a weekly basis since September 2020 to discuss the adoption of 3MDR in real-world contexts, hardware and software development, and software validation. The evolution of 3MDR hardware and software was undertaken followed by a mixed methods software validation study with triangulation of results to inform the further development of 3MDR. RESULTS: This study resulted in the identification, description, and evolution of hardware and software components and the development of new 3MDR software. Within the software validation, PSP participants widely acknowledged that the newly developed 3MDR software would be applicable and feasible for PSP affected by trauma within their professions. The key themes that emerged from the thematic analysis among the PSP included the desire for occupationally tailored environments, individually tailored immersion, and the applicability of 3MDR beyond military populations. CONCLUSIONS: Within the modified Delphi consultation and software validation study, support for 3MDR as an intervention was communicated. PSP participants perceived that 3MDR was relevant for populations affected by trauma beyond military members and veterans. The resulting hardware and software evolution addressed the recommendations and themes that arose from PSP participants. 3MDR is a novel, structured, exposure-based, virtual reality-supported therapy that is currently used to treat military members and veterans with PTSD. Going forward, it is necessary to innovate and adapt 3MDR, as well as other trauma interventions, to increase effectiveness, accessibility, cost-effectiveness, and efficacy among other populations affected by trauma.

Enhancing exposure therapy for posttraumatic stress disorder (PTSD): a randomized clinical trial of virtual reality and imaginal exposure with a cognitive enhancer.

Posttraumatic stress disorder (PTSD) is a significant public health issue. Yet, there are limited treatment options and no data to suggest which treatment will work for whom. We tested the efficacy of virtual reality exposure (VRE) or prolonged imaginal exposure (PE), augmented with D-cycloserine (DCS) for combat-related PTSD. As an exploratory aim, we examined whether brain-derived neurotrophic factor (BDNF) and fatty acid amide hydrolase (FAAH) moderated treatment response. Military personnel with PTSD (n = 192) were recruited into a multisite double-blind randomized controlled trial to receive nine weeks of VRE or PE, with DCS or placebo. Primary outcome was the improvement in symptom severity. Randomization was stratified by comorbid depression (MDD) and site. Participants in both VRE and PE showed similar meaningful clinical improvement with no difference between the treatment groups. A significant interaction (p = 0.45) suggested VRE was more effective for depressed participants (CAPS difference M = 3.51 [95% CI 1.17-5.86], p = 0.004, ES = 0.14) while PE was more effective for nondepressed participants (M = -8.87 [95% CI -11.33 to -6.40], p < 0.001, ES = -0.44). The main effect of DCS vs. placebo was not significant. Augmentation by MDD interaction (p = 0.073) suggested that depressed participants improved more on placebo (M = -8.43 [95% CI -10.98 to -5.88], p < 0.001, ES = -0.42); DCS and placebo were equally effective for nondepressed participants. There was an apparent moderating effect of BDNF Val66Met polymorphism on DCS augmentation (ES = 0.67). Met66 allele carriers improved more on DCS (ES = -0.25). FAAH 385 A carriers improved more than non-carriers (ES = 0.33), particularly those with MDD (ES = 0.62). This study provides a step toward precision therapeutics for PTSD by demonstrating that comorbid MDD and genetic markers may help guide treatment selection.ClinicalTrials.gov Identifier: NCT01352637.

Production and purification of the PEAK pseudokinases for structural and functional studies.

The PEAK family of pseudokinases, which comprises PEAK1, PEAK2 and PEAK3, are newly identified scaffolds that dynamically assemble oncogenic signaling pathways known to contribute to the development of several aggressive cancers. A striking feature of this unique family of pseudokinase scaffolds is their large multi-domain structure, which allows them to achieve protein complex assemblies through their structural plasticity and functional versatility. Recent structural advances have begun to reveal the critical regulatory elements that control their function. Specifically, the dimer-dependent scaffolding activity of PEAK pseudokinases is emerging as a critical mechanism for their signaling function, in addition to their ability to hetero-associate to form higher-order regulatory networks to diversify and amplify their signaling output. Here, we present a suite of techniques that enable the efficient expression and purification of PEAK proteins for functional characterization.

Distinct PEAK3 interactors and outputs expand the signaling potential of the PEAK pseudokinase family.

The pseudokinase scaffolds PEAK1 and PEAK2 are implicated in cancer cell migration and metastasis. We characterized the regulation and role of the third family member PEAK3 in cell signaling. Similar to PEAK1 and PEAK2, PEAK3 formed both homotypic and heterotypic complexes. In addition, like PEAK1, it bound to the adaptors Grb2 and CrkII. However, unlike PEAK1 and PEAK2, homodimerized PEAK3 also interacted with the ARF GTPase-activating protein ASAP1, the E3 ubiquitin ligase Cbl, and the kinase PYK2. Dimerization and subsequent phosphorylation on Tyr24, likely by a Src family kinase, were required for the binding of PEAK3 to Grb2 and ASAP1. Interactions with Grb2, CrkII, ASAP1, Cbl, and PYK2 exhibited contrasting dynamics upon cell stimulation with epidermal growth factor (EGF), in part due to PEAK3 dephosphorylation mediated by the phosphatase PTPN12. Overexpressing PEAK3 in mesenchymal-like MDA-MB-231 breast cancer cells enhanced cell elongation in a manner dependent on PEAK3 dimerization, and manipulation of PEAK3 expression demonstrated a positive role for this scaffold in regulating cell migration. Overexpressing PEAK3 in PEAK1/2 double-knockout MCF-10A breast epithelial cells enhanced acinar growth, impaired basement membrane integrity, and promoted invasion in three-dimensional cultures, with the latter two effects dependent on the binding of PEAK3 to Grb2 and ASAP1. PEAK1 and PEAK2 quantitatively and temporally influenced PEAK3 function. These findings characterize PEAK3 as an integral, signal-diversifying member of the PEAK family with scaffolding roles that promote cell proliferation, migration, and invasion.

Sex Differences in Behavioral Symptoms and the Levels of Circulating GFAP, Tau, and NfL in Patients With Traumatic Brain Injury.

Traumatic brain injury (TBI) affects millions of Americans each year and has been shown to disproportionately impact those subject to greater disparities in health. Female sex is one factor that has been associated with disparities in health outcomes, including in TBI, but sex differences in biomarker levels and behavioral outcomes after TBI are underexplored. This study included participants with both blunt and blast TBI with majority rating their TBI as mild. Time since injury was 5.4 (2.0, 15.5) years for females and 6.8 (2.4, 11.3) years for males. The aim of this cross sectional study is to investigate the relationship between postconcussive, depression, and post-traumatic stress disorder (PTSD) symptoms, as well as health related quality of life (HRQOL), and the levels of glial fibrillary acidic protein (GFAP), total tau (t-tau), neurofilament light chain (NfL), and ubiquitin C-terminal hydrolase-L1 (UCH-L1). Behavioral outcomes were evaluated with the Neurobehavioral Symptom Inventory (NSI), Patient Health Questionnaire-9 (PHQ-9), PTSD Checklist- Civilian Version (PCL-C), short form (SF)-36, and plasma levels of total tau, GFAP, NfL, and UCHL-1 measured with the Simoa-HDX. We observed that females had significantly higher levels of GFAP and tau (ps < 0.05), and higher PHQ-9 scores, NSI total scores, NSI- vestibular, NSI-somatosensory, NSI-affective sub-scale scores (ps < 0.05)), than males. In addition, females had lower scores in HRQOL outcomes of role limitations due to emotional problems, vitality, emotional well-being, social functioning, and pain compared to males (ps < 0.05). Correlation analysis showed positive associations between levels of tau and the NSI-total and NSI-cognitive sub-scale scores (ps < 0.05) in females. No significant associations were found for NfL or GFAP with NSI scores. For female participants, negative correlations were observed between tau and NfL concentrations and the SF-36 physical function subscale (ps < 0.05), as well as tau and the social function subscale (p < 0.001), while GFAP levels positively correlated with role limitations due to emotional problems (p = 0.004). No significant associations were observed in males. Our findings suggest that sex differences exist in TBI-related behavioral outcomes, as well as levels of biomarkers associated with brain injury, and that the relationship between biomarker levels and behavioral outcomes is more evident in females than males. Future studies are warranted to corroborate these results, and to determine the implications for prognosis and treatment. The identification of candidate TBI biomarkers may lead to development of individualized treatment guidelines.

Combat-Related Post-traumatic Stress Disorder: A Case Report of Virtual Reality Graded Exposure Therapy With Physiological Monitoring in a U.S. Navy Officer and a U.S. Army Officer.

The U.S. Department of Defense (DoD) and the Department of Veterans Affairs (DVA) seek to enhance the efficacy of treatments for warriors with post-traumatic stress disorder (PTSD) secondary to their combat deployments to Iraq and/or Afghanistan. Virtual Reality Graded Exposure Therapy (VR-GET) with arousal control has shown particular promise in reducing the symptom severity of PTSD in combat veterans. In this report, we describe the outcome of VR-GET for the treatment of combat-related PTSD in two combat veterans, neither of whom had received treatment for PTSD in the initial years after their return from combat duty.

Structural basis for small molecule targeting of Doublecortin Like Kinase 1 with DCLK1-IN-1.

Doublecortin-like kinase 1 (DCLK1) is an understudied bi-functional kinase with a proven role in tumour growth and development. However, the presence of tissue-specific spliced DCLK1 isoforms with distinct biological functions have challenged the development of effective strategies to understand the role of DCLK1 in oncogenesis. Recently, DCLK1-IN-1 was reported as a highly selective DCLK1 inhibitor, a powerful tool to dissect DCLK1 biological functions. Here, we report the crystal structures of DCLK1 kinase domain in complex with DCLK1-IN-1 and its precursors. Combined, our data rationalises the structure-activity relationship that informed the development of DCLK1-IN-1 and provides the basis for the high selectivity of DCLK1-IN-1, with DCLK1-IN-1 inducing a drastic conformational change of the ATP binding site. We demonstrate that DCLK1-IN-1 binds DCLK1 long isoforms but does not prevent DCLK1's Microtubule-Associated Protein (MAP) function. Together, our work provides an invaluable structural platform to further the design of isoform-specific DCLK1 modulators for therapeutic intervention.

Synthesis of C-Mannosylated Glycopeptides Enabled by Ni-Catalyzed Photoreductive Cross-Coupling Reactions.

The biological functions of tryptophan C-mannosylation are poorly understood, in part, due to a dearth of methods for preparing pure glycopeptides and glycoproteins with this modification. To address this issue, efficient and scalable methods are required for installing this protein modification. Here, we describe unique Ni-catalyzed cross-coupling conditions that utilize photocatalysis or a Hantzsch ester photoreductant to couple glycosyl halides with (hetero)aryl bromides, thereby enabling the α-C-mannosylation of 2-bromo-tryptophan, peptides thereof, and (hetero)aryl bromides more generally. We also report that 2-(α-d-mannopyranosyl)-L-tryptophan undergoes facile anomerization in the presence of acid: something that must be considered when preparing and handling peptides with this modification. These developments enabled the first automated solid-phase peptide syntheses of C-mannosylated glycopeptides, which we used to map the epitope of an antibody, as well as providing the first verified synthesis of Carmo-HrTH-I, a C-mannosylated insect hormone. To complement this approach, we also performed late-stage tryptophan C-mannosylation on a diverse array of peptides, demonstrating the broad scope and utility of this methodology for preparing glycopeptides.

Structure-Guided Development of Potent Benzoylurea Inhibitors of BCL-XL and BCL-2.

The BCL-2 family of proteins (including the prosurvival proteins BCL-2, BCL-XL, and MCL-1) is an important target for the development of novel anticancer therapeutics. Despite the challenges of targeting protein-protein interaction (PPI) interfaces with small molecules, a number of inhibitors (called BH3 mimetics) have entered the clinic and the BCL-2 inhibitor, ABT-199/venetoclax, is already proving transformative. For BCL-XL, new validated chemical series are desirable. Here, we outline the crystallography-guided development of a structurally distinct series of BCL-XL/BCL-2 inhibitors based on a benzoylurea scaffold, originally proposed as α-helix mimetics. We describe structure-guided exploration of a cryptic "p5" pocket identified in BCL-XL. This work yields novel inhibitors with submicromolar binding, with marked selectivity toward BCL-XL. Extension into the hydrophobic p2 pocket yielded the most potent inhibitor in the series, binding strongly to BCL-XL and BCL-2 (nanomolar-range half-maximal inhibitory concentration (IC50)) and displaying mechanism-based killing in cells engineered to depend on BCL-XL for survival.

BAK core dimers bind lipids and can be bridged by them.

BAK and BAX are essential mediators of apoptosis that oligomerize in response to death cues, thereby causing permeabilization of the mitochondrial outer membrane. Their transition from quiescent monomers to pore-forming oligomers involves a well-characterized symmetric dimer intermediate. However, no essential secondary interface that can be disrupted by mutagenesis has been identified. Here we describe crystal structures of human BAK core domain (α2-α5) dimers that reveal preferred binding sites for membrane lipids and detergents. The phospholipid headgroup and one acyl chain (sn2) associate with one core dimer while the other acyl chain (sn1) associates with a neighboring core dimer, suggesting a mechanism by which lipids contribute to the oligomerization of BAK. Our data support a model in which, unlike for other pore-forming proteins whose monomers assemble into oligomers primarily through protein-protein interfaces, the membrane itself plays a role in BAK and BAX oligomerization.

Excessive daytime sleepiness is associated with altered gene expression in military personnel and veterans with posttraumatic stress disorder: an RNA sequencing study.

STUDY OBJECTIVES: Posttraumatic stress disorder (PTSD) is a common condition for military personnel and veterans. PTSD has been shown to impact gene expression, however, to date no study has examined comorbid conditions which may also impact gene expression, for example, excessive daytime sleepiness (EDS). As such, this study sought to examine gene expression using RNA sequencing across three group comparisons of military personnel and veterans: (1) PTSD with EDS (PTSDwEDS) versus PTSD without EDS (PTSDw/outEDS), (2) Controls (no PTSD or EDS) versus PTSDwEDS, and (3) Controls versus PTSDw/outEDS. METHODS: We performed experimental RNA-seq using Illumina's HiSeq 2500 Sequencing System. We also used Ingenuity Pathway Analysis (IPA), a bioinformatics application, to identify gene pathways and networks which may be disrupted. RESULTS: There were only two genes that were significantly dysregulated between the Controls and PTSDw/outEDS, therefore IPA analysis was not conducted. However, comparisons revealed that there was significant gene dysregulation between Controls and the PTSDwEDS (251 genes), and the PTSDwEDS versus the PTSDw/outEDS (1,873 genes) groups. Four candidate networks were identified via the IPA software for analysis. Significantly dysregulated genes across the four candidate networks were associated with sleep and circadian function, metabolism, mitochondrial production and function, ubiquitination, and the glutamate system. CONCLUSIONS: These results suggest that PTSD with concurrent EDS is associated with gene dysregulation. This dysregulation may present additional biological and health consequences for these military personnel and veterans. Further research, to track these gene changes over time and to determine the cause of the EDS reported, is vital.