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OBJECTIVE: Resistance to chemo-radiation therapy is a substantial obstacle that compromises treatment of advanced cervical cancer. The objective of this study was to investigate if a proteomic panel associated with radioresistance could predict survival of patients with locally advanced cervical cancer. METHODS: A total of 181 frozen tissue samples were prospectively obtained from patients with locally advanced cervical cancer before chemoradiation. Expression levels of 22 total and phosphorylated proteins were evaluated using well-based reverse phase protein arrays. Selected proteins were validated with western blotting analysis and immunohistochemistry. Performances of models were internally and externally validated. RESULTS: Unsupervised clustering stratified patients into three major groups with different overall survival (OS, P = 0.001) and progression-free survival (PFS, P = 0.003) based on detection of BCL2, HER2, CD133, CAIX, and ERCC1. Reverse-phase protein array results significantly correlated with western blotting results (R(2) = 0.856). The C-index of model was higher than clinical model in the prediction of OS (C-index: 0.86 and 0.62, respectively) and PFS (C-index: 0.82 and 0.64, respectively). The Kaplan-Meier survival curve showed a dose-dependent prognostic significance of risk score for PFS and OS. Multivariable Cox proportional hazard model confirmed that the risk score was an independent predictor of PFS (HR: 1.6; 95% CI: 1.4–1.9; P < 0.001) and OS (HR: 2.1; 95% CI: 1.7–2.5; P < 0.001). CONCLUSION: A proteomic panel of BCL2, HER2, CD133, CAIX, and ERCC1 independently predicted survival in locally advanced cervical cancer patients. This prediction model can help identify chemoradiation responsive tumors and improve prediction for clinical outcome of cervical cancer patients.

Anti-tumor immune polarization is a key predictor of clinical outcomes to cancer therapy. An emerging concept influencing clinical outcome involves the spatial location of CD8+T cells, within the tumor. Our earlier work demonstrated immunosuppressive effects of NOS2/ COX2 tumor expression. Here, we show that NOS2/COX2 levels influence the polarization and spatial location of lymphoid cells including CD8+T cells. Importantly, elevated tumor NOS2/COX2 correlated with exclusion of CD8+T cells from the tumor epithelium. In contrast, tumors expressing low NOS2/COX2 had increased CD8+T cell penetration into the tumor epithelium. Consistent with a causative relationship between these observations, pharmacological inhibition of COX2 with indomethacin dramatically reduced tumor growth of the 4T1 model of TNBC in bothWTandNos2-/-mice. This regimen led to complete tumor regression in ∼20% of tumor-bearingNos2-/-mice, and these animals were resistant to tumor rechallenge. Th1 cytokines were elevated in the blood of treated mice and intratumoral CD4+and CD8+T cells were higher in mice that received indomethacin when compared to control untreated mice. Multiplex immunofluorescence imaging confirmed our phenotyping results and demonstrated that targeted Nos2/Cox2 blockade improved CD8+T cell penetration into the 4T1 tumor core. These findings are consistent with our observations in low NOS2/COX2 expressing breast tumors` proving that COX2 activity is responsible for limiting the spatial distribution of effector T cells in TNBC. Together these results suggest that clinically available NSAID’s may provide a cost-effective, novel immunotherapeutic approach for treatment of aggressive tumors including triple negative breast cancer.

The role of lymph node involvement and tumor size in metastatic disease including breast cancer is unclear. Here, nodal metastasis and T stage on the risk of mortality were investigated in de novo metastatic breast cancer population (35812 patients) in the Surveillance, Epidemiology, and End Results (SEER) Program database in the United States. We found an association between all-cause mortality and regional node involvement (adjusted hazard ratio [HR] = 1.45, 95% confidence interval [CI] 1.36–1.55, p p p +/HER2–), HR+/HER2+, HR–/HER2+ and triple-negative metastatic breast cancer subtypes. Thus, nodal involvement and T stage are independent risk factors for mortality in the population of de novo metastatic breast cancer.

Rationale: Despite longstanding national recommendations for lung cancer screening by low-dose computed tomography (LDCT), annual participation rates are below 15%. Cost, access, and uncertainty over individual-level benefits and harms of screening preclude greater uptake. The development of a low-cost initial blood test that detects lung cancer with high sensitivity could boost LDCT screening rates and improve screening efficiency, by identifying within the screening-eligible population those at relatively higher risk of lung cancer. DELFI (DNA evaluation of fragments for early interception) is a technology that uses low-coverage, whole-genome sequencing and machine learning to detect cancer signals in the blood. In an early study, DELFI demonstrated high sensitivity to detect stage I/II lung cancers [PMID 34417454]. CASCADE-LUNG (NCT05306288) is an ongoing study to clinically validate a DELFI-based test to detect lung cancers that would be found by chest LDCT. Study Design and Methods: CASCADE-LUNG is an event-driven, multisite, prospective, cross-sectional, observational, blood specimen collection study. Eligible individuals are ≥50 years old with ≥20 pack-years history of smoking who are scheduled for a first-time or annual lung cancer screening by chest CT scan within ~30 days after enrollment. Individuals are excluded for any history of hematologic malignancy or organ transplantation, prior cancer diagnosis or prior cancer treatment within 2 years before enrollment, and blood transfusion within 120 days before enrollment. Blood samples are collected within 30 days after enrollment for DELFI analysis. Medical record reviews are conducted at 4 months and 12 months post-enrollment, during which medical history (including social and family history and cancer screening history), demographics, imaging reports, diagnostic reports, surgery reports, pathology reports, and other diagnostic information are collected. Objectives are to evaluate the sensitivity and specificity of a DELFI-based test to detect lung cancer (primary), including within clinically relevant subgroups such as stage, nodule size, age, and race/ethnicity (secondary). Clinical endpoints include the presence or absence of histopathologically confirmed lung cancer diagnosed between the time of the enrollment chest CT scan and the 4-month follow-up visit (primary) and the 12-month follow-up visit (secondary), and adverse events associated with blood specimen collection (secondary). About 15,000 participants are expected to enroll to reach ~70 lung cancers that are suitable for analysis. Enrollment began in April 2022 and is expected to close in 2023. Citation Format: Julie A. Barta, Stephen Freedland, Peter J. Mazzone, Amy E. Isaacson, Alisa M. Hewitt, Debbie Jakubowski, Peter B. Bach, Victor E. Velculescu. CASCADE-LUNG: validation of a blood-based assay that evaluates cell-free DNA fragmentation patterns to detect lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT068.

Data sharing statement: PHOSP-COVID: The protocol, consent form, definition and derivation of clinical characteristics and outcomes, training materials, regulatory documents, requests for data access and other relevant study materials are available online at https://www.phosp.org. COVIDENCE UK: De-identified participant data will be made available upon reasonable request to the corresponding author. Supplementary data is available online at: https://www.sciencedirect.com/science/article/pii/S2666776223000546?via%3Dihub#appsec1 . Copyright © 2023 The Author(s). Background: The risk factors for recovery from COVID-19 dyspnoea are poorly understood. We investigated determinants of recovery from dyspnoea in adults with COVID-19 and compared these to determinants of recovery from non-COVID-19 dyspnoea. Methods: We used data from two prospective cohort studies: PHOSP-COVID (patients hospitalised between March 2020 and April 2021 with COVID-19) and COVIDENCE UK (community cohort studied over the same time period). PHOSP-COVID data were collected during hospitalisation and at 5-month and 1-year follow-up visits. COVIDENCE UK data were obtained through baseline and monthly online questionnaires. Dyspnoea was measured in both cohorts with the Medical Research Council Dyspnoea Scale. We used multivariable logistic regression to identify determinants associated with a reduction in dyspnoea between 5-month and 1-year follow-up. Findings: We included 990 PHOSP-COVID and 3309 COVIDENCE UK participants. We observed higher odds of improvement between 5-month and 1-year follow-up among PHOSP-COVID participants who were younger (odds ratio 1.02 per year, 95% CI 1.01–1.03), male (1.54, 1.16–2.04), neither obese nor severely obese (1.82, 1.06–3.13 and 4.19, 2.14–8.19, respectively), had no pre-existing anxiety or depression (1.56, 1.09–2.22) or cardiovascular disease (1.33, 1.00–1.79), and shorter hospital admission (1.01 per day, 1.00–1.02). Similar associations were found in those recovering from non-COVID-19 dyspnoea, excluding age (and length of hospital admission). Interpretation: Factors associated with dyspnoea recovery at 1-year post-discharge among patients hospitalised with COVID-19 were similar to those among community controls without COVID-19. Funding: PHOSP-COVID is supported by a grant from the MRC-UK Research and Innovation and the Department of Health and Social Care through the National Institute for Health Research (NIHR) rapid response panel to tackle COVID-19. The views expressed in the publication are those of the author(s) and not necessarily those of the National Health Service (NHS), the NIHR or the Department of Health and Social Care. COVIDENCE UK is supported by the UK Research and Innovation, the National Institute for Health Research, and Barts Charity. The views expressed are those of the authors and not necessarily those of the funders. PHOSP-COVID: This study would not be possible without all the participants who have given their time and support. We thank all the participants and their families. We thank the many research administrators, health-care and social-care professionals who contributed to setting up and delivering the study at all of the 65 NHS trusts/Health boards and 25 research institutions across the UK, as well as all the supporting staff at the NIHR Clinical Research Network, Health Research Authority, Research Ethics Committee, Department of Health and Social Care, Public Health Scotland, and Public Health England, and support from the ISARIC Coronavirus Clinical Characterisation Consortium. We thank Kate Holmes at the NIHR Office for Clinical Research Infrastructure (NOCRI) for her support in coordinating the charities group. The PHOSP-COVID industry framework was formed to provide advice and support in commercial discussions, and we thank the Association of the British Pharmaceutical Industry as well NOCRI for coordinating this. We are very grateful to all the charities that have provided insight to the study: Action Pulmonary Fibrosis, Alzheimer's Research UK, Asthma + Lung UK, British Heart Foundation, Diabetes UK, Cystic Fibrosis Trust, Kidney Research UK, MQ Mental Health, Muscular Dystrophy UK, Stroke Association Blood Cancer UK, McPin Foundations, and Versus Arthritis. We thank the NIHR Leicester Biomedical Research Centre patient and public involvement group and Long Covid Support. COVIDENCE UK: We thank all participants of COVIDENCE UK, and the following organisations who supported study recruitment: Asthma UK/British Lung Foundation, the British Heart Foundation, the British Obesity Society, Cancer Research UK, Diabetes UK, Future Publishing, Kidney Care UK, Kidney Wales, Mumsnet, the National Kidney Federation, the National Rheumatoid Arthritis Society, the North West London Health Research Register (DISCOVER), Primary Immunodeficiency UK, the Race Equality Foundation, SWM Health, the Terence Higgins Trust, and Vasculitis UK.

Dysregulation in neutrophil extracellular trap (NET) formation and degradation may play a role in the pathogenesis and severity of COVID-19; however, its role in the pediatric manifestations of this disease including MIS-C and chilblain-like lesions (CLL), otherwise known as “COVID toes”, remains unclear. Studying multinational cohorts, we found that, in CLL, NETs were significantly increased in serum and skin. There was geographic variability in the prevalence of increased NETs in MIS-C, in association with disease severity. MIS-C and CLL serum samples displayed decreased NET degradation ability, in association with C1q and G-actin or anti-NET antibodies, respectively, but not with genetic variants of DNases. In adult COVID-19, persistent elevations in NETs post-disease diagnosis were detected but did not occur in asymptomatic infection. COVID-19-affected adults displayed significant prevalence of impaired NET degradation, in association with anti-DNase1L3, G-actin, and specific disease manifestations, but not with genetic variants of DNases. NETs were detected in many organs of adult patients who died from COVID-19 complications. Infection with the Omicron variant was associated with decreased levels of NETs when compared to other SARS-CoV-2 strains. These data support a role for NETs in the pathogenesis and severity of COVID-19 in pediatric and adult patients.SummaryNET formation and degradation are dysregulated in pediatric and symptomatic adult patients with various complications of COVID-19, in association with disease severity. NET degradation impairments are multifactorial and associated with natural inhibitors of DNase 1, G-actin and anti-DNase1L3 and anti-NET antibodies. Infection with the Omicron variant is associated with decreased levels of NETs when compared to other SARS-CoV-2 strains.

Fast radio bursts (FRBs) are exceptionally luminous flashes of unknown physical origin, reaching us from other galaxies (Petroff et al. 2019). Most FRBs have only ever been seen once, while others flash repeatedly, though sporadically (Spitler et al. 2016, CHIME/FRB Collaboration et al. 2021). Many models invoke magnetically powered neutron stars (magnetars) as the engines producing FRB emission (Margalit & Metzger 2018, CHIME/FRB Collaboration et al. 2020). Recently, CHIME/FRB announced the discovery (Bhardwaj et al. 2021) of the repeating FRB 20200120E, coming from the direction of the nearby grand design spiral galaxy M81. Four potential counterparts at other observing wavelengths were identified (Bhardwaj et al. 2021) but no definitive association with these sources, or M81, could be made. Here we report an extremely precise localisation of FRB 20200120E, which allows us to associate it with a globular cluster (GC) in the M81 galactic system and to place it ~2pc offset from the optical center of light of the GC. This confirms (Bhardwaj et al. 2021) that FRB 20200120E is 40 times closer than any other known extragalactic FRB. Because such GCs host old stellar populations, this association strongly challenges FRB models that invoke young magnetars formed in a core-collapse supernova as powering FRB emission. We propose, instead, that FRB 20200120E is a highly magnetised neutron star formed via either accretion-induced collapse of a white dwarf or via merger of compact stars in a binary system (Margalit et al. 2019). Alternative scenarios involving compact binary systems, efficiently formed inside globular clusters, could also be responsible for the observed bursts., Submitted. Comments welcome

Nonalcoholic steatohepatitis (NASH) is the most common chronic liver disease in the US, partly due to the increasing incidence of metabolic syndrome, obesity, and type 2 diabetes. The roles of bile acids and their receptors, such as the nuclear receptor farnesoid X receptor (FXR) and the G protein-coupled receptor TGR5, on the development of NASH are not fully clear. C57BL/6J male mice fed a Western diet (WD) develop characteristics of NASH, allowing determination of the effects of FXR and TGR5 agonists on this disease. Here we show that the FXR-TGR5 dual agonist INT-767 prevents progression of WD-induced hepatic steatosis, inflammation, and fibrosis, as determined by histological and biochemical assays and novel label-free microscopy imaging techniques, including third harmonic generation, second harmonic generation, and fluorescence lifetime imaging microscopy. Furthermore, we show INT-767 decreases liver fatty acid synthesis and fatty acid and cholesterol uptake, as well as liver inflammation. INT-767 markedly changed bile acid composition in the liver and intestine, leading to notable decreases in the hydrophobicity index of bile acids, known to limit cholesterol and lipid absorption. In addition, INT-767 upregulated expression of liver p-AMPK, SIRT1, PGC-1α, and SIRT3, which are master regulators of mitochondrial function. Finally, we found INT-767 treatment reduced WD-induced dysbiosis of gut microbiota. Interestingly, the effects of INT-767 in attenuating NASH were absent in FXR-null mice, but still present in TGR5-null mice. Our findings support treatment and prevention protocols with the dual FXR-TGR5 agonist INT-767 arrest progression of WD-induced NASH in mice mediated by FXR-dependent, TGR5-independent mechanisms.

Background\ud\udThe quality of dermatology consultations is partly determined by how clinicians approach patient care. The term ‘Personal Models’ describes the explanatory frameworks of thoughts, feelings and experiences that drive behaviour. One study found that clinicians’ personal models, specifically their beliefs about autonomy and patient self-management, influenced the degree to which clinicians engage patients in shared decision-making during consultations. Further research is needed to further explore how clinicians’ personal models inform and affect the quality of patient care.\udAims & objectives\ud\udTo explore how clinicians’ personal models inform shared decision-making and consultation style in managing people living with psoriasis in the context of a new treatment, Apremilast.\udMethods\ud\udA Framework Analysis of qualitative semi-structured telephone interviews with 13 dermatologists from the UK and Germany who participated in a novel medicine trial for psoriasis called APPRECIATE.\udResults\ud\udTwo themes were derived from the data. Theme one, personal working models of patient care, comprised of two sub-themes: (1.1) patient-centeredness: a continuum; and (1.2) stereotypes and assumptions. Theme two, impact of personal working models on patient care, included three sub-themes: (2.1) shared decision-making: a continuum; (2.2) consultation skills; and (2.3) impact of concerns about Apremilast on prescribing behaviour.\udConclusions\ud\udAlthough many dermatologists endorsed a patient-centred approach, not all reported working in this way. Clinicians’ personal models, their beliefs, stereotypes, personal perceptions, and assumptions about patients, are likely to affect their prescribing behaviour and shared decision-making. Additional specialised training and education could increase patient-centeredness and whole person management.

HCC is a highly aggressive and heterogeneous cancer type with limited treatment options. Identifying drivers of tumor heterogeneity may lead to better therapeutic options and favorable patient outcomes. We investigated whether apoptotic cell death and its spatial architecture is linked to tumor molecular heterogeneity using single-cell in situ hybridization analysis.We analyzed 254 tumor samples from two HCC cohorts using tissue microarrays. We developed a mathematical model to quantify cellular diversity among HCC samples using two tumor markers, cyclin-dependent kinase inhibitor 3 and protein regulator of cytokinesis 1 as surrogates for heterogeneity and caspase 3 (CASP3) as an apoptotic cell death marker. We further explored the impact of potential dying-cell hubs on tumor cell diversity and patient outcome by density contour mapping and spatial proximity analysis. We also developed a selectively controlled in vitro model of cell death using CRISPR/CRISPR-associated 9 to determine therapy response and growth under hypoxic conditions. We found that increasing levels of CASP3These results are consistent with the hypothesis that increased apoptotic cell death may lead to greater tumor heterogeneity and thus worse patient outcomes.

We present 849 new bursts from FRB 20121102A detected with the 305-m Arecibo Telescope. Observations were conducted as part of our regular campaign to monitor activity and evolution of burst properties. The 10 reported observations were carried out between 1150 and 1730 MHz and fall in the active period around November 2018. All bursts were dedispersed at the same dispersion measure and are consistent with a single value of 562.4(1) pc/cm$^3$. The rate varies between 0 bursts and 218(16) bursts per hour, the highest rate observed to date. The times between consecutive bursts show a bimodal distribution. We find that a Poisson process with varying rate best describes arrival times with separations >0.1 s. Clustering on timescales of 22 ms reflects a characteristic timescale of the source and possibly the emission mechanism. We analyse the spectro-temporal structure of the bursts by fitting 2D Gaussians with a temporal drift to each sub-burst in the dynamic spectra. We find a linear relationship between the sub-burst's drift and its duration. At the same time, the drifts are consistent with coming from the sad-trombone effect. This has not been predicted by current models. The energy distribution shows an excess of high energy bursts and is insufficiently modelled by a single power-law even within single observations. We find long-term changes in the energy distribution, the average spectrum, and the sad-trombone drift, compared to earlier and later published observations. Despite the large burst rate, we find no strict short-term periodicity., 22 pages, 15 figures, published in MNRAS. Data available at https://doi.org/10.1093/mnras/stac3446

BackgroundOncolytic immunotherapy represents a unique therapeutic platform for the treatment of cancer. Here, we evaluated the safety and efficacy of the combination of pexastimogene devacirepvec (PexaVec) plus durvalumab (anti-programmed death ligand 1) with and without tremelimumab (anti-cytotoxic T-lymphocyte associated protein 4) in patients with standard chemotherapy refractory mismatch repair proficient (pMMR) metastatic colorectal cancer (mCRC) in a phase I/II trial.MethodsAdult patients with histologically confirmed advanced pMMR mCRC, who had progressed on at least two prior lines of systemic chemotherapy were studied in four cohorts. Patients received four doses of PexaVec IV at a dose of 3×108plaque forming units (pfu) (dose level 1) or 1×109 pfu (dose level 2) every 2 weeks. Twelve days after the first PexaVec administration, patients received either 1500 mg of durvalumab every 28 days alone or an additional single dose of 300 mg tremelimumab on day 1. Responses were assessed every 8 weeks by CT or MRI. AEs were recorded. The primary endpoints were safety and feasibility. Secondary endpoints included progression-free survival (PFS) and overall survival. Paired tumor samples and peripheral blood were collected to perform immune monitoring.ResultsThirty-four patients with mCRC enrolled on to the study: 16 patients in the PexaVec/durvalumab cohorts and 18 patients in the PexaVec/durvalumab/tremelimumab cohorts. Overall, the combination of PexaVec plus immune checkpoint inhibitors did not result in any unexpected toxicities. Most common toxicities observed were fever and chills after PexaVec infusion. Two cases of grade 3 colitis, one case of a grade 2 myositis and one case of grade 3 hypotension resulted in discontinuation of immune checkpoint inhibitor and PexaVec treatment, respectively. The median PFS in the PexaVec/durvalumab/tremelimumab cohorts was 2.3 months (95% CI: 2.2 to 3.2 months) vs 2.1 months (95% CI: 1.7 to 2.8 months; p=0.57) in the PexaVec/durvalumab cohorts. Flow cytometry analysis of peripheral blood mononuclear cells revealed an increase in Ki67+CD8+T cells on treatment.ConclusionPexaVec in combination with durvalumab and tremelimumab is safe and tolerable. No unexpected toxicities were observed. The combination of PexaVec/durvalumab/tremelimumab demonstrated potential clinical activity in patients with pMMR mCRC, but further studies are needed to identify the predictive biomarkers.Trial registration numberNCT03206073.

Medical image interpretation is central to detecting, diagnosing, and staging cancer and many other disorders. At a time when medical imaging is being transformed by digital technologies and artificial intelligence, understanding the basic perceptual and cognitive processes underlying medical image interpretation is vital for increasing diagnosticians’ accuracy and performance, improving patient outcomes, and reducing diagnostician burnout. Medical image perception remains substantially understudied. In September 2019, the National Cancer Institute convened a multidisciplinary panel of radiologists and pathologists together with researchers working in medical image perception and adjacent fields of cognition and perception for the “Cognition and Medical Image Perception Think Tank.” The Think Tank’s key objectives were to identify critical unsolved problems related to visual perception in pathology and radiology from the perspective of diagnosticians, discuss how these clinically relevant questions could be addressed through cognitive and perception research, identify barriers and solutions for transdisciplinary collaborations, define ways to elevate the profile of cognition and perception research within the medical image community, determine the greatest needs to advance medical image perception, and outline future goals and strategies to evaluate progress. The Think Tank emphasized diagnosticians’ perspectives as the crucial starting point for medical image perception research, with diagnosticians describing their interpretation process and identifying perceptual and cognitive problems that arise. This article reports the deliberations of the Think Tank participants to address these objectives and highlight opportunities to expand research on medical image perception.

While serum-circulating complement destroys invading pathogens, intracellularly active complement, termed the ‘complosome’, functions as a vital orchestrator of cell-metabolic events underlying T cell effector responses. Whether intracellular complement is also non-redundant for the activity of myeloid immune cells is currently unknown. Here, we show that monocytes and macrophages constitutively express complement component (C) 5 and generate autocrine C5a via formation of an intracellular C5 convertase. Cholesterol crystal-sensing by macrophages induced C5aR1 signaling on mitochondrial membranes, which shifted ATP production via reverse electron chain flux towards reactive oxygen species (ROS) generation and anaerobic glycolysis to favor IL-1β production, both at the transcriptional level and processing of pro-IL-1β. Consequently, atherosclerosis-prone mice lacking macrophage-specific C5ar1 had ameliorated cardiovascular disease on a high-cholesterol diet. Conversely, inflammatory gene signatures and IL-1β produced by cells in unstable atherosclerotic plaques of patients were normalized by a specific cell-permeable C5aR1 antagonist. Deficiency of the macrophage cell autonomous C5 system also protected mice from crystal nephropathy mediated by folic acid. These data demonstrate the unexpected intracellular formation of a C5 convertase and identify C5aR1 as a direct modulator of mitochondrial function and inflammatory output from myeloid cells. Together, these findings suggest that the complosome is a contributor to the biologic processes underlying sterile inflammation and indicate that targeting this system could be beneficial in macrophage-dependent diseases, such as atherosclerosis.

Nanobodies offer several potential advantages over mAbs for the control of SARS-CoV-2. Their ability to access cryptic epitopes conserved across SARS-CoV-2 variants of concern (VoCs) and feasibility to engineer modular, multimeric designs, make these antibody fragments ideal candidates for developing broad-spectrum therapeutics against current and continually emerging SARS-CoV-2 VoCs. Here we describe a diverse collection of 37 anti-SARS-CoV-2 spike glycoprotein nanobodies extensively characterized as both monovalent and IgG Fc-fused bivalent modalities. The panel of nanobodies were shown to have high intrinsic affinity; high thermal, thermodynamic and aerosolization stability; broad subunit/domain specificity and cross-reactivity across many VoCs; wide-ranging epitopic and mechanistic diversity; high and broad in vitro neutralization potencies; and high neutralization efficacies in hamster models of SARS-CoV-2 infection, reducing viral burden by up to six orders of magnitude to below detectable levels. In vivo protection was demonstrated with anti-RBD and previously unreported anti-NTD and anti-S2 nanobodies. This collection of nanobodies provides a therapeutic toolbox from which various cocktails or multi-paratopic formats could be built to tackle current and future SARS-CoV-2 variants and SARS-related viruses. Furthermore, the high aerosol-ability of nanobodies provides the option for effective needle-free delivery through inhalation.

Coronavirus disease 2019 (COVID-19) is known to cause multi-organ dysfunction

Background: Some of the obstacles to successful entrepreneurship in emerging economies are embedded in the internal operations and management of enterprises, especially women mid-scale enterprises. This includes the lack of effective strategies that promote the growth of women-owned mid-scale enterprises and their inclination towards necessity-based entrepreneurship, which greatly limits their growth potential. Aim: The study explored the experience and know-how of strategies applied by three high-profiled women-owned mid-scale enterprises in East Africa to stimulate their business growth. Methods: A qualitative research approach, using perspective seeking methods by applying unrestrictive semi-structured questioning was adopted. Results: Findings suggest that women must embrace, recognise and own their internal aspirations and ambition to grow their mid-scale enterprise. Conclusion: Women-owned mid-scale enterprises showed the need to map out their strategic growth plan and it must be built on a solid foundation to justify the need to grow for all stakeholders to buy into it.

A 26-year-old otherwise healthy man died of fulminant myocarditis. Nasopharyngeal specimens collected premortem tested negative for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Histopathological evaluation of the heart showed myocardial necrosis surrounded by cytotoxic T-cells and tissue-repair macrophages. Myocardial T-cell receptor (TCR) sequencing revealed hyper-dominant clones with highly similar sequences to TCRs that are specific for SARS-CoV-2 epitopes. SARS-CoV-2 RNA was detected in the gut, supporting a diagnosis of multisystem inflammatory syndrome in adults (MIS-A). Molecular targets of MIS-associated inflammation are not known. Our data indicate that SARS-CoV-2 antigens selected high-frequency T-cell clones that mediated fatal myocarditis.

Tissues and organs are composed of distinct cell types that must operate in concert to perform physiological functions. Efforts to create high-dimensional biomarker catalogs of these cells are largely based on transcriptomic single-cell approaches that lack the spatial context required to understand critical cellular communication and correlated structural organization. To probe in situ biology with sufficient coverage depth, several multiplexed protein imaging methods have recently been developed. Though these antibody-based technologies differ in strategy and mode of immunolabeling and detection tags, they commonly utilize antibodies directed against protein biomarkers to provide detailed spatial and functional maps of complex tissues. As these promising antibody-based multiplexing approaches become more widely adopted, new frameworks and considerations are critical for training future users, generating molecular tools, validating antibody panels, and harmonizing datasets. In this perspective, we provide essential resources and key considerations for obtaining robust and reproducible multiplexed antibody-based imaging data compiling specialized knowledge from domain experts and technology developers., Comment: {\ddag}authors contributed equally to this work

Very long baseline interferometric (VLBI) localisations of repeating fast radio bursts (FRBs) have demonstrated a diversity of local environments: from nearby star-forming regions to globular clusters. Here we report the VLBI localisation of FRB 20201124A using an ad-hoc array of dishes that also participate in the European VLBI Network (EVN). In our campaign, we detected 18 total bursts from FRB 20201124A at two separate epochs. By combining the visibilities from both observing epochs, we were able to localise FRB 20201124A with a 1-$\sigma$ error of 4.5 milliarcseconds (mas). We use the relatively large burst sample to investigate astrometric accuracy, and find that for $\gtrsim20$ baselines ($\gtrsim7$ dishes) that we can robustly reach milliarcsecond precision even using single-burst data sets. Sub-arcsecond precision is still possible for single bursts, even when only $\sim$ six baselines (four dishes) are available. We explore two methods for determining the individual burst positions: the peaks of the dirty maps and a Gaussian fit to the cross fringe pattern on the dirty maps. We found the latter to be more reliable due to the lower mean and standard deviation in the offsets from the FRB position. Our VLBI work places FRB 20201124A 705$\pm$26 mas (1-$\sigma$ errors) from the optical centre of the host galaxy, and consistent with originating from within the recently-discovered extended radio structure associated with star-formation in the host galaxy. Future high-resolution optical observations, e.g. with Hubble Space Telescope, can determine the proximity of our FRB 20201124A VLBI position to nearby knots of star formation., Comment: submitted, comments welcome

Multiplexed ion beam imaging by time-of-flight (MIBI-TOF) is a form of mass spectrometry imaging that uses metal labeled antibodies and secondary ion mass spectrometry to image dozens of proteins simultaneously in the same tissue section. Working with the National Cancer Institute's (NCI) Cancer Immune Monitoring and Analysis Centers (CIMAC), we undertook a validation study, assessing concordance across a dozen serial sections of a tissue microarray of 21 samples that were independently processed and imaged by MIBI-TOF or single-plex immunohistochemistry (IHC) over 12 days. Pixel-level features were highly concordant across all 16 targets assessed in both staining intensity (R

Cancer immunotherapy has emerged as a promising cancer treatment. However, the presence of immune-refractory tumor cells limits its clinical success by blocking amplification of anti-tumor immunity. Previously, we found that immune selection by immunotherapy drives the evolution of tumors toward multi-modal resistant and stem-like phenotypes via transcription induction of AKT co-activator TCL1A by NANOG. Here, we report a crucial role of HSP90A at the crossroads between NANOG-TCL1A axis and multi-aggressive properties of immune-edited tumor cells by identifying HSP90AA1 as a NANOG transcriptional target. Furthermore, we demonstrate that HSP90A potentiates AKT activation through TCL1A-stabilization, thereby contributing to the multi-aggressive properties in NANOGhigh tumor cells. Importantly, HSP90 inhibition sensitized immune-refractory tumor to adoptive T cell transfer as well as PD-1 blockade, and re-invigorated the immune cycle of tumor-reactive T cells. Our findings implicate that the HSP90A-TCL1A-AKT pathway ignited by NANOG is a central molecular axis and a potential target for immune-refractory tumor., Nanog can confer resistance to cancer immunotherapy by promoting AKT activity. Here, the authors demonstrate that HSP90A is a Nanog target that stabilizes the AKT coactivator TCL1, thereby activating AKT, and that HSP90A inhibition can enhance the anti-tumor efficacy of adoptive T cell transfer and checkpoint blockade.

Cutaneous malignant melanoma is an aggressive cancer of melanocytes with a strong propensity to metastasize. We posit that melanoma cells acquire metastatic capability by adopting an embryonic-like phenotype, and that a lineage approach would uncover metastatic melanoma biology. Using a genetically engineered mouse model to generate a rich melanoblast transcriptome dataset, we identify melanoblast-specific genes whose expression contribute to metastatic competence and derive a 43-gene signature that predicts patient survival. We identify a melanoblast gene, KDELR3, whose loss impairs experimental metastasis. In contrast, KDELR1 deficiency enhances metastasis, providing the first example of different disease etiologies within the KDELR-family of retrograde transporters. We show that KDELR3 regulates the metastasis suppressor, KAI1, and report an interaction with the E3 ubiquitin-protein ligase gp78, a regulator of KAI1 degradation. Our work demonstrates that the melanoblast transcriptome can be mined to uncover targetable pathways for melanoma therapy., Metastatic cells can mimic many of the phenotypic behaviors of embryonic cells. Here, the authors generate a melanoblast-specific transcriptome using a genetically engineered mouse model and identify KDELR3 as a pro-metastasis gene in melanoma.

Delayed terminal duct lobular unit (TDLU) involution is associated with elevated mammographic breast density (MD). Both are independent breast cancer risk factors among women with benign breast disease (BBD). Prior digital analyses of normal breast tissues revealed that epithelial nuclear density (END) and TDLU involution are inversely correlated. Accordingly, we examined associations of END, TDLU involution, and MD in BBD clinical biopsies. This study included digitized images of 262 representative image-guided hematoxylin and eosin–stained biopsies from 224 women diagnosed with BBD, enrolled within the cross-sectional BREAST-Stamp project that were visually assessed for TDLU involution (TDLU count/100 mm2, median TDLU span and median acini count per TDLU). A digital algorithm estimated nuclei count per unit epithelial area, or END. Single X-ray absorptiometry of prebiopsy ipsilateral craniocaudal digital mammograms measured global and localized MD surrounding the biopsy region. Adjusted ordinal logistic regression models assessed relationships between tertiles of TDLU and END measures. Analysis of covariance examined mean differences in MD across END tertiles. TDLU measures were positively associated with increasing END tertiles [TDLU count/100 mm2, ORT3vsT1: 3.42, 95% confidence interval (CI), 1.87–6.28; acini count/TDLUT3vsT1, OR: 2.40, 95% CI, 1.39–4.15]. END was significantly associated with localized, but not, global MD. Relationships were most apparent among patients with nonproliferative BBD. These findings suggest that quantitative END reflects different but complementary information to the histologic information captured by visual TDLU and radiologic MD measures and merits continued evaluation in assessing cellularity of breast parenchyma to understand the etiology of BBD.

Objective: To determine if a weight gain prevention intervention is acceptable to young women with a normal Body Mass Index and a moderately increased or high risk of breast cancer. Design: Qualitative semi-structured interview study involving 14 women aged 26–35 years who were registered with a Family History Clinic in Manchester, UK, due to family history of breast cancer. Participants’ views were analysed thematically. Results: Four themes were produced: (1) perceptions of a healthy lifestyle: women’s perceptions included health-related behaviours and subjective wellbeing; (2) construing a healthy weight: women rely on appearance, feelings and others opinions to construe weight instead of quantitative indicators; (3) configuring a useful programme: the idea of a programme that is remotely accessible; provides a point of contact; and promotes general wellbeing was appealing. Women believed information explaining the link between lifestyle and breast cancer would facilitate behaviour change; (4) the importance of will(power): women recognised that commitment to a programme is affected by time, money and readiness to change. Conclusion: A weight gain prevention intervention that focuses on wellbeing and behaviour change appears acceptable to many healthy weight women. Future research should examine whether women’s expressed acceptability translates into actual acceptability of such a programme.

PURPOSE Genetically engineered T-cell therapy is an emerging treatment of hematologic cancers with potential utility in epithelial cancers. We investigated T-cell therapy for the treatment of metastatic human papillomavirus (HPV)–associated epithelial cancers. METHODS This phase I/II, single-center trial enrolled patients with metastatic HPV16-positive cancer from any primary tumor site who had received prior platinum-based therapy. Treatment consisted of autologous genetically engineered T cells expressing a T-cell receptor directed against HPV16 E6 (E6 T-cell receptor T cells), a conditioning regimen, and systemic aldesleukin. RESULTS Twelve patients were treated in the study. No dose-limiting toxicities were observed in the phase I portion. Two patients, both in the highest-dose cohort, experienced objective tumor responses. A patient with three lung metastases experienced complete regression of one tumor and partial regression of two tumors, which were subsequently resected; she has no evidence of disease 3 years after treatment. All patients demonstrated high levels of peripheral blood engraftment with E6 T-cell receptor T cells 1 month after treatment (median, 30%; range, 4% to 53%). One patient’s resistant tumor demonstrated a frameshift deletion in interferon gamma receptor 1, which mediates response to interferon gamma, an essential molecule for T-cell–mediated antitumor activity. Another patient’s resistant tumor demonstrated loss of HLA-A*02:01, the antigen presentation molecule required for this therapy. A tumor from a patient who responded to treatment did not demonstrate genetic defects in interferon gamma response or antigen presentation. CONCLUSION Engineered T cells can induce regression of epithelial cancer. Tumor resistance was observed in the context of T-cell programmed death-1 expression and defects in interferon gamma and antigen presentation pathway components. These findings have important implications for development of cellular therapy in epithelial cancers.

PURPOSE: Forkhead box protein O1 (FOXO1) and paired box gene 3 (PAX3) have been reported to play an imported role in human cancers, but their role in cervical cancer has not yet been clarified. In this study, we evaluated the functional role of FOXO1 in cervical cancer cells and investigated the expression and clinical significance of FOXO1 and PAX3 in cervical lesions. METHODS: In vitro assessment of cell function by cell viability, migration, and invasion assays were performed on FOXO1-knockdown cervical cancer cells. Immunohistochemical (IHC) staining analyses of FOXO1 and PAX3 were performed with a tissue microarray (TMA). The clinical significance was evaluated by comparing the data with various clinicopathologic characteristics, including survival of patients with cervical cancer. RESULTS: In vitro results revealed that knockdown of FOXO1 is associated with decreased cell viability (p < 0.001), migration (p < 0.001), and invasion (p < 0.05), supporting the oncogenic role of FOXO1 in cervical cancer. FOXO1 and PAX3 expression was significantly higher in CIN (both p < 0.001) and cancer tissue (both p < 0.001) than in normal tissue. Multivariate analysis indicated that FOXO1 expression (hazard ratio 4.01 [95% CI 1.22–13.10], p = 0.021) and an advanced FIGO stage (hazard ratio 3.89 [95% CI 1.35–11.19], p = 0.012) were independent prognostic factors for overall survival. CONCLUSIONS: This study reveals increased FOXO1 and PAX3 expression in cervical cancers and indicates an oncogenic role of FOXO1 in cervical cancer cells that correlates with poor patient survival.

Insights from a number of INCO cyanide destruction process reviews and studies are presented and discussed. In particular, the performance of three plants are discussed in terms of reagent (sulfite...

The surface mining of oil sands north of Fort McMurray, Alberta produces considerable tailings waste that is stored in large tailings ponds on industrial lease sites. Viable strategies for the detoxification of oil sands process affected water (OSPW) are under investigation. In order to assess the toxic potential of the suite of dissolved organics in OSPW, a method for their extraction and fractionation was developed using solid phase extraction. The method successfully isolated organic compounds from 180 L of an aged OSPW source. Using acidic- or alkaline-conditioned non-polar ENV+ resin and soxhlet extraction with ethyl acetate and methanol, three fractions (F1–F3) were generated. Chemical characterization of the generated fractions included infusion to electrospray ionization ultrahigh-resolution mass spectrometry (ESI-UHRMS), liquid chromatography quadrupole time-of-flight mass spectrometry, gas chromatography triple quadrupole time-of-flight mass spectrometry, and synchronous fluorescence spectroscopy (SFS). Additionally, ESI-UHRMS class distribution data and SFS identified an increased degree of oxygenation and aromaticity, associated with increased polarity. Method validation, which included method and matrix spikes with surrogate and labelled organic mono carboxylic acid standards, confirmed separation according to acidity and polarity with generally good recoveries (average 76%). Because this method is capable of extracting large sample volumes, it is amenable to thorough chemical characterization and toxicological assessments with a suite of bioassays. As such, this protocol will facilitate effects-directed analysis of toxic components within bitumen-influenced waters from a variety of sources.

RNA has attracted considerable attention as a target for small molecules. However, methods to identify, study, and characterize suitable RNA targets have lagged behind strategies for protein targets. One approach that has received considerable attention for protein targets has been to utilize computational analysis to investigate ligandable “pockets” on proteins that are amenable to small molecule binding. These studies have shown that selected physical properties of pockets are important parameters that govern the ability of a structure to bind to small molecules. This work describes a similar analysis to study pockets on all RNAs in the Protein Data Bank (PDB). Using parameters such as buriedness, hydrophobicity, volume, and other properties, the set of all RNAs is analyzed and compared to all proteins. Considerable overlap is observed between the properties of pockets on RNAs and proteins. Thus, many RNAs are capable of populating conformations with pockets that are likely suitable for small molecule binding. Further, principal moment of inertia (PMI) calculations reveal that liganded RNAs exist in diverse structural space, much of which overlaps with protein structural space. Taken together, these results suggest that complex folded RNAs adopt unique structures with pockets that may represent viable opportunities for small molecule targeting.

The process of surface mining and extracting bitumen from oil sand produces large quantities of tailings and oil sands process-affected water (OSPW). The industry is currently storing OSPW on-site while investigating strategies for their detoxification. One such strategy relies on the biodegradation of organic compounds by indigenous microbes, resulting in aged tailings waters with reduced toxicity. This study assessed the toxicity of OSPW aged statically for approximately 18 years. Dissolved organics in aged OSPW were fractionated using a preparative solid-phase extraction method that generated three organic fractions (F1–F3) of increasing polarity. Eight aquatic species from different trophic levels were exposed to whole OSPW (WW) and the derived OSPW organic fractions to assess toxicity: Pimephales promelas, Oryzias latipes, Vibrio fischeri, Daphnia magna, Lampsilis cardium, Hyalella azteca, Ceriodaphnia dubia, and Hexagenia spp. Broad comparisons revealed that P. promelas and H. azteca were most sensitive to dissolved organics within aged OSPW, while WW was most toxic to L. cardium and H. azteca. Three cases of possible contaminant interactions occurred within whole OSPW treatments, as toxicity was higher than organic fractions for H. azteca and L. cardium, and lower for P. promelas. As such, the drivers of toxicity appeared to be dependent on the species exposed. Of the organic fractions assessed, F3 (most polar) was the most toxic overall while F2 (intermediate polarity) displayed little toxicity to all species evaluated. This presents strong evidence that classical mono-carboxylic naphthenic acids, mostly present in F1 (least polar), are not primarily responsible for the toxicity in aged tailings. The current study indicates that although the aged tailings source (≥18 years) did not display acute toxicity to the majority of organisms assessed, inorganic components and polyoxygenated organics may pose a persistent concern to some aquatic organisms.

Proteus syndrome is a mosaic, progressive overgrowth disorder caused by a somatic activating variant c.49G > A p.(E17K) in AKT1. The presentation in affected individuals is variable, with a diversity of tissues demonstrating abnormalities. Common manifestations include skin and bony overgrowth, vascular malformations (VMs), cysts and benign tumors. We used two methods to create mouse models that had endogenously-regulated mosaic expression of the Proteus syndrome variant. Variant allele fractions (VAFs) ranged from 0% to 50% across numerous tissues in 44 Proteus syndrome mice. Mice were phenotypically heterogeneous with lesions rarely observed before 12 months of age. VMs were the most frequent finding with a total of 69 found in 29 of 44 Proteus syndrome mice. Twenty-eight cysts and ectasia, frequently biliary, were seen in 22 of 44 Proteus syndrome mice. Varying levels of mammary hyperplasia were seen in 10 of 16 female Proteus syndrome mice with other localized regions of hyperplasia and stromal expansion noted in several additional animals. Interestingly, 27 of 31 Proteus syndrome animals had non-zero blood VAF that is in contrast to the human disorder where it is rarely seen in peripheral blood. Identification of variant-positive cells by green fluorescent protein (GFP) staining in chimeric Proteus syndrome mice showed that in some lesions, hyperplastic cells were predominantly GFP/Akt1E17K-positive and showed increased pAKT signal compared to GFP-negative cells. However, hyperplastic mammary epithelium was a mixture of GFP/Akt1E17K-positive and negative cells with some GFP/Akt1E17K-negative cells also having increased pAKT signal suggesting that the variant-positive cells can induce lesion formation in a non-cell autonomous manner.

Aim: We have shown that IL-4 fused to Pseudomonas exotoxin (IL-4-PE) is cytotoxic to ovarian cancer cell lines. The antineoplastic properties of IFN-α, IFN-γ and IL-4-PE have been studied and showed some promise in the clinical trials. Here, we investigated whether the combination of IL-4-PE, IFN-α and IFN-γ will result in increased ovarian cancer cell death in vitro and in vivo. Materials & Methods: Ovarian cancer cells were tested in vitro to analyze the cytotoxic effects of IL-4-PE, IFN-α and IFN-γ, and the combination of all three. Tumor-bearing xenograft mice were treated with the combination of IL-4-PE, IFN-α and IFN-γ to monitor their overall survival. The JAK/STAT phosphorylation signaling pathways were studied to delineate the mechanism of synergistic antitumor activity. Results: The combination of IL-4-PE with IFN-α and IFN-γ resulted in increased ovarian cancer cell death in vitro and in vivo. Mechanistically, the synergistic antitumor effect was dependent on interferon signaling, but not IL-4-PE signaling as determined by signaling specific chemical inhibitors. The combination therapy induced the activation of critical mediators of apoptosis. Conclusion: The combination of IL-4-PE with interferons increased overall survival of mice with human ovarian cancer xenograft. These data suggest that this novel combination could provide a unique approach to treating ovarian cancer.