The TRAF3-DYRK1A-RAD54L2 complex maintains ACE2 expression to promote SARS-CoV-2 infection.

Angiotensin converting enzyme 2 (ACE2), the host receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, is differentially expressed in a wide variety of tissues and cell types. The expression of ACE2 is under tight regulation, but the mechanisms regulating ACE2 expression have not yet been well defined. Through a genome-wide CRISPR knockout screen, we discovered that host factors TRAF3, DYRK1A, and RAD54L2 (TDR) form a complex to regulate the expression of ACE2. Knockout of TRAF3, DYRK1A, or RAD54L2 reduces the mRNA levels of ACE2 and inhibits the cellular entry of SARS-CoV-2. On the other hand, SARS-CoV-2 continuously evolves by genetic mutations for the adaption to the host. We have identified mutations in spike (S) (P1079T) and nucleocapsid (N) (S194L) that enhance the replication of SARS-CoV-2 in cells that express ACE2 at a low level. Our results have revealed the mechanisms for the transcriptional regulation of ACE2 and the adaption of SARS-CoV-2. IMPORTANCE: The expression of ACE2 is essential for the entry of SARS-CoV-2 into host cells. We identify a new complex-the TDR complex-that acts to maintain the abundance of ACE2 in host cells. The identification and characterization of the TDR complex provide new targets for the development of therapeutics against SARS-CoV-2 infection. By analysis of SARS-CoV-2 virus replicating in cells expressing low levels of ACE2, we identified mutations in spike (P1079T) and nucleocapsid (S194L) that overcome the restriction of limited ACE2. Functional analysis of these key amino acids in S and N extends our knowledge of the impact of SARS-CoV-2 variants on virus infection and transmission.

Effects of N-glycan modifications on spike expression, virus infectivity, and neutralization sensitivity in ancestral compared to Omicron SARS-CoV-2 variants.

The SARS-CoV-2 spike glycoprotein has 22 potential N-linked glycosylation sites per monomer that are highly conserved among diverse variants, but how individual glycans affect virus entry and neutralization of Omicron variants has not been extensively characterized. Here we compared the effects of specific glycan deletions or modifications in the Omicron BA.1 and D614G spikes on spike expression, processing, and incorporation into pseudoviruses, as well as on virus infectivity and neutralization by therapeutic antibodies. We found that loss of potential glycans at spike residues N717 and N801 each conferred a loss of pseudovirus infectivity for Omicron but not for D614G or Delta variants. This decrease in infectivity correlated with decreased spike processing and incorporation into Omicron pseudoviruses. Oligomannose-enriched Omicron pseudoviruses generated in GnTI- cells or in the presence of kifunensine were non-infectious, whereas D614G or Delta pseudoviruses generated under similar conditions remained infectious. Similarly, growth of live (authentic) SARS-CoV-2 in the presence of kifunensine resulted in a greater reduction of titers for the BA.1.1 variant than Delta or D614G variants relative to their respective, untreated controls. Finally, we found that loss of some N-glycans, including N343 and N234, increased the maximum percent neutralization by the class 3 S309 monoclonal antibody against D614G but not BA.1 variants, while these glycan deletions altered the neutralization potency of the class 1 COV2-2196 and Etesevimab monoclonal antibodies without affecting maximum percent neutralization. The maximum neutralization by some antibodies also varied with the glycan composition, with oligomannose-enriched pseudoviruses conferring the highest percent neutralization. These results highlight differences in the interactions between glycans and residues among SARS-CoV-2 variants that can affect spike expression, virus infectivity, and susceptibility of variants to antibody neutralization.

Prognosis and risk of suicide after cancer diagnosis.

INTRODUCTION: Suicide rates are elevated after cancer diagnosis. Existential distress caused by awareness of one's impending death is well-described in patients with cancer. The authors hypothesized that suicide risk is associated with cancer prognosis, and the impact of prognosis on suicide risk is greatest for populations with higher baseline suicide risk. METHODS: The authors identified patients (≥16 years old) with newly diagnosed cancers from 2000 to 2019 in the Surveillance, Epidemiology, and End Results database, representing 27% of US cancers. Multiple primary-standardized mortality ratios (SMR) were used to estimate the relative risk of suicide within 6 months of diagnosis compared to the general US population, adjusted for age, sex, race, and year of follow-up. Suicide rates by 20 most common cancer sites were compared with respective 2-year overall survival rates (i.e., prognosis) using a weighted linear regression model. RESULTS: Among 6,754,704 persons diagnosed with cancer, there were 1610 suicide deaths within 6 months of diagnosis, three times higher than the general population (SMR = 3.1; 95% confidence interval, 3.0-3.3). Suicide risk by cancer site was closely associated with overall prognosis (9.5%/percent survival deficit, R2  = 0.88, p < .0001). The association of prognosis with suicide risk became attenuated over time. For men, the risk of suicide increased by 2.8 suicide deaths per 100,000 person-years (p < .0001) versus 0.3 in women (p < .0001). The risk was also higher for persons ≥60 old and for the White (vs. Black) race. CONCLUSIONS: Poorer prognosis was closely associated with suicide risk early after cancer diagnosis and had a greater effect on populations with higher baseline risks of suicide. This model highlights the need for enhanced psychiatric surveillance and continued research in this patient population.

Integration of just-in-time learning with variational autoencoder for cell culture process monitoring based on Raman spectroscopy.

Protein production in the biopharmaceutical industry necessitates the utilization of multiple analytical techniques and control methodologies to ensure both safety and consistency. To facilitate real-time monitoring and control of cell culture processes, Raman spectroscopy has emerged as a versatile analytical technology. This technique, categorized as a Process Analytical Technology, employs chemometric models to establish correlations between Raman signals and key variables of interest. One notable approach for achieving real-time monitoring is through the application of just-in-time learning (JITL), an industrial soft sensor modeling technique that utilizes Raman signals to estimate process variables promptly. The conventional Raman-based JITL method relies on the K-nearest neighbor (KNN) algorithm with Euclidean distance as the similarity measure. However, it falls short of addressing the impact of data uncertainties. To rectify this limitation, this study endeavors to integrate JITL with a variational autoencoder (VAE). This integration aims to extract dominant Raman features in a nonlinear fashion, which are expressed as multivariate Gaussian distributions. Three experimental runs using different cell lines were chosen to compare the performance of the proposed algorithm with commonly utilized methods in the literature. The findings indicate that the VAE-JITL approach consistently outperforms partial least squares, convolutional neural network, and JITL with KNN similarity measure in accurately predicting key process variables.

Cell culture product quality attribute prediction using convolutional neural networks and Raman spectroscopy.

Advanced process control in the biopharmaceutical industry often lacks real-time measurements due to resource constraints. Raman spectroscopy and Partial Least Squares (PLS) models are often used to monitor bioprocess cultures in real-time. In spite of the ease of training, the accuracy of the PLS model is impacted if it is not used to predict quality attributes for the cell lines it is trained on. To address this issue, a deep convolutional neural network (CNN) is proposed for offline modeling of metabolites using Raman spectroscopy. By utilizing asymmetric least squares smoothing to adjust Raman spectra baselines, a generic training data set is created by amalgamating spectra from various cell lines and operating conditions. This data set, combined with their derivatives, forms a two-dimensional model input. The CNN model is developed and validated for predicting different quality variables against measurements from various continuous and fed-batch experimental runs. Validation results confirm that the deep CNN model is an accurate generic model of the process to predict real-time quality attributes, even in experimental runs not included in the training data. This model is robust and versatile, requiring no recalibration when deployed at different sites to monitor various cell lines and experimental runs.

MGMT promoter methylation in 1p19q-intact gliomas.

OBJECTIVE: Standard-of-care for 1p19q-intact anaplastic gliomas is defined by the international randomized phase III CATNON trial, which found an overall survival (OS) benefit for adjuvant temozolomide (TMZ) when added to radiotherapy. Paradoxically, TMZ did not appear to benefit patients with IDH-wildtype gliomas, regardless of MGMT promoter status. The authors concluded that well-powered prospective study on the clinical efficacy of TMZ for patients with IDH-wildtype anaplastic gliomas (meeting criteria for glioblastoma) is warranted. Given that the prognostic and predictive role of MGMT status for grade 2-3 gliomas is unresolved, we determined the effect of MGMT status on OS in patients with 1p19q-intact gliomas in the National Cancer Database (NCDB). METHODS: We queried the NCDB from 2018 to 2019 for patients with diffuse (grade 2) and anaplastic (grade 3) IDH-wildtype or -mutant astrocytomas who received chemotherapy with follow-up through 2022. The Kaplan-Meier method and Cox proportional hazards regressions models were used to determine the association of MGMT with OS. RESULTS: We identified 1514 patients who were newly diagnosed with IDH-wildtype (n = 802, 33% methylated) or -mutant astrocytomas (n = 712, 48% methylated) and received chemotherapy during initial management. An unmethylated promoter was associated with poorer survival in patients with IDH-wildtype (3-year OS 34% [95%CI 29-39%] vs. 46% [95%CI 39-54%], p < .001, adjusted HR 1.53 [95%CI 1.24-1.89]) but not IDH-mutant astrocytomas (3-year OS 79% [95%CI 74-84%] vs. 80% [95%CI 75-86%], p =0 .81, HR 1.04 [95%CI 0.73-1.50]). CONCLUSIONS: This ancillary analysis supports conclusions from the CATNON trial for adjuvant TMZ as standard-of-care for anaplastic astrocytomas (IDH-mutant and 1p19q-intact), irrespective of MGMT status. Determining the optimal strategy for diffuse gliomas that are IDH-wildtype will be particularly important. MGMT promoter methylation should be considered as a stratification factor in future clinical trials for these patients.

Relationship between glycated hemoglobin levels and three-month outcomes in acute ischemic stroke patients with or without diabetes: a prospective Korean cohort study.

OBJECTIVE: In patients experiencing acute ischemic stroke, there is ongoing debate surrounding the connection between chronic hyperglycemic status and their initial clinical outcomes. Our objective was to examine the connection between glycated hemoglobin (HbA1c) levels and adverse clinical outcomes at both 3-months adverse clinical outcomes in individuals with acute ischemic stroke (AIS) with and without diabetes. METHODS: The present prospective cohort study involved 896 AIS patients without diabetes and 628 with diabetes treated at a South Korean hospital from January 2010 to December 2016. The target independent variable is HbA1c. The outcome variable is a modified Rankin scale score ≥ 3. A binary logistic regression model was applied to assess the connection between HbA1c levels and 3-month poor clinical outcomes in AIS patients with and without diabetes. Additionally, a generalized additive model and smoothed curve fitting were utilized to explore potential nonlinear associations between HbA1c levels and 3-month adverse clinical outcomes in AIS patients with and without diabetes. RESULTS: The binary logistic regression model could not identify any statistically significant connection between HbA1c and 3-month adverse clinical outcomes in AIS patients, both those with and without diabetes, after correcting for various factors. However, a nonlinear relationship emerged between HbA1c and 3-month adverse clinical outcomes in AIS patients with diabetes. The inflection point for HbA1c was determined to be 6.1%. For HbA1c values ≤ 6.1%, an inverse association was observed between HbA1c and 3-month adverse clinical outcomes in diabetic AIS patients, and each 1% increase in HbA1c in AIS patients with DM was associated with an 87% reduction in 3-month adverse clinical outcomes (OR = 0.13, 95% CI: 0.02-0.81). Conversely, when HbA1c exceeded 6.1%, a positive association between HbA1c and 3-month adverse clinical outcomes became apparent in diabetic AIS patients, and each 1% increase in HbA1c in AIS patients with DM was associated with a 23% increase in 3-month adverse clinical outcomes (OR = 1.23, 95%CI: 1.03-1.47). However, it's important to note that no significant linear or nonlinear relationships were observed between HbA1c levels and 3-month adverse clinical outcomes in AIS patients without diabetes. CONCLUSION: Our findings suggest a nonlinear connection and threshold effect between HbA1c and 3-month adverse clinical outcomes in AIS patients with diabetes. AIS patients with diabetes had a lower risk of 3-month adverse clinical outcomes when their HbA1c control was close to 6.1%. Our findings may aid treatment decision-making and potentially guide interventions to optimize glycemic control in AIS patients.

Differences in New Variant of Concern Replication at Physiological Temperatures In Vitro.

Using multiple cell types and isolates of Delta and Omicron variants of SARS-CoV-2, we report differences in virus production, replication, and infectivity in vitro. Ancestral and Delta SARS-CoV-2 variant exhibit reduced virus production and replication at 34°C compared to 37°C while Omicron replication is balanced between temperatures.

Evolution of cisplatin resistance through coordinated metabolic reprogramming of the cellular reductive state.

BACKGROUND: Cisplatin (CDDP) is a mainstay treatment for advanced head and neck squamous cell carcinomas (HNSCC) despite a high frequency of innate and acquired resistance. We hypothesised that tumours acquire CDDP resistance through an enhanced reductive state dependent on metabolic rewiring. METHODS: To validate this model and understand how an adaptive metabolic programme might be imprinted, we performed an integrated analysis of CDDP-resistant HNSCC clones from multiple genomic backgrounds by whole-exome sequencing, RNA-seq, mass spectrometry, steady state and flux metabolomics. RESULTS: Inactivating KEAP1 mutations or reductions in KEAP1 RNA correlated with Nrf2 activation in CDDP-resistant cells, which functionally contributed to resistance. Proteomics identified elevation of downstream Nrf2 targets and the enrichment of enzymes involved in generation of biomass and reducing equivalents, metabolism of glucose, glutathione, NAD(P), and oxoacids. This was accompanied by biochemical and metabolic evidence of an enhanced reductive state dependent on coordinated glucose and glutamine catabolism, associated with reduced energy production and proliferation, despite normal mitochondrial structure and function. CONCLUSIONS: Our analysis identified coordinated metabolic changes associated with CDDP resistance that may provide new therapeutic avenues through targeting of these convergent pathways.

Stable Cell Clones Harboring Self-Replicating SARS-CoV-2 RNAs for Drug Screen.

The development of antivirals against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been hampered by the lack of efficient cell-based replication systems that are amenable to high-throughput screens in biosafety level 2 laboratories. Here we report that stable cell clones harboring autonomously replicating SARS-CoV-2 RNAs without spike (S), membrane (M), and envelope (E) genes can be efficiently derived from the baby hamster kidney (BHK-21) cell line when a pair of mutations were introduced into the non-structural protein 1 (Nsp1) of SARS-CoV-2 to ameliorate cellular toxicity associated with virus replication. In a proof-of-concept experiment we screened a 273-compound library using replicon cells and identified three compounds as novel inhibitors of SARS-CoV-2 replication. Altogether, this work establishes a robust, cell-based system for genetic and functional analyses of SARS-CoV-2 replication and for the development of antiviral drugs. IMPORTANCE SARS-CoV-2 replicon systems that have been reported up to date were unsuccessful in deriving stable cell lines harboring non-cytopathic replicons. The transient expression of viral sgmRNA or a reporter gene makes it impractical for industry-scale screening of large compound libraries using these systems. Here, for the first time, we derived stable cell clones harboring the SARS-CoV-2 replicon. These clones may now be conveniently cultured in a standard BSL-2 laboratory for high throughput screen of compound libraries. Additionally, our stable replicon cells represent a new model system to study SARS-CoV-2 replication.

Characterization of Entry Pathways, Species-Specific Angiotensin-Converting Enzyme 2 Residues Determining Entry, and Antibody Neutralization Evasion of Omicron BA.1, BA.1.1, BA.2, and BA.3 Variants.

The SARS-CoV-2 Omicron variants were first detected in November 2021, and several Omicron lineages (BA.1, BA.2, BA.3, BA.4, and BA.5) have since rapidly emerged. Studies characterizing the mechanisms of Omicron variant infection and sensitivity to neutralizing antibodies induced upon vaccination are ongoing by several groups. In the present study, we used pseudoviruses to show that the transmembrane serine protease 2 (TMPRSS2) enhances infection of BA.1, BA.1.1, BA.2, and BA.3 Omicron variants to a lesser extent than ancestral D614G. We further show that Omicron variants have higher sensitivity to inhibition by soluble angiotensin-converting enzyme 2 (ACE2) and the endosomal inhibitor chloroquine compared to D614G. The Omicron variants also more efficiently used ACE2 receptors from 9 out of 10 animal species tested, and unlike the D614G variant, used mouse ACE2 due to the Q493R and Q498R spike substitutions. Finally, neutralization of the Omicron variants by antibodies induced by three doses of Pfizer/BNT162b2 mRNA vaccine was 7- to 8-fold less potent than the D614G. These results provide insights into the transmissibility and immune evasion capacity of the emerging Omicron variants to curb their ongoing spread. IMPORTANCE The ongoing emergence of SARS-CoV-2 Omicron variants with an extensive number of spike mutations poses a significant public health and zoonotic concern due to enhanced transmission fitness and escape from neutralizing antibodies. We studied three Omicron lineage variants (BA.1, BA.2, and BA.3) and found that transmembrane serine protease 2 has less influence on Omicron entry into cells than on D614G, and Omicron exhibits greater sensitivity to endosomal entry inhibition compared to D614G. In addition, Omicron displays more efficient usage of diverse animal species ACE2 receptors than D614G. Furthermore, due to Q493R/Q498R substitutions in spike, Omicron, but not D614G, can use the mouse ACE2 receptor. Finally, three doses of Pfizer/BNT162b2 mRNA vaccination elicit high neutralization titers against Omicron variants, although the neutralization titers are still 7- to 8-fold lower those that against D614G. These results may give insights into the transmissibility and immune evasion capacity of the emerging Omicron variants to curb their ongoing spread.

Machine learning-based model predictive controller design for cell culture processes.

The biopharmaceutical industry continuously seeks to optimize the critical quality attributes to maintain the reliability and cost-effectiveness of its products. Such optimization demands a scalable and optimal control strategy to meet the process constraints and objectives. This work uses a model predictive controller (MPC) to compute an optimal feeding strategy leading to maximized cell growth and metabolite production in fed-batch cell culture processes. The lack of high-fidelity physics-based models and the high complexity of cell culture processes motivated us to use machine learning algorithms in the forecast model to aid our development. We took advantage of linear regression, the Gaussian process and neural network models in the MPC design to maximize the daily protein production for each batch. The control scheme of the cell culture process solves an optimization problem while maintaining all metabolites and cell culture process variables within the specification. The linear and nonlinear models are developed based on real cell culture process data, and the performance of the designed controllers is evaluated by running several real-time experiments.

Stereotactic and Functional Neurosurgery Convection-Enhanced Delivery of Autologous Cerebrospinal Fluid Enhances Basal Ganglia Visualization during MRI-Guided Deep Brain Stimulation Surgery.

INTRODUCTION: The aim of this study was to determine the safety and feasibility of convection-enhanced delivery of autologous cerebrospinal fluid (CSF) for enhancing intraoperative magnetic resonance imaging (MRI) of the basal ganglia during stereotactic neurosurgery. METHODS: This pilot study was conducted in 4 patients with Parkinson's disease (PD) who underwent MRI-guided deep brain stimulation of the globus pallidus internus (GPi). CSF was obtained via lumbar puncture after general anesthesia and prior to incision. A frameless stereotaxy system was installed, and an infusion catheter was inserted to the GPi using intraoperative MRI. Infusion of autologous CSF was performed at a convective rate of 5 µL/min with a maximum volume of infusion (Vi) of 500 mL. T2-weighted MRI scans were obtained every 15 min up to a maximum of 105 min in order to calculate the volume of distribution (Vd). Safety was assessed with adverse event monitoring, and clinical outcomes were measured with changes in unmedicated UPDRS part III and PDQ-39 scores from baseline to 6 months postoperatively. RESULTS: All four infusions were safe and without adverse events. The mean unmedicated UPDRS part III and PDQ-39 scores improved by 24% and 26%, respectively. The Vd:Vi ratio ranged from 2.2 to 2.8 and peaked 45 min from the onset of infusion, which is when the borders of the GPi could generally be visualized based on T2-weighted MRI. Two patients underwent refinement of the stereotactic targeting based on infusion-enhanced images. CONCLUSIONS: The convective administration of autologous CSF to deep brain structures appears safe and feasible for enhancing intraoperative MRI during stereotactic procedures. Infusion-enhanced imaging with target-specific infusates could be developed to visualize neurochemical circuits or cellular regions that currently are not seen with anatomic/structural MRI.

Pharmacokinetics and Efficacy of Human Hyperimmune Intravenous Immunoglobulin Treatment of Severe Acute Respiratory Syndrome Coronavirus 2 Infection in Adult Syrian Hamsters.

BACKGROUND: After the failure of antibody therapies in treating hospitalized patients with coronavirus disease 2019 (COVID-19), we investigated the impact of viral replication on the pharmacokinetics and efficacy of a hyperimmune severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) immunoglobulin (CoVIG) product in treating SARS-CoV-2 infection using an adult Syrian hamster model. METHODS: The CoVIG was manufactured from plasma donors who had recovered from COVID-19. The dose used (400 mg/kg) was based on the dose given in clinical trials to hospitalized patients with COVID-19. Hamsters were given a single dose of CoVIG 2 days after challenge with the SARS-CoV-2 virus (isolate NY/PV08410/2020), followed by sampling of blood, nasal, tracheal, and lung tissues at different time points. The blood samples were assayed for anti-SARS-CoV-2 spike binding and used to calculate pharmacokinetic (PK) parameters. Nasal wash, tracheal, and lung tissue samples were assayed for viral replication by polymerase chain reaction (subgenomic messenger RNA). RESULTS: CoVIG-treated hamsters showed a reduction in viral replication in the lower respiratory tract, but minimal reduction in the upper respiratory tract, after challenge with SARS-CoV-2. Challenge resulted in altered PK parameters proportionate to viral replication, resulting in decreased area under the curve, accelerated clearance, and shorter half-life of CoVIG. CONCLUSIONS: These data indicate that in the presence of actively replicating SARS-CoV-2 virus, PK parameters are altered and should trigger an adjustment in CoVIG dosing.

The PRRA insert at the S1/S2 site modulates cellular tropism of SARS-CoV-2 and ACE2 usage by the closely related Bat RaTG13.

Biochemical and structural analyses suggest that SARS-CoV-2 is well-adapted to infecting humans and the presence of four residues (PRRA) at the S1/S2 site within the spike (S) protein, which may lead to unexpected tissue or host tropism. Here we report that SARS-CoV-2 efficiently utilized ACE2 of 9 species to infect 293T cells. Similarly, pseudoviruses bearing S protein derived from either the bat RaTG13 or pangolin GX, two closely related animal coronaviruses, utilized ACE2 of a diverse range of animal species to gain entry. Removal of PRRA from SARS-CoV-2 S protein displayed distinct effects on pseudoviral entry into different cell types. Unexpectedly, insertion of PRRA into the RaTG13 S protein selectively abrogated the usage of horseshoe bat and pangolin ACE2 but enhanced the usage of mouse ACE2 by the relevant pseudovirus to enter cells. Together, our findings identified a previously unrecognized effect of the PRRA insert on SARS-CoV-2 and RaTG13 S proteins.ImportanceThe four-residue insert (PRRA) at the boundary between the S1and S2 subunits of SARS-CoV-2 has been widely recognized since day 1 for its role in SARS-CoV-2 S protein processing and activation. As this PRRA insert is unique to SARS-CoV-2 among group b betacoronaviruses, it is thought to affect the tissue and species tropism of SARS-CoV-2. We compared the usage of 10 ACE2 orthologs and found that the presence of PRRA not only affects the cellular tropism of SARS-CoV-2 but also modulates the usage of ACE2 orthologs by the closely related bat RaTG13 S protein. The binding of pseudovirions carrying RaTG13 S with a PRRA insert to mouse ACE2 was nearly 2-fold higher than that of pseudovirions carrying RaTG13 S.

Long-term immunity in convalescent Syrian hamsters provides protection against new-variant SARS-CoV-2 infection of the lower but not upper respiratory tract.

COVID-19 vaccines provide high levels of protection against severe disease and hospitalization due to severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) infection. Vaccination may be less effective in preventing shedding of infectious viruses from otherwise immune patients. In this study, we describe breakthrough infections and shedding of infectious viruses in convalescent hamsters without significant replication in the lower respiratory tract following reinfection by Alpha and Delta variants despite high levels of circulating antibodies in sera. Using convalescent hamsters with long-term immunity (up to 1 year) following infection by ancestral SARS-CoV-2, we can model aspects of recurring COVID-19 in the context of preexisting immunity.

Feasibility of fractionated gamma knife radiosurgery in the management of newly diagnosed Glioblastoma.

BACKGROUND: Glioblastoma (GBM) is the most common primary malignant brain tumor in adults, with overall survival remaining poor despite ongoing efforts to explore new treatment paradigms. Given these outcomes, efforts have been made to shorten treatment time. Recent data report on the safety of CyberKnife (CK) fractionated stereotactic radiosurgery (SRS) in the management of GBM using a five-fraction regimen. The latest Gamma Knife (GK) model also supports frameless SRS, and outcomes using GK SRS in the management of primary GBM have not yet been reported. OBJECTIVE: To report on the feasibility of five-fraction SRS with the GammaKnife ICON in the management of newly diagnosed GBM. METHODS: In this single institutional study, we retrospectively reviewed all patients from our medical center from January 2017 through December 2021 who received fractionated SRS with Gamma Knife ICON for newly diagnosed GBM. Patient demographics, upfront surgical margins, molecular subtyping, radiation treatment volumes, systemic therapies, and follow-up imaging findings were extracted to report on oncologic outcomes. RESULTS: We identified six patients treated within the above time frame. Median age at diagnosis was 73.5 years, 66% were male, and had a median Karnofsky Performance Status (KPS) of 70. All tumors were IDH wild-type, and all but one were MGMT methylated and received concurrent temozolomide (TMZ). Within this group, progression free survival was comparable to that of historical data without significant radiation-induced toxicities. CONCLUSION: Gamma Knife ICON may be discussed as a potential treatment option for select GBM patients and warrants further investigation in the prospective setting.

Patient-Reported Outcomes and Predictive Factors following Focused Ultrasound Thalamotomy for Essential Tremor.

INTRODUCTION: The objectives of this study were to determine long-term patient-reported outcomes with magnetic resonance-guided focused ultrasound (MRgFUS) thalamotomy for medication-refractory essential tremor (ET) and to identify risk factors for a poor clinical outcome. METHODS: We administered a telephone or mail-in survey to patients who consecutively underwent unilateral MRgFUS thalamotomy for ET at our institution over an 8-year period. Patients were asked to self-report measures of hand tremor improvement, degree of overall postprocedure improvement, activities of daily life, side effects, and willingness to undergo the procedure again. Specific patient characteristics, ultrasound treatment parameters, and postoperative radiological findings from magnetic resonance imaging performed 1 day after the procedure were analyzed, and multivariable linear regression was used to determine if these factors could serve as predictors of clinical outcome. RESULTS: A total of 85 patients were included in this study with a mean follow-up time of 3.0 years (range 2 months to 1 8.4 years). The mean patient-reported improvement in hand tremor at last follow-up was 66%, and 73% of patients reported meaningful change in their overall condition after the procedure. The percentages of patients reporting normal or only minimal limitations with feeding, drinking, and writing ability at last follow-up were 60%, 71%, and 48%, respectively. In the position of their former selves, 89% of patients would again choose to undergo the procedure. Larger lesions were correlated with a higher risk of adverse events. DISCUSSION/CONCLUSION: While subjective hand tremor improvement declines with time, willingness to undergo the procedure again following MRgFUS thalamotomy for ET remains very high even several years after the procedure.

HIF-1α-Dependent Metabolic Reprogramming, Oxidative Stress, and Bioenergetic Dysfunction in SARS-CoV-2-Infected Hamsters.

The mechanistic interplay between SARS-CoV-2 infection, inflammation, and oxygen homeostasis is not well defined. Here, we show that the hypoxia-inducible factor (HIF-1α) transcriptional pathway is activated, perhaps due to a lack of oxygen or an accumulation of mitochondrial reactive oxygen species (ROS) in the lungs of adult Syrian hamsters infected with SARS-CoV-2. Prominent nuclear localization of HIF-1α and increased expression of HIF-1α target proteins, including glucose transporter 1 (Glut1), lactate dehydrogenase (LDH), and pyruvate dehydrogenase kinase-1 (PDK1), were observed in areas of lung consolidation filled with infiltrating monocytes/macrophages. Upregulation of these HIF-1α target proteins was accompanied by a rise in glycolysis as measured by extracellular acidification rate (ECAR) in lung homogenates. A concomitant reduction in mitochondrial respiration was also observed as indicated by a partial loss of oxygen consumption rates (OCR) in isolated mitochondrial fractions of SARS-CoV-2-infected hamster lungs. Proteomic analysis further revealed specific deficits in the mitochondrial ATP synthase (Atp5a1) within complex V and in the ATP/ADP translocase (Slc25a4). The activation of HIF-1α in inflammatory macrophages may also drive proinflammatory cytokine production and complement activation and oxidative stress in infected lungs. Together, these findings support a role for HIF-1α as a central mediator of the metabolic reprogramming, inflammation, and bioenergetic dysfunction associated with SARS-CoV-2 infection.

Robust Wet Milling Technique for Producing Micronized Ibuprofen Particles with Improved Solubility and Dissolution.

This study investigated a systematic approach for producing ibuprofen (IBF) particles with leucine by wet milling. Using a high shear homogenizer, the particles size of the IBF was reduced. Prepared IBF microparticles were freeze-dried and characterized by using Mastersizer, SEM, DSC, XRD, ATR-FTIR, and TGA. The drug saturation solubility and in-vitro dissolution performance were carried out in phosphate buffer solution (PBS, pH 7.4) at 37°C temperature and IBF were determined using a validated HPLC method. The wet-milled method reduced the particle size from 71.3 to 1.7 µm. The minimum particle size of IBF was obtained in 0.05% Tween 80 solution homogenized at 17,000 rpm for 15 min. The saturated solubility (168.7 µg/mL) of the micronized IBF particles with leucine showed higher compared to that of the original IBF (147.4 µg/mL) in PBS solution. The prepared IBF particles containing 2.5-6.25% leucine showed significantly higher IBF release (100%) compared to that of original drug particles (55.9%) in 120 min. The excipient leucine played a major role in enhancing the solubility and dissolution profile of the prepared IBF particles probably by the formation of hydrogen bonding. The developed wet milling was an efficient and robust technique for reducing the particle size of IBF and could be a useful method for manufacturing drug particles with enhanced solubility and dissolution.