Clinical trial links oncolytic immunoactivation to survival in glioblastoma.

Immunotherapy failures can result from the highly suppressive tumour microenvironment that characterizes aggressive forms of cancer such as recurrent glioblastoma (rGBM)1,2. Here we report the results of a first-in-human phase I trial in 41 patients with rGBM who were injected with CAN-3110-an oncolytic herpes virus (oHSV)3. In contrast to other clinical oHSVs, CAN-3110 retains the viral neurovirulence ICP34.5 gene transcribed by a nestin promoter; nestin is overexpressed in GBM and other invasive tumours, but not in the adult brain or healthy differentiated tissue4. These modifications confer CAN-3110 with preferential tumour replication. No dose-limiting toxicities were encountered. Positive HSV1 serology was significantly associated with both improved survival and clearance of CAN-3110 from injected tumours. Survival after treatment, particularly in individuals seropositive for HSV1, was significantly associated with (1) changes in tumour/PBMC T cell counts and clonal diversity, (2) peripheral expansion/contraction of specific T cell clonotypes; and (3) tumour transcriptomic signatures of immune activation. These results provide human validation that intralesional oHSV treatment enhances anticancer immune responses even in immunosuppressive tumour microenvironments, particularly in individuals with cognate serology to the injected virus. This provides a biological rationale for use of this oncolytic modality in cancers that are otherwise unresponsive to immunotherapy (ClinicalTrials.gov: NCT03152318 ).

Novel Approach for Pancreas Transcriptomics Reveals the Cellular Landscape in Homeostasis and Acute Pancreatitis.

BACKGROUND & AIMS: Acinar cells produce digestive enzymes that impede transcriptomic characterization of the exocrine pancreas. Thus, single-cell RNA-sequencing studies of the pancreas underrepresent acinar cells relative to histological expectations, and a robust approach to capture pancreatic cell responses in disease states is needed. We sought to innovate a method that overcomes these challenges to accelerate study of the pancreas in health and disease. METHODS: We leverage FixNCut, a single-cell RNA-sequencing approach in which tissue is reversibly fixed with dithiobis(succinimidyl propionate) before dissociation and single-cell preparation. We apply FixNCut to an established mouse model of acute pancreatitis, validate findings using GeoMx whole transcriptome atlas profiling, and integrate our data with prior studies to compare our method in both mouse and human pancreas datasets. RESULTS: FixNCut achieves unprecedented definition of challenging pancreatic cells, including acinar and immune populations in homeostasis and acute pancreatitis, and identifies changes in all major cell types during injury and recovery. We define the acinar transcriptome during homeostasis and acinar-to-ductal metaplasia and establish a unique gene set to measure deviation from normal acinar identity. We characterize pancreatic immune cells, and analysis of T-cell subsets reveals a polarization of the homeostatic pancreas toward type-2 immunity. We report immune responses during acute pancreatitis and recovery, including early neutrophil infiltration, expansion of dendritic cell subsets, and a substantial shift in the transcriptome of macrophages due to both resident macrophage activation and monocyte infiltration. CONCLUSIONS: FixNCut preserves pancreatic transcriptomes to uncover novel cell states during homeostasis and following pancreatitis, establishing a broadly applicable approach and reference atlas for study of pancreas biology and disease.

Epidemiology of Craniomaxillofacial Fractures Over a 5-year Period at a Midwestern Level 1 Trauma Center Serving a Large Rural Population.

BACKGROUND: Though the epidemiology of craniomaxillofacial (CMF) fractures has been well documented at urban hospitals, the characteristics of these fractures in rural hospitals have not been well studied. PURPOSE: The purpose of this study is to report on the epidemiology of CMF fractures at a regional Level 1 trauma center serving a large rural population in central Illinois. STUDY DESIGN, SETTING, SAMPLE: This is a retrospective cohort study at a community-based regional tertiary referral center that serves a predominantly rural population. Inclusion criteria comprised patients with radiologically confirmed CMF fractures between 2015 and 2019. Patients with incomplete medical records were excluded. PREDICTOR VARIABLE: Predictor variables included demographics (age, admission source, race, and sex) and etiology of CMF fracture (assault/domestic violence, all-terrain vehicle/off-road, falls, farm-related, motor vehicle collisions, gunshot wound, and others). MAIN OUTCOME VARIABLE: The primary outcome variable was the CMF anatomic location including nasal bone, orbit, mandible, malar/maxillary, and other CMF fractures. COVARIATES: The covariates are not applicable. ANALYSES: Descriptive statistics were used to summarize a sample of the population characteristics. Wilcoxon ranked sign tests and χ2 tests of independence were used to assess for statistically significant associations between select variables of interest. Statistical significance was defined as P < .05. RESULTS: Between 2015 and 2019, a total of 2,334 patients presented to the emergency department with a CMF fracture. After applying the inclusion/exclusion criteria, the final sample was composed of 1,844 patients for the management of 2,405 CMF fractures. The majority of patients were male(62.0%) and young adults (aged 18-39) had the highest number of CMF fractures (819) relative to all other age groups. The most common fracture etiology was fall(37.3%), and nasal bone fractures represented the most common fracture location(41.6%). χ2 analyses revealed statistically significant associations between the anatomic location of CMF fracture incurred, and differing categories of age, admission source, race, sex, and etiology. CONCLUSION AND RELEVANCE: Our study shows that patients seen at our Midwestern Level 1 trauma center are more likely to present with nasal bone and malar/maxillary fractures due to falls. In studies based in urban centers, patients are likely to present with orbital and mandibular fractures due to falls and assault.

Evaluation of the drug disposition of RO7049389 with in vitro data and human mass balance supported by physiologically based pharmacokinetic modelling.

AIMS: RO7049389 (linvencorvir) is a developmental oral treatment for chronic hepatitis B virus infection. The aim of this work was to conduct mass balance (MB) and absolute bioavailability (BA) analyses in healthy volunteers, alongside in vitro evaluations of the metabolism of RO7049389 and a major circulating active metabolite M5 in human hepatocytes, and physiologically based pharmacokinetic (PBPK) modelling to refine the underlying drug disposition paradigm. METHODS: Participants in the clinical study (MB: Caucasian, male, n = 6; BA: Caucasian and Asian, male and female, n = 16, 8 in each ethnic groups) received oral [14 C] or unlabelled RO7049389 (600/1000 mg) followed by 100 µg intravenous [13 C]RO7049389. Metabolic pathways with fractions metabolized-obtained from the in vitro incubation results of 10 µM [14 C]RO7049389 and 1 µM M5 with (long-term cocultured) human hepatocytes in the absence and presence of the cytochrome P450 3A4 (CYP3A4) inhibitor itraconazole-were used to complement the PBPK models, alongside the clinical MB and BA data. RESULTS: The model performance in predicting the pharmacokinetic profiles of RO7049389 and M5 aligned with clinical observations in Caucasians and was also successfully applied to Asians. Accordingly, the drug disposition pathways for RO7049389 were postulated with newly characterized estimates of the fractions: biliary excretion by P-glycoprotein (~41%), direct glucuronidation via uridine 5'-diphosphoglucuronosyltransferase 1A3 (~11%), hexose conjugation (~6%), oxidation by CYP3A4 (~28%) and other oxidation reactions (~9%). CONCLUSION: These results support the ongoing clinical development program for RO7049389 and highlight the broader value of PBPK and MB analyses in drug development.

Design and Synthesis of Stem Cell-Laden Keratin/Glycol Chitosan Methacrylate Bioinks for 3D Bioprinting.

With the advancements in tissue engineering and three-dimensional (3D) bioprinting, physiologically relevant three-dimensional structures with suitable mechanical and bioactive properties that mimic the biological tissue can be designed and fabricated. However, the available bioinks are less than demanded. In this research, the readily available biomass sources, keratin and glycol chitosan, were selected to develop a UV-curable hydrogel that is feasible for the 3D bioprinting process. Keratin methacrylate and glycol chitosan methacrylate were synthesized, and a hybrid bioink was created by combining this protein-polysaccharide cross-linked hydrogel. While human hair keratin could provide biological functions, the other composition, glycol chitosan, could further enhance the mechanical strength of the construct. The mechanical properties, degradation profile, swelling behavior, cell viability, and proliferation were investigated with various ratios of keratin methacrylate to glycol chitosan methacrylate. The composition of 2% (w/v) keratin methacrylate and 2% (w/v) chitosan methacrylate showed a significantly higher cell number and swelling percentage than other compositions and was designated as the bioink for 3D printing afterward. The feasibility of stem cell loading in the selected formula was examined with an extrusion-based bioprinter. The cells and spheroids can be successfully printed with the synthesized bioink into a specific shape and cultured. This work provides a potential option for bioinks and delivers insights into personalization research on stem cell-laden biofabricated hydrogels in the future.

Intelligent Electromagnetic Sensors for Non-Invasive Trojan Detection.

Rapid growth of sensors and the Internet of Things is transforming society, the economy and the quality of life. Many devices at the extreme edge collect and transmit sensitive information wirelessly for remote computing. The device behavior can be monitored through side-channel emissions, including power consumption and electromagnetic (EM) emissions. This study presents a holistic self-testing approach incorporating nanoscale EM sensing devices and an energy-efficient learning module to detect security threats and malicious attacks directly at the front-end sensors. The built-in threat detection approach using the intelligent EM sensors distributed on the power lines is developed to detect abnormal data activities without degrading the performance while achieving good energy efficiency. The minimal usage of energy and space can allow the energy-constrained wireless devices to have an on-chip detection system to predict malicious attacks rapidly in the front line.

K-means clustering of coherent Raman spectra from extracellular vesicles visualized by label-free multiphoton imaging.

Extracellular vesicles (EVs) have emerged as potential biomarkers in cancer research and for clinical diagnosis. Little is known, however, about their spatial distributions in tissue and the different subpopulations that may exist. Here we report the use of label-free nonlinear optical imaging techniques to provide spatially resolved chemical information of EVs within untreated tissues. A multimodal nonlinear optical imaging system incorporating multiphoton autofluorescence and hyperspectral coherent anti-Stokes Raman scattering (CARS) imaging was built to visualize the spatial tissue distribution and probe the spectra of EVs. K-means clustering is performed on the CARS spectra from EVs in rat mammary tissues and human breast tumor tissue to reveal both the spatial distribution of EV clusters and their different chemical signatures. Correlations are identified between EV clusters and metabolic information.

Home Safety Fall and Accident Risk Among Prematurely Aging, Formerly Homeless Adults.

OBJECTIVE: Homelessness prematurely ages people. A large subgroup of formerly homeless adults between ages 40 and 64 yr have health conditions similar to or worse than people categorized as elderly. Little is known about the impact of this group's chronic health conditions on their ability to safely function in supportive housing. METHOD: Home safety visits were carried out with 25 formerly homeless adults, ages 40-64 yr, now residing in supportive housing. RESULTS: Participants had physical, cognitive, and mental health problems that significantly interfered with their ability to perform daily life skills, safely function in an apartment, and manage chronic health conditions. Home safety hazards included cluttered walking paths, the presence of steps, and the lack of grab bars and nonskid flooring. CONCLUSION: The homeless population would benefit from aging specialists, such as occupational therapists, who could help people to maintain and function more safely in their homes. Without such services, this population may be at risk for home safety events leading to hospitalization and mortality.

Tunable Chemochromism and Elastic Properties in Intercalated MoO3: Au-, Cr-, Fe-, Ge-, Mn-, and Ni-MoO3.

Chemical tunability of the elastic constants of α-MoO3, a two-dimensional layered oxide, is demonstrated with mutability on the order of tens of GPa, simply by choice of a metal intercalant including Au, Cr, Fe, Ge, Mn, and Ni. Using Brillouin laser light scattering from confined acoustic phonons in nanometer-thick materials, the in-plane angular dispersion of the quantized acoustic phonon branches of 2D layered, intercalated MoO3 is measured and used to determine the bulk modulus (K), Young's moduli (E11, E22, and E33), each of the nine independent elastic tensor elements (cij), and the thickness. Intercalation of metals generally reduces the anisotropy in MoO3 except in Ge-MoO3, for which the in-plane longitudinal elastic anisotropy is unaffected. Chemochromism from transparent white (MoO3 and Fe-MoO3) to near black (Ni-MoO3) to brilliant dark blue (Ge-MoO3) is demonstrated and is associated with a reduction in electronic band gap with intercalation and an increase in absorption >600 nm for some intercalants (Cr-, Ge-, and Mn-MoO3).

Lessons learned from phase 3 trials of immunotherapy for glioblastoma: Time for longitudinal sampling?

Glioblastoma (GBM)'s median overall survival is almost 21 months. Six phase 3 immunotherapy clinical trials have recently been published, yet 5/6 did not meet approval by regulatory bodies. For the sixth, approval is uncertain. Trial failures result from multiple factors, ranging from intrinsic tumor biology to clinical trial design. Understanding the clinical and basic science of these 6 trials is compelled by other immunotherapies reaching the point of advanced phase 3 clinical trial testing. We need to understand more of the science in human GBMs in early trials: the "window of opportunity" design may not be best to understand complex changes brought about by immunotherapeutic perturbations of the GBM microenvironment. The convergence of increased safety of image-guided biopsies with "multi-omics" of small cell numbers now permits longitudinal sampling of tumor and biofluids to dissect the complex temporal changes in the GBM microenvironment as a function of the immunotherapy.

Sequence clustering confounds AlphaFold2.

Though typically associated with a single folded state, some globular proteins remodel their secondary and/or tertiary structures in response to cellular stimuli. AlphaFold21 (AF2) readily generates one dominant protein structure for these fold-switching (a.k.a. metamorphic) proteins2, but it often fails to predict their alternative experimentally observed structures3,4. Wayment-Steele, et al. steered AF2 to predict alternative structures of a few metamorphic proteins using a method they call AF-cluster5. However, their Paper lacks some essential controls needed to assess AF-cluster's reliability. We find that these controls show AF-cluster to be a poor predictor of metamorphic proteins. First, closer examination of the Paper's results reveals that random sequence sampling outperforms sequence clustering, challenging the claim that AF-cluster works by "deconvolving conflicting sets of couplings." Further, we observe that AF-cluster mistakes some single-folding KaiB homologs for fold switchers, a critical flaw bound to mislead users. Finally, proper error analysis reveals that AF-cluster predicts many correct structures with low confidence and some experimentally unobserved conformations with confidences similar to experimentally observed ones. For these reasons, we suggest using ColabFold6-based random sequence sampling7-augmented by other predictive approaches-as a more accurate and less computationally intense alternative to AF-cluster.

Tailoring Ultrafast Near-Band Gap Photoconductive Response in GeS by Zero-Valent Cu Intercalation.

Zero-valent intercalation of atomic metals into the van der Waals gap of layered materials can be used to tune their electronic, optical, thermal, and mechanical properties. Here, we report the impact of intercalating ∼3 atm percent of zero-valent copper into germanium sulfide (GeS). Advanced many-body calculations predict that copper introduces quasi-localized intermediate band states, and time-resolved THz spectroscopy studies demonstrate that those states have prominent effects on the photoconductivity of GeS. Cu-intercalated GeS exhibits a faster rise of transient photoconductivity and a shorter lifetime of optically injected carriers following near-gap excitation with 800 nm pulses. At the same time, Cu intercalation improves free carrier mobility from 1100 to 1300 cm2 V-1 s-1, which we attribute to the damping of acoustic phonons observed in Brillouin scattering and consequent reduction of phonon scattering.

Recent advances using MXenes in biomedical applications.

An MXene is a novel two-dimensional transition metal carbide or nitride, with a typical formula of Mn+1XnTx (M = transition metals, X = carbon or nitrogen, and T = functional groups). MXenes have found wide application in biomedicine and biosensing, owing to their high biocompatibility, abundant reactive surface groups, good conductivity, and photothermal properties. Applications include photo- and electrochemical sensors, energy storage, and electronics. This review will highlight recent applications of MXene and MXene-derived materials in drug delivery, tissue engineering, antimicrobial activity, and biosensors (optical and electrochemical). We further elaborate on recent developments in utilizing MXenes for photothermal cancer therapy, and we explore multimodal treatments, including the integration of chemotherapeutic agents or magnetic nanoparticles for enhanced therapeutic efficacy. The high surface area and reactivity of MXenes provide an interface to respond to the changes in the environment, allowing MXene-based drug carriers to respond to changes in pH, reactive oxygen species (ROS), and electrical signals for controlled release applications. Furthermore, the conductivity of MXene enables it to provide electrical stimulation for cultured cells and endows it with photocatalytic capabilities that can be used in antibiotic applications. Wearable and in situ sensors incorporating MXenes are also included. Major challenges and future development directions of MXenes in biomedical applications are also discussed. The remarkable properties of MXenes will undoubtedly lead to their increasing use in the applications discussed here, as well as many others.

Dual crosslinking silk fibroin/pectin-based bioink development and the application on neural stem/progenitor cells spheroid laden 3D bioprinting.

The human nervous system is an incredibly intricate physiological network, and neural cells lack the ability to repair and regenerate after a brain injury. 3-dimensional (3D) bioprinting technology offers a promising strategy for constructing biomimetic organ constructs and in vitro brain/disease models. The bioink serves as a pivotal component that emulates the microenvironment of biomimetic construct and exerts a profound influence on cellular behaviors. In this study, a series of mechanically adjustable and dual crosslinking bioinks were developed using photocrosslinkable methacrylated silk fibroin (SilMA) in combination with the ionic crosslinking material, pectin, or pectin methacryloyl (PecMA) with silk fibroin (SF) supplementation. SilMA/pectin exhibited superior properties, with SilMA providing biocompatibility and adjustable mechanical properties, while the addition of pectin enhanced printability. The porous structure supported neural cell growth, and 15 % SilMA/0.5 % pectin bioinks displayed excellent printability and shape fidelity. Neural stem/progenitor cells (NSPCs)-loaded bioinks were used to construct a 3D brain model, demonstrating sustained vitality and high neuronal differentiation without the need for growth factors. The SilMA/pectin bioinks demonstrated adjustable mechanical properties, favorable biocompatibility, and an environment highly conducive to neural induction, offering an alternative approach for neural tissue engineering applications or in vitro brain models.

Efficacy, safety, and pharmacokinetics of capsid assembly modulator linvencorvir plus standard of care in chronic hepatitis B patients.

BACKGROUND/AIMS: Four-week treatment of linvencorvir (RO7049389) was generally safe and well tolerated, and showed anti-viral activity in chronic hepatitis B (CHB) patients. This study evaluated the efficacy, safety, and pharmacokinetics of 48-week treatment with linvencorvir plus standard of care (SoC) in CHB patients. METHODS: This was a multicentre, non-randomized, non-controlled, open-label phase 2 study enrolling three cohorts: nucleos(t)ide analogue (NUC)-suppressed patients received linvencorvir plus NUC (Cohort A, n=32); treatment-naïve patients received linvencorvir plus NUC without (Cohort B, n=10) or with (Cohort C, n=30) pegylated interferon-α (Peg-IFN-α). Treatment duration was 48 weeks, followed by NUC alone for 24 weeks. RESULTS: 68 patients completed the study. No patient achieved functional cure (sustained HBsAg loss and unquantifiable HBV DNA). By Week 48, 89% of treatment-naïve patients (10/10 Cohort B; 24/28 Cohort C) reached unquantifiable HBV DNA. Unquantifiable HBV RNA was achieved in 92% of patients with quantifiable baseline HBV RNA (14/15 Cohort A, 8/8 Cohort B, 22/25 Cohort C) at Week 48 along with partially sustained HBV RNA responses in treatment-naïve patients during follow-up period. Pronounced reductions in HBeAg and HBcrAg were observed in treatment-naïve patients, while HBsAg decline was only observed in Cohort C. Most adverse events were grade 1-2, and no linvencorvir-related serious adverse events were reported. CONCLUSION: 48-week linvencorvir plus SoC was generally safe and well tolerated, and resulted in potent HBV DNA and RNA suppression. However, 48-week linvencorvir plus NUC with or without Peg-IFN did not result in the achievement of functional cure in any patient.

SSDraw: Software for generating comparative protein secondary structure diagrams.

The program SSDraw generates publication-quality protein secondary structure diagrams from three-dimensional protein structures. To depict relationships between secondary structure and other protein features, diagrams can be colored by conservation score, B-factor, or custom scoring. Diagrams of homologous proteins can be registered according to an input multiple sequence alignment. Linear visualization allows the user to stack registered diagrams, facilitating comparison of secondary structure and other properties among homologous proteins. SSDraw can be used to compare secondary structures of homologous proteins with both conserved and divergent folds. It can also generate one secondary structure diagram from an input protein structure of interest. The source code can be downloaded (https://github.com/ncbi/SSDraw) and run locally for rapid structure generation, while a Google Colab notebook allows easy use.

SSDraw: software for generating comparative protein secondary structure diagrams.

The program SSDraw generates publication-quality protein secondary structure diagrams from three-dimensional protein structures. To depict relationships between secondary structure and other protein features, diagrams can be colored by conservation score, B-factor, or custom scoring. Diagrams of homologous proteins can be registered according to an input multiple sequence alignment. Linear visualization allows the user to stack registered diagrams, facilitating comparison of secondary structure and other properties among homologous proteins. SSDraw can be used to compare secondary structures of homologous proteins with both conserved and divergent folds. It can also generate one secondary structure diagram from an input protein structure of interest. The source code can be downloaded (https://github.com/ethanchen1301/SSDraw) and run locally for rapid structure generation, while a Google Colab notebook allows easy use.

ColabFold predicts alternative protein structures from single sequences, coevolution unnecessary for AF-cluster.

Though typically associated with a single folded state, globular proteins are dynamic and often assume alternative or transient structures important for their functions1,2. Wayment-Steele, et al. steered ColabFold3 to predict alternative structures of several proteins using a method they call AF-cluster4. They propose that AF-cluster "enables ColabFold to sample alternate states of known metamorphic proteins with high confidence" by first clustering multiple sequence alignments (MSAs) in a way that "deconvolves" coevolutionary information specific to different conformations and then using these clusters as input for ColabFold. Contrary to this Coevolution Assumption, clustered MSAs are not needed to make these predictions. Rather, these alternative structures can be predicted from single sequences and/or sequence similarity, indicating that coevolutionary information is unnecessary for predictive success and may not be used at all. These results suggest that AF-cluster's predictive scope is likely limited to sequences with distinct-yet-homologous structures within ColabFold's training set.

Hafnium, Titanium, and Zirconium Intercalation in 2D Layered Nanomaterials.

Altering the physical and chemical properties of a layered material through intercalation has emerged as a unique strategy toward tunable applications. In this work, we demonstrate a wet chemical method to intercalate titanium, hafnium, and zirconium into 2D layered nanomaterials. The metals are intercalated using bis-tetrahydrofuran metal halide complexes. Metal intercalation is demonstrated in nanomaterials of Bi2Se3, Si2Te3, MoO3, and GeS. This strategy intercalates, on average, 3 atm % or less of Hf, Ti, and Zr that share charge with the host nanomaterial. This methodology is used to chemochromically alter MoO3 from transparent white to dark blue.

AlphaFold2 has more to learn about protein energy landscapes.

Recent work suggests that AlphaFold2 (AF2)-a deep learning-based model that can accurately infer protein structure from sequence-may discern important features of folded protein energy landscapes, defined by the diversity and frequency of different conformations in the folded state. Here, we test the limits of its predictive power on fold-switching proteins, which assume two structures with regions of distinct secondary and/or tertiary structure. Using several implementations of AF2, including two published enhanced sampling approaches, we generated >280,000 models of 93 fold-switching proteins whose experimentally determined conformations were likely in AF2's training set. Combining all models, AF2 predicted fold switching with a modest success rate of ~25%, indicating that it does not readily sample both experimentally characterized conformations of most fold switchers. Further, AF2's confidence metrics selected against models consistent with experimentally determined fold-switching conformations in favor of inconsistent models. Accordingly, these confidence metrics-though suggested to evaluate protein energetics reliably-did not discriminate between low and high energy states of fold-switching proteins. We then evaluated AF2's performance on seven fold-switching proteins outside of its training set, generating >159,000 models in total. Fold switching was accurately predicted in one of seven targets with moderate confidence. Further, AF2 demonstrated no ability to predict alternative conformations of two newly discovered targets without homologs in the set of 93 fold switchers. These results indicate that AF2 has more to learn about the underlying energetics of protein ensembles and highlight the need for further developments of methods that readily predict multiple protein conformations.