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Cancer cells are especially sensitive to perturbations in ribosome biogenesis as they rely on finely tuned protein homeostasis to facilitate their rapid growth and proliferation. While ribosome synthesis and cancer have a well-established relationship, ribosome biogenesis has only recently drawn interest as a cancer therapeutic target. In this study, we exploited the relationship between ribosome biogenesis and cancer cell proliferation by using a potent ribosome biogenesis inhibitor, RBI2 (Ribosome Biogenesis Inhibitor 2), to perturb cancer cell growth and viability. We demonstrate herein that RBI2 significantly decreases cell viability in malignant melanoma cells and breast cancer cell lines. Treatment with RBI2 dramatically and rapidly decreased ribosomal RNA (rRNA) synthesis, without affecting the occupancy of RNA polymerase I (Pol I) on the ribosomal DNA template. Next-generation RNA sequencing (RNA-seq) revealed that RBI2 and previously described ribosome biogenesis inhibitor CX-5461 induce distinct changes in the transcriptome. An investigation of the content of the pre-rRNAs through RT-qPCR revealed an increase in the polyadenylation of cellular rRNA after treatment with RBI2, constituting a known pathway by which rRNA degradation occurs. Northern blotting revealed that RBI2 does not appear to impair or alter rRNA processing. Collectively, these data suggest that RBI2 inhibits rRNA synthesis differently from other previously described ribosome biogenesis inhibitors, potentially acting through a novel pathway that upregulates the turnover of premature rRNAs.

Since the US Food and Drug Administration (FDA) began monitoring the quality of pharmaceutical manufacturing by enforcing current good manufacturing practices roughly 60 years ago, forces related to the global economy have changed, rendering the task of monitoring quality more difficult. Alternative strategies by groups like Valisure, LLC, and the University of Kentucky Drug Quality Study to monitor the quality of the currently circulated US drug supply through end-product testing and screening have resulted in several concerning findings. Given the successful approaches of identifying quality defects in pharmaceuticals by non-regulatory bodies, and considering the changing landscape and pressures on manufacturing, the FDA, large buying groups, and the US Department of Defense should consider these alternative strategies as a means to augment current regulatory activities.

Chronologies generated from core profiles to apply dates to environmental changes commonly use the measurement of the activity of radionuclides deposited and stratified with physical environmental material. The most commonly reported nuclide to define chronologies covering the last 150 years is Pb-210, for which accepted data processing methodologies in the literature have focussed on the constant rate of supply (CRS) model and the more recently published Bayesian Plum model. This short communication describes a validation approach using defined sediment layers referred to as ‘varve’ counting, which provide known points of reference to account for uncertainty between generated dates from each model using published Pb-210 measurements. A significant improvement in the chronologies was observed when applying reference date corrections to the models. This was shown to be essential in providing confidence in reported datasets and accuracy of predicted chronologies, which will better inform the interpretation of environmental change, e.g. sedimentation rates, climate change, pollution pathways and land degradation. Generated chronologies from both the CRS and Plum methods showed good agreement with the established varve dates (typically

Low-nanomolar binding constants were recorded for a series of six 2′-fluoro-(carbamoylpyridinyl)deschloroepibatidine analogues with acetylcholine-binding protein (AChBP). The crystal structures of three complexes with AChBP reveal details of molecular recognition in the orthosteric binding site and imply how the other three ligands bind. Comparisons exploiting AChBP as a surrogate for α4β2 and α7 nicotinic acetylcholine receptors (nAChRs) suggest that the key interactions are conserved. The ligands interact with the same residues as the archetypal nAChR agonist nicotine yet display greater affinity, thereby rationalizing their in vivo activity as potent antagonists of nicotine-induced antinociception. An oxyanion-binding site is formed on the periphery of the AChBP orthosteric site by Lys42, Asp94, Glu170 and Glu210. These residues are highly conserved in the human α4, β2 and α7 nAChR sequences. However, specific sequence differences are discussed that could contribute to nAChR subtype selectivity and in addition may represent a point of allosteric modulation. The ability to engage with this peripheral site may explain, in part, the function of a subset of ligands to act as agonists of α7 nAChR.

HIV-1 Nef is an attractive target for antiretroviral drug discovery because of its roles promoting HIV-1 infectivity, replication, and host immune system avoidance. Here we applied a screening strategy in which recombinant HIV-1 Nef protein was coupled to activation of the Src-family tyrosine kinase Hck, which enhances the HIV-1 life cycle in macrophages. Nef stimulates recombinant Hck activity in vitro, providing a robust assay for chemical library screening. High-throughput screening of more than 730,000 compounds using the Nef•Hck assay identified six unique hit compounds that bound directly to recombinant Nef by SPR in vitro and inhibited HIV-1 replication in primary macrophages in the 0.04 to 5 μM range without cytotoxicity. Eighty-four analogs were synthesized around an isothiazolone scaffold from this series, many of which bound to recombinant Nef and inhibited HIV-1 infectivity in the low to submicromolar range. Compounds in this series restored MHC-I to the surface of HIV-infected primary cells and disrupted a recombinant protein complex of Nef with the C-terminal tail of MHC-I and the μ1 subunit of the AP-1 endocytic trafficking protein. Nef inhibitors in this class have the potential to block HIV-1 replication in myeloid cells and trigger recognition of HIV-infected cells by the adaptive immune system in vivo.

Delivering cargo to the central nervous system (CNS) remains a pharmacological challenge. For infectious diseases such as HIV, the CNS acts as a latent reservoir that is inadequately managed by systemic antiretrovirals (ARTs). ARTs thus cannot eradicate HIV, and given CNS infection, patients experience an array of neurological deficits that are collectively referred to as ‘neuroHIV’. Herein we report the development of bioinspired ionic liquid-coated nanoparticles (IL-NPs) for in situ hitchhiking on red blood cells (RBCs), which enabled 48% delivery of intravenously infused cargo to the brain. Moreover, the ionic liquid (IL) choline trans-2-hexenoate (CA2HA 1:2) demonstrated preferential accumulation in parenchymal microglia over endothelial cells post-delivery. We further demonstrate the successful loading of abacavir (ABC), an ART that is challenging to encapsulate, into the IL-coated NPs and verify the retention of antiviral efficacy in vitro. IL-NPs were not cytotoxic to primary human peripheral blood mononuclear cells (PBMCs) and the CA2HA 1:2 coating conferred notable anti-viremic capacity on its own. In addition, in vitro cell culture assays showed markedly increased uptake of IL-coated nanoparticles into neuronal cells compared to bare nanoparticles. This work debuts bioinspired ionic liquids as promising nanoparticle coatings to assist CNS biodistribution and has the potential to revolutionize the delivery of cargos (i.e., drugs, viral vectors) through compartmental barriers such as the blood-brain-barrier (BBB), illustrated in the graphical abstract below.

The KMT2 family of genes is essential epigenetic regulators promoting gene expression. The gene family contains three subgroups, each with two paralogues: KMT2A and KMT2B; KMT2C and KMT2D; KMT2F and KMT2G. KMT2A-D are among the most frequent somatically altered genes in several different cancer types. Somatic KMT2A rearrangements are well-characterized in infant leukemia (IL), and growing evidence supports the role of additional family members (KMT2B, KMT2C, and KMT2D) in leukemogenesis. Enrichment of rare heterozygous frameshift variants in KMT2A and C has been reported in acute myeloid leukemia (AML), IL, and solid tumors. Currently, the non-synonymous variation, prevalence, and penetrance of these four genes are unknown.This study determined the prevalence of pathogenic/likely pathogenic (P/LP) germline KMT2A-D variants in a cancer-free adult population from the Genome Aggregation Database (gnomAD). Two methods of variant interpretation were utilized: a manual genomic variant interpretation and an automated ACMG pipeline.The ACMG pipeline identified considerably fewer P/LP variants (n = 89) compared to the manual method (n = 660) in all 4 genes. Consequently, the total P/LP prevalence and allele frequency (AF) were higher in the manual method (1:112, AF = 4.46E-03) than in ACMG (1:832, AF = 6.01E-04). Multiple ancestry-exclusive P/LP variants were identified along with an increased frequency in males compared to females. Many of these variants identified in this population database are also associated with severe juvenile conditions.These data demonstrate that putatively functional germline variation in these developmentally important genes is more common than previously appreciated and identification in cancer-free adults may indicate incomplete penetrance for many of these variants. Future research should examine a genetic predisposing role in IL and other pediatric cancers.

Background In 2021 the UK Foundation Programme introduced a new curriculum for UK Foundation Doctors (FDs). For the first time, frailty has been designated as a core learning topic. This means that all FDs are expected to undertake dedicated frailty training. Subsequently, a virtual frailty workshop was developed and delivered by Zoom to all F2 doctors in Northern Ireland. Entitled ‘Taking a Better Approach to the Frailer Adult’, the three hour workshop familiarises FDs with core frailty concepts including Comprehensive Geriatric Assessment (CGA), delirium, and deprescribing. Methods All attendees completed an online survey after workshops. They were asked to provide free text comments about what they felt were the most successful and useful aspects of the workshop. Attendees were also were asked to numerically rate the workshop relevance, handout quality and venue suitability. Results The frailty workshop was delivered to 222 F2 doctors over eight months. On average, attendees rated the relevance of the workshop as 4.8/5. Attendees rated the quality of the handouts and the venue suitability at 4.4/5 and 4.2/5 respectively. Thematic analysis of qualitative feedback on the most successful elements of the workshop showed that attendees found the session to be useful, relevant, and engaging, with particular preference for use of cases. For example, one attendee wrote that, ‘use of cases and breakout rooms to discuss CGA [allowed] us to get a feel of how this applies to patients in clinical settings”. Thematic analysis of qualitative feedback on what could be done to make the workshop better showed attendees had preferences for greater use of cases and smaller breakout rooms. This feedback was used to develop and improve the workshop. Conclusion FDs value teaching on frailty and consider it highly relevant for clinical practice. Frailty education can be delivered successfully in a virtual setting. Junior doctors prefer interactive case based learning, with use of breakout rooms with small numbers of participants in each room.

While non-suicidal self-injury (NSSI) occurs in the general population at a surprisingly high rate, with higher rates among certain clinical populations, its etiology is not well-understood. Consequently, the DSM-5 lists NSSI as requiring further research. This study utilizes a translational model of naturally-occurring NSSI to assess the role of early parental neglect and variation in the serotonin transporter genotype (5-HTT) in the etiology of NSSI. Subjects (N = 161) were rhesus macaques (Macaca mulatta) reared in one of three conditions (mother-reared (MR), peer-reared (PR), or surrogate peer-reared (SPR)), and classified as NSSI (n = 18) or non-NSSI (n = 143). Subjects were genotyped for 5-HTT and their behaviors were recorded during an ecologically-meaningful, stress-evoking, intruder paradigm. Two weeks prior to testing, blood samples were obtained and assayed for plasma cortisol and adrenocorticotropic hormone (ACTH) concentrations. NSSI subjects were more likely to be SPR, paralleling human studies showing that individuals that exhibit NSSI tend to have experienced abuse or neglect early in life. Results also indicated that variation in the 5-HTT genotype differentiated the NSSI subjects. NSSI subjects that were homozygous for the L allele exhibited high plasma ACTH and high rates of stress-induced stereotypies; whereas NSSI subjects with the s allele exhibited impulsive behaviors, including frequently approaching the potentially dangerous intruder, high rates of aggressive vocal threats, and more activity. These results suggest that there may be different 5-HTT genotype-mediated NSSI typologies and that both early experiences and variation in the 5-HTT genotype may be important factors in understanding the etiology of NSSI.

Temperament is an individual’s nature and is widely believed to have a heritable foundation. Few studies, however, have evaluated paternal and maternal contributions to the triadic dimensions of temperament. Rhesus monkeys are widely utilized to model genetic contributions to human development due to their close genetic-relatedness and common temperament structure, providing a powerful translational model for investigating paternal and maternal genetic influences on temperament. The temperament of rhesus monkey infants born to 19 different sires and 50 different dams was assessed during the first month of life by comparing the temperament of paternal or maternal half-siblings reared with their mothers in species-normative conditions or reared in a neonatal nursery. Factor scores from three dimensions of temperament were obtained (Orienting/Regulation, Negative Affectivity, and Surgency/Extraversion) and ANOVAs were used to assess genetic effects. For paternal half-siblings, results showed a statistically significant paternal contribution to Orienting/Regulation, Negative Affectivity, and Surgency/Extraversion factor scores. For maternal half-siblings, results showed a statistically significant contribution to Orienting/Regulation factor scores. When parsed by early rearing condition, results showed a paternal contribution Orienting/Regulation, Negative Affectivity, and Surgency/Extraversion scores for paternal half-siblings reared in the neonatal nursery, while there was only a paternal contribution to Surgency/Extraversion for paternal half-siblings reared by their mothers. There was only a maternal contribution to Orienting/Regulation for maternal half-siblings reared by their mothers. These results show that paternal and maternal contributions to temperament vary by environmental context, and that mothers may environmentally buffer their infants from paternal contributions to their temperament.

The affinity and thermodynamic parameters for the interactions of two naturally occurring neurotoxins, (+)-anatoxin-a and (−)-hosieine-A, with acetylcholine-binding protein were investigated using a fluorescence-quenching assay and isothermal titration calorimetry. The crystal structures of their complexes with acetylcholine-binding protein from Aplysia californica (AcAChBP) were determined and reveal details of molecular recognition in the orthosteric binding site. Comparisons treating AcAChBP as a surrogate for human α4β2 and α7 nicotinic acetylcholine receptors (nAChRs) suggest that the molecular features involved in ligand recognition and affinity for the protein targets are conserved. The ligands exploit interactions with similar residues as the archetypal nAChR agonist nicotine, but with greater affinity. (−)-Hosieine-A in particular has a high affinity for AcAChBP driven by a favorable entropic contribution to binding. The ligand affinities help to rationalize the potent biological activity of these alkaloids. The structural data, together with comparisons with related molecules, suggest that there may be opportunities to extend the hosieine-A scaffold to incorporate new interactions with the complementary side of the orthosteric binding site. Such a strategy may guide the design of new entities to target human α4β2 nAChR that may have therapeutic benefit.

Mg(OH)2 suspensions were floated utilising sodium dodecyl sulphate (SDS) and sodium lauroyl isethionate (SLI) collectors, for rapid dewatering of radwaste suspensions. Freundlich adsorption isotherms were first used to compare the adsorption densities of SDS and SLI on Mg(OH)2 surfaces, to determine the maximum monolayer coverage capacity, and were found to be 0.11 μmol m−2 at a dosed concentration of 172 μM for SDS and 0.05 μmol m−2 at a dosed concentration of 188 μM for SLI. The natural and salt induced coagulation kinetics of Mg(OH)2 were examined using static light scattering, where the influence of collector adsorption on particle size distributions was also investigated, to probe potential hydrodynamic limitations of flotation. Particle stabilised foam formation was then characterised using a Bikerman column test, where the dynamic foamability indices (DFIs) of SDS and SLI were determined to be 49 × 103 s L mol−1 and 321 × 103 s L mol−1 respectively. Flotation performance was measured, and a collection efficiency factor used to compare the solid–liquid separation ability of mixed 2.5 vol% suspensions with SDS or SLI, as well as MIBC frother. Optimal performance aligned with collector concentrations relating to predicted maximum monolayer coverage, and whilst both surfactants were effective, SDS systems performed better than SLI in all metrics. Recoveries of >80% of the Mg(OH)2 wastes were achieved, whilst only transferring 35% of the water mass at the optimum SDS dosed concentration of 82 μM, likely due to its denser surface adsorption and minimised lamella water entrainment.

We have used magnetization transfer NMR experiments to measure the exchange rate constant (kex) of the imino protons in the unbound, cocaine-bound, and quinine-bound forms of the cocaine-binding DNA aptamer. Both long-stem 1 (MN4) and short-stem 1 (MN19) variants were analyzed, corresponding to structures with a prefolded secondary structure and ligand-induced-folding versions of this aptamer, respectively. The kex values were measured as a function of temperature from 5 to 45°C to determine the thermodynamics of the base pair opening for MN4. We find that the base pairs close to the ligand-binding site become stronger upon ligand binding, whereas those located away from the binding site do not strengthen. With the buffer conditions used in this study, we observe imino 1H signals in MN19 not previously seen, which leads us to conclude that in the free form, both stem 2 and parts of stem 3 are formed and that the base pairs in stem 1 become structured or more rigid upon binding. This is consistent with the kex values for MN19 decreasing in both stem 1 and at the ligand-binding site. Based on the temperature dependence of the kex values, we find that MN19 is more dynamic than MN4 in the free and both ligand-bound forms. For MN4, ligand-binding results in the reduction of dynamics that are localized to the binding site. These results demonstrate that an aptamer in which the base pairs are preformed also experiences a reduction in dynamics with ligand binding.

Cortical gamma oscillations are believed to be involved in mental processes which are disturbed in schizophrenia. For example, the magnitudes of sensory-evoked oscillations, as measured by auditory steady-state responses (ASSRs) at 40 Hz, are robustly diminished, whereas the baseline gamma power is enhanced in schizophrenia. Such dual gamma oscillation abnormalities are also present in a mouse model of N-methyl-D-aspartate receptor hypofunction (Ppp1r2cre/Grin1 knockout mice). However, it is unclear whether the abnormal gamma oscillations are associated with dysfunction in schizophrenia. We found that glycogen synthase kinase-3 (GSK3) is overactivated in corticolimbic parvalbumin-positive GABAergic interneurons in Grin1 mutant mice. Here we addressed whether GSK3β inhibition reverses both abnormal gamma oscillations and behavioral deficits with high correlation by pharmacological and genetic approach. We demonstrated that the paralog selective-GSK3β inhibitor, but not GSK3α inhibitor, normalizes the diminished ASSRs, excessive baseline gamma power, and deficits in spatial working memory and prepulse inhibition (PPI) of acoustic startle in Grin1 mutant mice. Cell-type specific GSK3B knockdown, but not GSK3A knockdown, also reversed abnormal gamma oscillations and behavioral deficits. Moreover, GSK3B knockdown, but not GSK3A knockdown, reverses the mutants’ in vivo spike synchrony deficits. Finally, ex vivo patch-clamp recording from pairs of neighboring cortical pyramidal neurons showed a reduction of synchronous spontaneous inhibitory-postsynaptic-current events in mutants, which was reversed by GSK3β inhibition genetically and pharmacologically. Together, GSK3β inhibition in corticolimbic interneurons ameliorates the deficits in spatial working memory and PPI, presumably by restoration of synchronous GABA release, synchronous spike firing, and evoked-gamma power increase with lowered baseline power.

Research and development related to the processing of recycled materials has grown progressively in recent times, as scientists endeavour to integrate sustainability criteria, depletion of natural resources, reducing the energy of technological processes involving extraction and processing of raw materials, etc. When it comes to processing slate, industries have to pass over several steps ranging from the extraction of rock blocks until the final products consisting of plates, leaving behind a considerable amount of waste in the form of a mud composed mainly of water, lubricants and crushed rock. This waste with no defined destination accumulates in yards, reservoirs and streams, affecting the environment. Slate powder has great prospects for recovery, recycling and further applications once it has chemical composition and components similar to raw materials used in ceramic and building construction industries. Therefore, a complete characterization of this powder is important and here we present some results of XRD, SEM, EDS, G3 morphology and thermal analysis of slate powder samples from the southeastern state of Minas Gerais in Brazil.

Genetic disorders are historically defined through phenotype-first approaches. However, risk estimates derived from phenotype-linked ascertainment may overestimate severity and penetrance. NF1 is a large tumor suppressor gene and related disorders are characterized by several major features, age of onset, variability of severity and occurrence of complications. Pathogenic variants are associated with increased risks of rare and common neoplasms and a large array of clinical phenotypes in adults and children. This study compared two variant interpretation approaches: a publicly available population schematic and the clinical gold standard of the ACMG criteria. The population utilized 134,187 individuals who were ascertained to not have cancer in a cancer study from gnomAD (v2.1). A total of 103 pathogenic and 20 likely pathogenic short variants were identified in the population approach and 6 pathogenic and 13 likely pathogenic were identified through the ACMG criteria. Fifty pathogenic variants were identified to have been previously published with associated clinical phenotypes. Malignant tumors and hematologic malignancies such as rhabdomyosarcomas, malignant peripheral nerve sheath tumors, Watson Syndrome, Familial Spinal NF1, optic glioma and JMML were the most severe phenotypes identified. The recent inclusion of genetic testing in the clinical diagnosis of NF1 syndrome increases the need to fully understand the full scope of genotype-phenotype correlations associated with pathogenic NF1 variants across ethnicities. The ascertainment of individuals with germline pathogenic NF1 variants in this analysis highlights cases with potentially increased personal adult-onset cancer risk and unknown reproductive risk of hereditary juvenile-onset cancers.

Intra-lot and inter-lot variability in cosyntropin was detected in the DQS using Fourier transform near-infrared spectrometry. One vial of 12 sampled from Sandoz Lot0200062 appeared 6.9 SDs from the center of the rest of the vials, and another was 4.3 SDs away. Spectra of 102 vials from 7 lots in the library clustered in two groups (p=0.02).

Pemetrexed is a folate analog metabolic inhibitor used in treatment of locally advanced or metastatic nonsquamous non-small cell lung cancer and for mesothelioma. Intra-lot and inter-lot variability in the spectra of ALIMTA® was detected in the Drug Quality Study (DQS) using Fourier transform near-infrared spectrometry (FTNIR). One vial of 12 (8%) sampled from lot S20I013A appeared 3.0 multidimensional SDs from the other vials, suggesting that it represents a different material. Consequently, additional spectra from other lots were analyzed. Spectra of 147 vials from 23 lots in the spectral library contained 14 vials that were outside the main group (26.4 SDs using a subcluster detection test), suggesting that the 14 library vials (9.5% of the total) also contain differing materials.

The parasite Cryptosporidium is responsible for diarrheal disease in young children causing death, malnutrition, and growth delay. Cryptosporidium invades enterocytes where it develops in a unique intracellular niche. Infected cells exhibit profound changes in morphology, physiology, and transcriptional activity. How the parasite effects these changes is poorly understood. We explored the localization of highly polymorphic proteins and found members of the Cryptosporidium parvum MEDLE protein family to be translocated into the cytosol of infected cells. All intracellular life stages engage in this export, which occurs after completion of invasion. Mutational studies defined an N-terminal host-targeting motif and demonstrated proteolytic processing at a specific leucine residue. Direct expression of MEDLE2 in mammalian cells triggered an ER stress response, which was also observed during infection. Taken together, our studies reveal the presence of a Cryptosporidium secretion system capable of delivering parasite proteins into the infected enterocyte.