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Biallelic loss of cyclin-dependent kinase 12 (CDK12) defines a metastatic castration-resistant prostate cancer (mCRPC) subtype. It remains unclear, however, whether CDK12 loss drives prostate cancer (PCa) development or uncovers pharmacologic vulnerabilities. Here, we show Cdk12 ablation in murine prostate epithelium is sufficient to induce preneoplastic lesions with lymphocytic infiltration. In allograft-based CRISPR screening, Cdk12 loss associates positively with Trp53 inactivation but negatively with Pten inactivation. Moreover, concurrent Cdk12/Trp53 ablation promotes proliferation of prostate-derived organoids, while Cdk12 knockout in Pten-null mice abrogates prostate tumor growth. In syngeneic systems, Cdk12/Trp53-null allografts exhibit luminal morphology and immune checkpoint blockade sensitivity. Mechanistically, Cdk12 inactivation mediates genomic instability by inducing transcription-replication conflicts. Strikingly, CDK12-mutant organoids and patient-derived xenografts are sensitive to inhibition or degradation of the paralog kinase, CDK13. We therein establish CDK12 as a bona fide tumor suppressor, mechanistically define how CDK12 inactivation causes genomic instability, and advance a therapeutic strategy for CDK12-mutant mCRPC.

For over a century, early researchers sought to study biological organisms in a laboratory setting, leading to the generation of both in vitro and in vivo model systems. Patient-derived models of cancer (PDMCs) have more recently come to the forefront of preclinical cancer models and are even finding their way into clinical practice as part of functional precision medicine programs. The PDMC Consortium, supported by the Division of Cancer Biology in the National Cancer Institute of the National Institutes of Health, seeks to understand the biological principles that govern the various PDMC behaviors, particularly in response to perturbagens, such as cancer therapeutics. Based on collective experience from the consortium groups, we provide insight regarding PDMCs established both in vitro and in vivo, with a focus on practical matters related to developing and maintaining key cancer models through a series of vignettes. Although every model has the potential to offer valuable insights, the choice of the right model should be guided by the research question. However, recognizing the inherent constraints in each model is crucial. Our objective here is to delineate the strengths and limitations of each model as established by individual vignettes. Further advances in PDMCs and the development of novel model systems will enable us to better understand human biology and improve the study of human pathology in the lab.

Abstract Recent developments have broadened our perception of SARS-CoV-2, indicating its capability to affect the body systemically beyond its initial recognition as a mere respiratory pathogen. However, the pathways of its widespread are not well understood. Employing a dual-modality approach, we integrated findings from a Murine Hepatitis Virus (MHV) infection model with corroborative clinical data to investigate the pervasive reach of Coronaviruses. The novel presence of viral particles within red blood cells (RBCs) was demonstrated via high-resolution transmission electron microscopy, with computational modeling elucidating a potential heme-mediated viral entry mechanism via Spike protein affinity. Our data affirm viral localization in RBCs, suggesting heme moieties as facilitators for cellular invasion. Exacerbation of MHV pathology upon hemin administration, contrasted with chloroquine-mediated amelioration, underscoring a heme-centric pathway in disease progression. These observations extend the paradigm of Coronavirus pathogenicity to include hemoprotein interactions. This study casts new light on the systemic invasion capabilities of Coronaviruses, linking RBC hemoproteins with viral virulence. The modulation of disease severity through heme-interacting agents heralds a promising avenue for COVID-19 therapeutics. Our findings propose a paradigm shift in the treatment approach, leveraging the virus-heme interplay as a strategic hinge for intervention.

IntroductionGalvanic vestibular stimulation (GVS) is a simple, safe, and noninvasive method of neurostimulation that can be used to improve body balance. Several central nervous system diseases cause alterations in body balance, including HTLV-1-associated myelopathy (HAM).ObjectiveTo test GVS as a balance rehabilitation strategy for HAM.MethodsThis study is a quasi-experimental clinical trial in which postural balance was compared before and after a GVS rehabilitation protocol applied to 20 patients with HAM, 12 women and 8 men, average age of 78 and 79 years, respectively. They were followed for nine months after the end of the GVS protocol, which consisted of one GVS session per week for 12 consecutive weeks. The GVS current intensity was progressively increased from 1.0 milliamperes (mA) to 3.5 mA until the third session and maintained at 3.5 mA until the 12th session. The electrical stimulation time progressively increased from 9 min in the first session to 18 min in the second session and maintained at 30 min from the third session onwards. Postural balance was assessed by Time up and go test (TUG), Berg balance scale (BBS) and posturography that were performed before the beginning of the intervention, during the intervention (6th week), at the end of the intervention (12th week) and after 9 months of follow-up without electrical stimulation.ResultsIn a blind comparison, in the 12th week of stimulation, improvement was observed in all the tests. In TUG, time in seconds changed from 28 before to 18 after GVS (p

Summary: Stress can have profound consequences on mental health. While much is known about the neural circuits supporting associative memories of stressful events, our understanding of the circuits underlying the non-associative impacts of stress, such as heightened stress sensitivity and anxiety-related behavior, is limited. Here, we demonstrate that the ventral hippocampus (vHC) and basolateral amygdala (BLA) support distinct non-associative behavioral changes following stress. Inhibiting stress-induced protein synthesis in the BLA blocked subsequent increases in stress sensitivity but not anxiety-related behaviors. Conversely, inhibiting stress-induced protein synthesis in the vHC blocked subsequent increases in anxiety-related behavior but not stress sensitivity. Inhibiting neuronal activity in the BLA and vHC during the assessment of stress sensitivity or anxiety-related behavior recapitulated these structures’ dissociable contributions to defensive behavior. Lastly, blocking the associative memory of a stressor had no impact on stress-induced changes in anxiety-related behavior. These findings highlight that multiple memory systems support the long-lasting effects of stress.

Background: Safety and outcome of atezolizumab/bevacizumab in Child-Pugh B patients with hepatocellular carcinoma (HCC) have not been completely characterized. Objectives: In this study, we aimed at addressing safety and efficacy of atezolizumab/bevacizumab in Child-Pugh B patients by reviewing the available data and analyzing them by meta-analysis. Methods: We compared the safety and efficacy of atezolizumab/becavizumab treatment in patients with unresectable HCC and various degrees of liver dysfunction. A total of 8 retrospective, non-randomized, cohort studies were included in this meta-analysis, for a total of 1,071 Child-Pugh A and 225 Child-Pugh B patients. The albumin-bilirubin (ALBI) grade was also used to assess liver function, when available. Results: Grade ≥3 adverse events were observed in 11.8% of Child-Pugh class A and 26.8% class B patients (p = 0.0001), with an odds ratio (OR) of 0.43 (confidence interval [CI] 0.21–0.90; p = 0.02). Progression-free survival (PFS) at both 6 months (4.90 ± 2.08 vs. 4.75 ± 2.08 months; p = 0.0004) and 12 months (8.83 ± 2.32 vs. 7.26 ± 2.33 months; p = 0.002) was lower in Child-Pugh class B patients. A trend toward a higher objective response rate (ORR) was observed in Child-Pugh class A patients (219/856, 25.6%) as compared to Child-Pugh class B patients (25/138, 18.1%; p = 0.070), while the probability of obtaining an ORR was significantly greater in Child-Pugh A patients (OR 1.79, CI 1.12–2.86; p = 0.02). Median overall survival (OS) was 16.8 ± 2.0 and 6.8 ± 3.2 months in Child-Pugh A and B patients, respectively (mean difference 9.06 months, CI 7.01–11.1, p < 0.0001). Lastly, OS was longer in patients with ALBI grades 1–2 than in those with grade 3 (8.3 ± 11.4 vs. 3.3 ± 5.0 months, p = 0.0008). Conclusions: Oncological efficacy of atezolizumab/bevacizumab is moderate in Child-Pugh class B patients, and the shorter PFS and OS associated with the greater likelihood of experiencing treatment-related adverse events observed in these patients suggest great caution and individualization of treatment, possibly with the support of the ALBI grade.

The aging process is commonly accompanied by a general or specific loss of muscle mass, force and/or function that inevitably impact on a person’s quality of life. To date, various clinical tests and assessments are routinely performed to evaluate the biomechanical status of an individual, to support and inform the clinical management and decision-making process (e.g., to design a tailored rehabilitation program). However, these assessments (e.g., gait analysis or strength measures on a dynamometer) are typically conducted independently from one another or at different time points, providing clinicians with valuable yet fragmented information. We hereby describe a comprehensive protocol that combines both in vivo measurements (maximal voluntary isometric contraction test, superimposed neuromuscular electrical stimulation, electromyography, gait analysis, magnetic resonance imaging, and clinical measures) and in silico methods (musculoskeletal modeling and simulations) to enable the full characterization of an individual from the biomechanical standpoint. The protocol, which requires approximately 4 h and 30 min to be completed in all its parts, was tested on twenty healthy young participants and five elderlies, as a proof of concept. The implemented data processing and elaboration procedures allowing for the extraction of several biomechanical parameters (including muscle volumes and cross-sectional areas, muscle activation and co-contraction levels) are thoroughly described to enable replication. The main parameters extracted are reported as mean and standard deviation across the two populations, to highlight the potential of the proposed approach and show some preliminary findings (which were in agreement with previous literature).

Thrombotic thrombocytopenic purpura (TTP) is a potentially life-threatening, rare acute thrombotic microangiopathy (TMA), caused by a severe ADAMTS13 deficiency. As the COVID-19 pandemic rapidly spread around the globe, much data about the pathogenicity of this virus were published. Soon after the detection of the first cases of COVID-19, it was clear that there was a wide range of COVID coagulopathy manifestations, such as deep venous thrombosis, pulmonary thromboembolism, and thrombotic microangiopathies. In the literature, little data have been reported about the association between TTP and COVID-19, and the treatment of COVID-19-associated TTP is still under debate. Here we present the case of a 46-year-old woman who developed a COVID-associated TTP, successfully treated with plasma exchange (PEX), steroids, and caplacizumab.

Exhausted olive pomace (EOP) represents the principal residue of olive pomace. Several studies have optimized the extraction of specialized metabolites from the EOP of Olea europaea L., but a comparison between different extractive methods has not been made. For this reason, the present investigation aims to compare four different extractive methods by using water and 15% ethanol/water as extractive solvents. Specifically, based on extract antioxidant activity, the methods compared were maceration (MAC), microwave-assisted extraction (MAE), ultrasound-assisted extraction (UAE), and Accelerated Solvent Extraction (ASE). Between these, the UAE and ASE hydroalcoholic EOP extracts were demonstrated to have the highest antioxidant activity. Subsequently, these extracts were investigated for their hypoglycemic and antiradical activity using in vitro cell-free and cell-based assays, respectively. ASE hydroalcoholic EOP extract demonstrated the greatest ability to inhibit the α-amylase enzyme and an in vitro antioxidant activity comparable to N-acetyl cysteine in HepG2 cells. UAE and ASE extracts’ phytochemical characterization was also performed, identifying seven phenolic compounds, including 3-hydroxytyrosol, tyrosol, and, for the first time, salidroside. The ASE hydroalcoholic EOP extract was the richest from a phytochemical point of view, thus confirming its major biological activity. Therefore, ASE and 15% ethanol/water may represent the best extractive method for EOP nutraceutical valorization.

Background Cerebral changes occur in individuals with human T-cell leukemia virus type 1 (HTLV-1)-associated myelopathy (HAM) and seem to predominate in subcortical areas. Little is known about the cognitive decline in the elderly living with HTLV-1.

The aim of this study was to investigate balance performance and muscle synergies during a Single-Limb Stance (SLS) task in individuals with Chronic Ankle Instability (CAI) and a group of healthy controls. Twenty individuals with CAI and twenty healthy controls were asked to perform a 30-second SLS task in Open-Eyes (OE) and Closed-Eyes (CE) conditions while standing on a force platform with the injured or the dominant limb, respectively. The activation of 13 muscles of the lower limb, hip, and back was recorded by means of surface electromyography. Balance performance was assessed by identifying the number and the duration of SLS epochs, and the Root-Mean-Square (RMS) in Antero-Posterior (AP) and Medio-Lateral (ML) directions of the body-weight normalized ground reaction forces. The optimal number of synergies, weight vectors, and activation coefficients were also analyzed. CAI group showed a higher number and a shorter duration of SLS epochs and augmented ground reaction force RMS in both AP and ML directions compared to controls. Both groups showed an increase in the RMS in AP and ML forces in CE compared to OE. Both groups showed 4 optimal synergies in CE, while controls showed 5 synergies in OE. CAI showed a significantly higher weight of knee flexor muscles in both OE and CE. In conclusion, muscle synergies analysis provided an in-depth knowledge of motor control mechanisms in CAI individuals. They showed worse balance performance, a lower number of muscle synergies in a CE condition and abnormal knee flexor muscle activation compared to healthy controls.

IntroductionTreatment-free remission (TFR) in patients with chronic myeloid leukemia in chronic phase is considered a safe option if suitable molecular monitoring is available. However, the question arises as to which factors can contribute to the maintenance of TFR, and immunologic surveillance of the remaining leukemic cells is believed to be one of them. Argentina Stop Trial is an open-label, single-arm, multicenter trial assessing TFR after tyrosine kinase inhibitors interruption, that after more than 4 years showed a successful TFR rate of 63%.MethodsIn this context, we set up an immunological study by flow cytometry in order to analyze specific NK cell subsets from peripheral blood patient samples both at the time of discontinuation as well as during the subsequent months.ResultsAt the time of discontinuation, patients show a mature NK cell phenotype, probably associated to TKI treatment. However, 3 months after discontinuation, significant changes in several NK cell receptors occurred. Patients with a higher proportion of CD56dim NK and PD-1+ NK cells showed better chances of survival. More interestingly, non-relapsing patients also presented a subpopulation of NK cells with features associated with the expansion after cytomegalovirus infection (expression of CD57+NKG2C+), and higher proportion of NKp30 and NKp46 natural cytotoxicity receptors, which resulted in greater degranulation and associated with better survival (p

Macrophage-based co-cultures are used to test the immunomodulatory function of candidate cells for clinical use. This study aimed to characterize a macrophage polarization model using human platelet lysate (hPL) as a GMP-compliant alternative to Fetal Bovine Serum (FBS). Primary human monocytes were differentiated into unpolarized (M0) or polarized (M1, M2a, and M2c) macrophages in an hPL- or FBS-based medium. The protein secretion profiles and expression of phenotypic markers (CD80 for M1, CD206 for M2a, and CD163 for M2c) were analyzed. Subsequently, chondrocytes were tested in an hPL-based co-culture model to assess their immunomodulatory function in view of their possible use in patients with osteoarthritis. The results showed similar marker regulation between hPL and FBS cultures, but lower basal levels of CD206 and CD163 in hPL-cultured macrophages. Functional co-culture experiments with chondrocytes revealed increased CD206 expression both in hPL and in FBS, indicating an interaction between macrophages and chondrocytes. While markers in FBS-cultured macrophages were confirmed in hPL-cultured cells, the interpretation of marker modulation in immunomodulatory assays with hPL-based cultures should be carried out cautiously due to the observed differences in the basal marker levels for CD206 and CD163. This research underscores the utility of hPL as a GMP-compliant alternative to FBS for macrophage-based co-cultures and highlights the importance of understanding marker expressions in different culture conditions.

Abstract Background The efficacy of early treatment with convalescent plasma in patients with COVID-19 is debated. Nothing is known about the potential effect of other plasma components other than anti-SARS-CoV-2 antibodies. Methods To determine whether convalescent or standard plasma would improve outcomes for adults in early phase of Covid19 respiratory impairment we designed this randomized, three-arms, clinical trial (PLACO COVID) blinded on interventional arms that was conducted from June 2020 to August 2021. It was a multicentric trial at 19 Italian hospitals. We enrolled 180 hospitalized adult patients with COVID-19 pneumonia within 5 days from the onset of respiratory distress. Patients were randomly assigned in a 1:1:1 ratio to standard of care (n = 60) or standard of care + three units of standard plasma (n = 60) or standard of care + three units of high-titre convalescent plasma (n = 60) administered on days 1, 3, 5 after randomization. Primary outcome was 30-days mortality. Secondary outcomes were: incidence of mechanical ventilation or death at day 30, 6-month mortality, proportion of days with mechanical ventilation on total length of hospital stay, IgG anti-SARS-CoV-2 seroconversion, viral clearance from plasma and respiratory tract samples, and variations in Sequential Organ Failure Assessment score. The trial was analysed according to the intention-to-treat principle. Results 180 patients (133/180 [73.9%] males, mean age 66.6 years [IQR 57–73]) were enrolled a median of 8 days from onset of symptoms. At enrollment, 88.9% of patients showed moderate/severe respiratory failure. 30-days mortality was 20% in Control arm, 23% in Convalescent (risk ratio [RR] 1.13; 95% confidence interval [CI], 0.61–2.13, P = 0.694) and 25% in Standard plasma (RR 1.23; 95%CI, 0.63–2.37, P = 0.544). Time to viral clearance from respiratory tract was 21 days for Convalescent, 28 for Standard plasma and 23 in Control arm but differences were not statistically significant. No differences for other secondary endpoints were seen in the three arms. Serious adverse events were reported in 1.7%, 3.3% and 5% of patients in Control, Standard and Convalescent plasma arms respectively. Conclusions Neither high-titer Convalescent nor Standard plasma improve outcomes of COVID-19 patients with acute respiratory failure. Trial Registration Clinicaltrials.gov Identifier: NCT04428021. First posted: 11/06/2020

Background International studies performed during the periods of social isolation highlighted the potential loss of student's learning skills. The present study fills a gap in Brazilian research on this topic and focuses on the development of reading fluency. Objective To investigate the development of the reading fluency of students in the early years of elementary school during e-learning as a result of the social distancing measures put into effect due to the coronavirus disease 2019 (COVID-19) pandemic. Methods Students from grades 2 to 5 were recorded. The number of words read per minute and of those read correctly per minute were analyzed. Descriptive statistical analysis was performed, using analysis of variance (ANOVA) for repeated measures with Bonferroni correction in the longitudinal study, and the t-test in the cross-sectional study. Results In the cross-sectional study, 162 students participated. Only the comparison between the 2nd grade classes of 2020 and 2021 showed a statistically significant difference. In the prepandemic classes, the students had better results in reading accuracy than the students assessed during the pandemic. The longitudinal study included 75 students, who improved in fluency rate and accuracy as expected between March and December 2020. In March 2021, the results showed a drop, which may be related to school closures during the Brazilian summer vacation. Conclusions The present research demonstrates the results of Brazilian students in terms of the development of reading fluency during the pandemic. There was an expressive development in grades 2 and 3, with stability in the following grades. The 2nd grade class of 2021 suffered a major impact due to the pandemic.

El examen clínico objetivo estructurado (ECOE) es una herramienta válida para medir competencias clínicas. En el contexto de la pandemia por COVID-19 se debieron adaptar las evaluaciones, habitualmente presenciales, a un formato virtual. El objetivo de este artículo es comunicar la experiencia de utilizar una modalidad virtual del ECOE para residentes de primer año de pediatría durante la pandemia por COVID-19. El ECOE comprendió 12 estaciones utilizando casos simulados en línea y consultas virtuales a distancia. Participaron del ECOE 115 residentes, aprobaron 110 (95,7%) y solo 17 participantes aprobaron el 100% de las estaciones. Las estaciones con mejor rendimiento fueron las referidas a alimentación complementaria, abordaje del traumatismo encéfalo craneano, interpretación de resultados de laboratorio y de imágenes radiológicas. El menor rendimiento se observó en uso de equipo de protección personal y vacunas. La encuesta de satisfacción fue respondida por 80/115 participantes (69,5%). El 91% de los participantes encontró fácil de utilizar la plataforma virtual. La mayoría (73/80) consideró que los temas fueron acordes al nivel de formación. La estación de pautas madurativas fue identificada como la más difícil. El 57% refirió que el ECOE les resultó útil para identificar puntos débiles en su formación. La implementación de una modalidad virtual del ECOE fue posible y presentó buena aceptación de los participantes.

ABSTRACT Galvanic vestibular stimulation (GVS) influences body balance and has proved to be useful to improve patients’ mood, quality of life, and cognitive skills. This study aimed to present three cases of patients with Parkinson’s disease and postural instability who had been submitted to GVS to improve their balance, by assessing the impact of this intervention on their cognition, mood, and quality of life. Patients were assessed before and after GVS sessions concerning P300 latency and scores on the 15-item Geriatric Depression Scale (GDS-15) and the 39-item quality-of-life Parkinson’s Disease Questionnaire (PDQ-39). The three patients’ P300 latency improved, possibly indicating improved attention. Their PDQ-39 score also improved, possibly indicating a positive impact on their quality of life. Their GDS-15 score did not change before and after the intervention. None of the patients had any intervention side effects. This three-case experimental pilot study has shown that GVS is a safe method, possibly useful to improve attention and, therefore, the quality of life of patients presented with Parkinson’s disease.