SCN as a local probe of protein structural dynamics.

The dynamics of lysozyme is probed by attaching -SCN to all alanine residues. The one-dimensional infrared spectra exhibit frequency shifts in the position of the maximum absorption of 4 cm-1, which is consistent with experiments in different solvents and indicates moderately strong interactions of the vibrational probe with its environment. Isotopic substitution 12C → 13C leads to a redshift by -47 cm-1, which agrees quantitatively with experiments for CN-substituted copper complexes in solution. The low-frequency, far-infrared part of the protein spectra contains label-specific information in the difference spectra when compared with the wild type protein. Depending on the position of the labels, local structural changes are observed. For example, introducing the -SCN label at Ala129 leads to breaking of the α-helical structure with concomitant change in the far-infrared spectrum. Finally, changes in the local hydration of SCN-labeled alanine residues as a function of time can be related to the reorientation of the label. It is concluded that -SCN is potentially useful for probing protein dynamics, both in the high-frequency part (CN-stretch) and in the far-infrared part of the spectrum.

Effectiveness and safety of tenofovir amibufenamide in chronic hepatitis B patients.

This letter to the editor relates to the study entitled "Tenofovir amibufenamide vs tenofovir alafenamide for treating chronic hepatitis B: A real-world study", which was recently published by Peng et al. Hepatitis B virus infection represents a significant health burden worldwide and can lead to cirrhosis and even liver cancer. The antiviral drugs currently used to treat patients with chronic hepatitis B infection still have many side effects, so it is crucial to identify safe and effective drugs to inhibit viral replication.

Real world community-based HIV Rapid Start Antiretroviral with B/F/TAF versus prior models of antiretroviral therapy start - the RoCHaCHa study, a pilot study.

BACKGROUND: The rapid start of antiretroviral therapy (RSA) model initiates antiretroviral therapy (ART) as soon as possible after a new or preliminary diagnosis of HIV, in advance of HIV-1 RNA and other baseline laboratory testing. This observational study aims to determine if RSA with a single tablet regimen of bictegravir, emtricitabine, and tenofovir alafenamide (B/F/TAF) is an effective regimen for achieving viral suppression and accepted by patients at the time of diagnosis. METHODS: Adults newly or preliminarily diagnosed with HIV were enrolled from October 2018 through September 2021. Real world advantage, measured in days between clinical milestones and time to virologic suppression, associated with B/F/TAF RSA was compared to historical controls. RESULTS: All Study RSA participants (n = 45) accepted treatment at their first visit and 43(95.6%) achieved virologic suppression by week 48. Study RSA participants had a significantly shorter time (median 32 days) from diagnosis to ART initiation and virologic suppression, in comparison to historical controls (median 181 days) (n = 42). Qualitative feedback from study RSA participants showed high acceptance positive response to RSA. CONCLUSIONS: RSA is feasible and well accepted by patients in a real-world community-based clinic setting. Promoting RSA in community-based clinics is an important tool in ending the HIV epidemic.

Origin and Roles of Alanine and Glutamine in Gluconeogenesis in the Liver, Kidneys, and Small Intestine under Physiological and Pathological Conditions.

Alanine and glutamine are the principal glucogenic amino acids. Most originate from muscles, where branched-chain amino acids (valine, leucine, and isoleucine) are nitrogen donors and, under exceptional circumstances, a source of carbons for glutamate synthesis. Glutamate is a nitrogen source for alanine synthesis from pyruvate and a substrate for glutamine synthesis by glutamine synthetase. The following differences between alanine and glutamine, which can play a role in their use in gluconeogenesis, are shown: (i) glutamine appearance in circulation is higher than that of alanine; (ii) the conversion to oxaloacetate, the starting substance for glucose synthesis, is an ATP-consuming reaction for alanine, which is energetically beneficial for glutamine; (iii) most alanine carbons, but not glutamine carbons, originate from glucose; and (iv) glutamine acts a substrate for gluconeogenesis in the liver, kidneys, and intestine, whereas alanine does so only in the liver. Alanine plays a significant role during early starvation, exposure to high-fat and high-protein diets, and diabetes. Glutamine plays a dominant role in gluconeogenesis in prolonged starvation, acidosis, liver cirrhosis, and severe illnesses like sepsis and acts as a substrate for alanine synthesis in the small intestine. Interactions among muscles and the liver, kidneys, and intestine ensuring optimal alanine and glutamine supply for gluconeogenesis are suggested.

Baricitinib and Pulse Steroids Combination Treatment in Hyperinflammatory COVID-19: A Rheumatological Approach in the Intensive Care Unit.

Hyperinflammatory Coronavirus disease 2019 (COVID-19) and rapidly-progressive interstitial lung diseases (RP-ILD) secondary to inflammatory myopathies (IIM) present important similarities. These data support the use of anti-rheumatic drugs for the treatment of COVID-19. The aim of this study was to compare the efficacy of combining baricitinib and pulse steroids with the Standard of Care (SoC) for the treatment of critically ill COVID-19 patients. We retrospectively enrolled consecutive patients admitted to the Intensive Care Unit (ICU) with COVID-19-pneumonia. Patients treated with SoC (dexamethasone plus remdesivir) were compared to patients treated with baricitinib plus 6-methylprednisolone pulses (Rheuma-group). We enrolled 246 patients: 104/246 in the SoC and 142/246 in the Rheuma-group. All patients presented laboratory findings suggestive of hyperinflammatory response. Sixty-four patients (26.1%) died during ICU hospitalization. The mortality rate in the Rheuma-group was significantly lower than in the SoC-group (15.5 vs. 40.4%, p < 0.001). Compared to the SoC-group, patients in the Rheuma-group presented significantly lower inflammatory biomarker levels after one week of treatment. Higher ferritin levels after one week of treatment were strongly associated with mortality (p < 0.001). In this large real-life COVID-19 cohort, baricitinib and pulse steroids led to a significant reduction in mortality, paralleled by a prompt reduction in inflammatory biomarkers. Our experience supports the similarities between hyperinflammatory COVID-19 and the IIM-associated RP-ILD.

Mechanism of Abnormal Activation of MEK1 Induced by Dehydroalanine Modification.

Mitogen-activated protein kinase kinase 1 (MAPK kinase 1, MEK1) is a key kinase in the mitogen-activated protein kinase (MAPK) signaling pathway. MEK1 mutations have been reported to lead to abnormal activation that is closely related to the malignant growth and spread of various tumors, making it an important target for cancer treatment. Targeting MEK1, four small-molecular drugs have been approved by the FDA, including Trametinib, Cobimetinib, Binimetinib, and Selumetinib. Recently, a study showed that modification with dehydroalanine (Dha) can also lead to abnormal activation of MEK1, which has the potential to promote tumor development. In this study, we used molecular dynamics simulations and metadynamics to explore the mechanism of abnormal activation of MEK1 caused by the Dha modification and predicted the inhibitory effects of four FDA-approved MEK1 inhibitors on the Dha-modified MEK1. The results showed that the mechanism of abnormal activation of MEK1 caused by the Dha modification is due to the movement of the active segment, which opens the active pocket and exposes the catalytic site, leading to sustained abnormal activation of MEK1. Among four FDA-approved inhibitors, only Selumetinib clearly blocks the active site by changing the secondary structure of the active segment from α-helix to disordered loop. Our study will help to explain the mechanism of abnormal activation of MEK1 caused by the Dha modification and provide clues for the development of corresponding inhibitors.

DA-9601 has protective effects comparable to those of proton pump inhibitor and rebamipide against nonsteroidal anti-inflammatory drugs-induced upper and lower gastrointestinal bleeding in patients with rheumatoid arthritis: A nationwide study using Korean Health Insurance Review and Assessment Service database.

DA-9601 extracted from Artemisia asiatica contains a bioactive compound - eupatilin - that can protect against gastric mucosal damage through anti-inflammatory and anti-oxidative properties and is approved for treating acute and chronic gastritis in Korea, but their ability to protect gastrointestinal (GI) bleeding caused by nonsteroidal anti-inflammatory drugs (NSAIDs) is unclear. We aimed to compare the protective effects of DA-9601 to those of proton pump inhibitors (PPI) and rebamipide against upper and lower GI bleeding in patients with rheumatoid arthritis (RA) undergoing long-term NSAIDs therapy using the Korean Health Insurance Review and Assessment database. In this nationwide retrospective cohort study, we evaluated patients with RA who concurrently received NSAIDs for >3 months with DA-9601, PPI, or rebamipide between January 2015 and December 2017. The index date was the date of NSAIDs initiation, and all patients were followed up until December 2020 to detect upper and lower GI bleeding. In total, 24,258 patients with RA were eligible, and 5468 (22.5%), 4417 (18.2%), and 14,373 (59.3%) received DA-9601, PPI, or rebamipide, respectively, on the index date. During follow-up, upper and lower GI bleeding occurred in 508 (2.1%) and 402 (1.6%) patients with RA, respectively. The incidence rate of upper and lower GI bleeding was 615/100,000 and 485/100,000 person-years, respectively. Among patients with RA receiving DA-9601, PPI, or rebamipide, the frequencies of NSAIDs-induced upper GI bleeding were 0.5%, 0.4%, and 1.2%, respectively. The frequencies of NSAIDs-induced lower GI bleeding were 0.4%, 0.4%, and 0.9%, respectively. The incidence of NSAIDs-induced upper GI bleeding in patients with RA receiving DA-9601, PPI, and rebamipide was 601/100,000, 705/100,000, and 596/100,000 person-years, respectively, while the incidence of NSAIDs-induced lower GI bleeding in the same groups was 449/100,000, 608/100,000, and 465/100,000 person-years, respectively. In the multivariate Cox regression analysis, no significant difference was observed in lower and upper GI bleeding hazards between patients with RA using DA-9601, PPI, and rebamipide. Our results suggest that DA-9601 may exhibit protection against NSAIDs-induced GI bleeding that is comparable to those of PPI and rebamipide in patients with RA.

Alanine and glutamate catabolism collaborate to ensure the success of Bacillus subtilis sporulation.

Sporulation as a typical bacterial differentiation process has been studied for decades. However, two crucial aspects of sporulation, (i) the energy sources supporting the process, and (ii) the maintenance of spore dormancy throughout sporulation, are scarcely explored. Here, we reported the crucial role of RocG-mediated glutamate catabolism in regulating mother cell lysis, a critical step for sporulation completion of Bacillus subtilis, likely by providing energy metabolite ATP. Notably, rocG overexpression resulted in an excessive ATP accumulation in sporulating cells, leading to adverse effects on future spore properties, e.g. increased germination efficiency, reduced DPA content, and lowered heat resistance. Additionally, we revealed that Ald-mediated alanine metabolism was highly related to the inhibition of premature germination and the maintenance of spore dormancy during sporulation, which might be achieved by decreasing the typical germinant L-alanine concentration in sporulating environment. Our data inferred that sporulation of B. subtilis was a highly orchestrated biological process requiring a delicate balance in diverse metabolic pathways, hence ensuring both the completion of sporulation and production of high-quality spores.

Blood pressure increases are associated with weight gain and not antiretroviral regimen or kidney function: a secondary analysis from the ADVANCE trial in South Africa.

INTRODUCTION: Recent evidence has raised questions about whether newer HIV treatment regimens, including dolutegravir (DTG) and tenofovir alafenamide (TAF), are associated with increases in blood pressure (BP). METHODS: We assessed changes in BP by treatment regimen and evaluated the relative contribution of kidney function and weight gain to these changes among participants in the ADVANCE phase-3 trial clinical trial in South Africa (study dates: January 2017-February 2022). Our primary outcome of interest was a change in systolic BP (SBP) at 96 and 192 weeks, among those not receiving antihypertensive medication. The secondary outcome was treatment-emergent hypertension at these same time points, defined as BP ≥140/90 mmHg on two occasions, or initiation of antihypertensive medication after week 4 among individuals without hypertension at enrolment. We used linear regression to evaluate the relationship between change in estimated glomerular filtration rate (eGFR) and change in SBP; and Poisson regression to evaluate the relationship between change in eGFR and treatment-emergent hypertension at each time point. All models were adjusted for age, sex, treatment group and change in body mass index (BMI). RESULTS: Over 96 weeks, the average changes in SBP were 1.7 mmHg (95% CI: 0.0-3.4), -0.5 mmHg (95% CI: -2.2 to 1.7) and -2.1 mmHg (95% CI: -3.8 to 0.4) in the TAF/emtricitabine (FTC)/DTG, tenofovir disoproxil fumarate (TDF)/FTC/DTG and TDF/FTC/efavirenz (EFV) groups, respectively. This difference was significant for the TAF/FTC/DTG compared to the TDF/FTC/EFV group (p = 0.002). Over 96 weeks, 18.2% (95% CI: 13.4-22.9), 15.4% (95% CI: 11.0-19.9) and 13.3% (95% CI: 8.9-17.6) of participants developed treatment-emergent hypertension, respectively. In adjusted models, there was no significant relationship between change in eGFR and either outcome. Change in BMI was significantly associated with an increase in SBP, while age was associated with an increased risk of treatment-emergent hypertension. Adjustment for BMI also mitigated the unadjusted relationship between HIV treatment regimen and SBP where present. CONCLUSIONS: In the ADVANCE cohort, weight gain and age accounted for increases in BP and risk of treatment-emergent hypertension. HIV treatment programmes may need to integrate the management of obesity and hypertension into routine care. CLINICAL TRIAL NUMBER: NCT03122262.

Early progression during or after cyclophosphamide, doxorubicin, vincristine, and prednisone chemotherapy indicates poor outcome with rescue protocols in dogs with multicentric lymphoma.

BACKGROUND: Dogs with lymphoma that fail cyclophosphamide, doxorubicin, vincristine, and prednisone chemotherapy (CHOP) before completion of their protocol are commonly thought to have poor long-term outcome, but no previous studies have evaluated the effect of early relapse on progression-free interval (PFI) or overall survival time (OST) for patients undergoing rescue chemotherapy. OBJECTIVE: Correlate rescue treatment outcomes in dogs with multicentric lymphoma with outcomes after 1st-line CHOP chemotherapy. METHODS: Data were collected from 6 previous retrospective or prospective studies in 187 dogs with multicentric lymphoma that received 1st-line CHOP chemotherapy and then received either lomustine (CCNU), L-asparaginase and prednisone (LAP), or rabacfosadine (RAB, Tanovea), with or without prednisone or L-asparaginase. RESULTS: The PFI after initiation of CHOP chemotherapy was significantly associated with response rate postprogression, PFI, and postrescue survival time (ST) for both rescue protocols. Immunophenotype (B- vs T-cell) was not significantly associated with response, PFI or OST for LAP but was significantly associated with response and PFI for RAB. CONCLUSION: Dogs that experience short PFI during or after 1st-line CHOP chemotherapy had lower response rates to rescue treatment, with shorter PFI and ST. Immunophenotype did not significantly affect outcome with LAP but was associated with PFI for RAB.

Remdesivir alleviates skin fibrosis by suppressing TGF-ß1 signaling pathway.

Fibrotic skin diseases, such as keloids, are pathological results of aberrant tissue healing and are characterized by overgrowth of dermal fibroblasts. Remdesivir (RD), an antiviral drug, has been reported to have pharmacological activities in a wide range of fibrotic diseases. However, whether RD function on skin fibrosis remains unclear. Therefore, in our study, we explored the potential effect and mechanisms of RD on skin fibrosis both in vivo and in vitro. As expected, the results demonstrated that RD alleviated BLM-induced skin fibrosis and attenuates the gross weight of keloid tissues in vivo. Further studies suggested that RD suppressed fibroblast activation and autophagy both in vivo and in vitro. In addition, mechanistic research showed that RD attenuated fibroblasts activation by the TGF-ß1/Smad signaling pathway and inhibited fibroblasts autophagy by the PI3K/Akt/mTOR signaling pathway. In summary, our results demonstrate therapeutic potential of RD for skin fibrosis in the future.

Relationship between adherence to bictegravir/emtricitabine/tenofovir alafenamide fumarate and clinical outcomes in people with HIV in Japan: a claims database analysis.

A lack of adherence to long-term antiretroviral therapy may impact viral suppression. The current study examined the relationship between medication adherence and clinical outcomes in people with human immunodeficiency virus infection (PWH) receiving bictegravir, emtricitabine, and tenofovir alafenamide fumarate (B/F/TAF). A retrospective cohort study using two Japanese claims databases was conducted. Adherence was measured by the proportion of days covered (PDC). Patients were grouped into 3 PDC category and persistence was estimated by Kaplan-Meier method. Cox regression analysis was performed to investigate whether the PDC was associated with treatment discontinuation. Among 952 patients, 820 (86.1%), 95 (10.0%), and 37 (3.9%) patients were grouped into the PDC ≥ 90%, 80- < 90%, and < 80% groups, respectively. Across all PDC groups, more than 90% of patients who received B/F/TAF were receiving treatment at 1 year. There was no significant difference in the risk of discontinuation between the lower PDC groups (80- < 90% and < 80%) and the PDC ≥ 90% group (0.400 [0.096, 1.661]; 2.244 [0.663, 7.594], hazard ratio [95% confidence interval], respectively). A drug resistance test was implemented for 15 patients, none of whom discontinued B/F/TAF after the test. The results suggest that events that could cause discontinuation, such as virologic failure, were not associated with PDC.

Exploring How System Dimensions and Periodic Boundary Conditions Influence the Molecular Dynamics Simulation of A6H Peptide Self-Assembly Nanostructures.

This work presents a study on the effects of periodic boundary conditions (PBC) on the energetic/structural properties and hydrogen bond dynamics (HB) using molecular dynamics (MD) simulations of peptide membranes composed of alanine and histidine. Our results highlight that simulations using small surface areas for the peptide membrane may result in nonconvergent values for membrane properties, which are only observed in regions simulated at a certain distance from the PBCs. Specifically, regarding hydrogen bonds, a property pervasive in peptide membranes, our findings indicate a significant increase in the lifetime of these interactions, reaching values ∼19% higher when observed in structures free from PBCs. For peptide mobility in these nanomembranes, our results compare regions simulated directly under the influence of PBCs with regions free from these conditions, emphasizing greater mobility of amino acid psi/phi angles in the latter model.

The effect of early versus late remdesivir treatment in hospitalized mild to moderate COVID-19 patients in the Omicron era: A retrospective study.

Although real-world studies have found that remdesivir is effective in preventing poor prognosis, more information is needed on the optimal timing of remdesivir administration in high-risk coronavirus disease 2019 (COVID-19) patients in the Omicron era. From February 2022 to January 2023, a single-center retrospective study was performed in Korea. We compared the clinical characteristics and treatment outcomes between early (remdesivir treatment within 0-3 days from symptom onset) and late (≥ 4 days from symptom onset) treatment groups of patients who received remdesivir monotherapy. Of 284 patients, 225 were classified into the early treatment group and 59 were classified into the late treatment group. The early treatment group had a lower rate of 28-day progression to severe disease than the late treatment group (1.4% vs 7.4%, P = .03). Delaying remdesivir treatment ≥ 4 days from symptom onset (adjusted odds ratio [aOR], 6.17; 95% CI, 1.18-32.44; P = .03) and Charlson comorbidity index ≥ 3 (aOR, 9.62; 95% CI, 1.65-56.10; P = .01) were independent risk factors for 28-day progression to severe disease. Our results suggest that early administration of remdesivir could be associated with better prognosis in COVID-19 patients with the Omicron variant, and within 3 days from symptom onset seems to be the appropriate timing.

Alanine, a potential amino acid biomarker of pediatric sepsis: a pilot study in PICU.

Sepsis is characterized by a metabolic disorder of amino acid occurs in the early stage; however, the profile of serum amino acids and their alterations associated with the onset of sepsis remain unclear. Thus, our objective is to identify the specific kinds of amino acids as diagnostic biomarkers in pediatric patients with sepsis. Serum samples were collected from patients with sepsis admitted to the pediatric intensive care unit (PICU) between January 2019 and December 2019 on the 1st, 3rd and 7th day following admission. Demographic and laboratory variables were also retrieved from the medical records specified times. Serum amino acid concentrations were detected by UPLC-MS/MS system. PLS-DA (VIP > 1.0) and Kruskal-Wallis test (p < 0.05) were employed to identify potential biomarkers. Spearman's rank correlation analysis was conducted to find the potential association between amino acid levels and clinical features. The diagnostic utility for pediatric sepsis was assessed using receiver operating characteristic (ROC) curve analysis. Most of amino acid contents in serum were significantly decreased in patients with sepsis, but approached normal levels by the seventh day post-diagnosis. Threonine (THR), lysine (LYS), valine (VAL) and alanine (ALA) emerged as potential biomarkers related for sepsis occurrence, though they were not associated with PELOD/PELOD-2 scores. Moreover, alterations in serum THR, LYS and ALA were linked to complications of brain injury, and serum ALA levels were also related to sepsis-associated acute kidney injury. Further analysis revealed that ALA was significantly correlated with the Glasgow score, serum lactate and glucose levels, C-reactive protein (CRP), and other indicators for liver or kidney dysfunction. Notably, the area under the ROC curve (AUC) for ALA in distinguishing sepsis from healthy controls was 0.977 (95% CI: 0.925-1.000). The serum amino acid profile of children with sepsis is significantly altered compared to that of healthy controls. Notably, ALA shows promise as a potential biomarker for the early diagnosis in septic children.

Attenuated replication and damaging effects of SARS-CoV-2 Omicron variants in an intestinal epithelial barrier model.

Many COVID-19 patients suffer from gastrointestinal symptoms and impaired intestinal barrier function is thought to play a key role in Long COVID. Despite its importance, the impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on intestinal epithelia is poorly understood. To address this, we established an intestinal barrier model integrating epithelial Caco-2 cells, mucus-secreting HT29 cells and Raji cells. This gut epithelial model allows efficient differentiation of Caco-2 cells into microfold-like cells, faithfully mimics intestinal barrier function, and is highly permissive to SARS-CoV-2 infection. Early strains of SARS-CoV-2 and the Delta variant replicated with high efficiency, severely disrupted barrier function, and depleted tight junction proteins, such as claudin-1, occludin, and ZO-1. In comparison, Omicron subvariants also depleted ZO-1 from tight junctions but had fewer damaging effects on mucosal integrity and barrier function. Remdesivir, the fusion inhibitor EK1 and the transmembrane serine protease 2 inhibitor Camostat inhibited SARS-CoV-2 replication and thus epithelial barrier damage, while the Cathepsin inhibitor E64d was ineffective. Our results support that SARS-CoV-2 disrupts intestinal barrier function but further suggest that circulating Omicron variants are less damaging than earlier viral strains.

Remdesivir-Related Cardiac Adverse Effects in COVID-19 Patients: A Case-Control Study.

BACKGROUND: There have been reports of serious side effects of Remdesivir, including cardiovascular complications. The present study aimed to determine the adverse cardiovascular effects of Remdesivir and the factors affecting them in COVID-19 patients. METHODS: The patients were classified into two groups: those receiving Remdesivir without cardiac complications and those receiving Remdesivir with cardiovascular complications. After reviewing the patient's medical records, the relationship of some factors with the incidence of adverse cardiovascular effects was measured. RESULTS: Chi-square test showed that the distribution of complications in men was significantly higher than in women (P=0.001). The independent t-test revealed that the mean age in the group with complications was significantly higher than the group without complications (P=0.013). Fisher's exact test demonstrated a significant relationship between smoking and cardiovascular complications (P=0.05). According to the Mann-Whitney test, a significant difference was found in the mean changes of Bilirubin (P=0.02) and ALKP (P=0.01) before and after treatment in the groups with and without heart complications. CONCLUSION: Our findings indicated that most of the COVID-19 patients suffered from sinus bradycardia, and the distribution of complications was more pronounced in men than in women. The mean age in the group with complications was higher than the group without complications. Smoking was found to be associated with the occurrence of cardiovascular complications and the mean changes of Bilirubin and ALKP before and after treatment were significantly different in the groups with and without cardiovascular complications.

HIV-1 infection kinetics, drug resistance, and long-term safety of pre-exposure prophylaxis with emtricitabine plus tenofovir alafenamide (DISCOVER): week 144 open-label extension of a randomised, controlled, phase 3 trial.

BACKGROUND: Data characterising the long-term use and safety of emtricitabine plus tenofovir disoproxil fumarate as daily oral pre-exposure prophylaxis (PrEP) are scarce and there are uncertainties regarding the value of routine HIV-1 RNA testing during oral PrEP follow-up. METHODS: The DISCOVER trial was a randomised, controlled, phase 3 trial in which cisgender men and transgender women aged 18 years and older with a high likelihood of acquiring HIV were recruited from 94 clinics in Europe and North America and randomly assigned to receive either emtricitabine plus tenofovir disoproxil fumarate (200/25 mg) tablets daily, with matched placebo tablets, or emtricitabine plus tenofovir alafenamide (200/300 mg) tablets daily, with matched placebo tablets, for at least 96 weeks. After completion of the trial, participants were offered enrolment in this 48-week open-label extension study of emtricitabine plus tenofovir alafenamide. In participants diagnosed with HIV during the randomised and open-label phases of the study, we characterised HIV-1 test results and measured HIV-1 RNA viral load retrospectively when available. Adherence based on tenofovir diphosphate concentrations in dried blood spots and genotypic resistance were assessed in participants diagnosed with HIV. Safety assessments included adverse events, laboratory parameters, and, in a subset of participants, bone mineral density. HIV-1 incidence in participants initially randomly assigned to receive emtricitabine plus tenofovir alafenamide was estimated using a Poisson distribution. Changes from baseline in safety endpoints were described in participants assigned to received emtricitabine plus tenofovir alafenamide and in those who switched from emtricitabine plus tenofovir disoproxil fumarate during the open-label phase. This trial is registered with ClinicalTrials.gov, NCT02842086, and is ongoing. FINDINGS: Between Sept 13, 2016, and June 30, 2017, 5399 participants were enrolled and randomly assigned in DISCOVER. 2699 were assigned to receive emtricitabine plus tenofovir disoproxil fumarate and 2700 were assigned to receive emtricitabine plus tenofovir alafenamide, of whom 2693 and 2694, respectively, received at least one dose of study drug. 2115 (79%) assigned to emtricitabine plus tenofovir disoproxil fumarate switched to emtricitabine plus tenofovir alafenamide in the open-label phase, and 2070 (77%) continued with emtricitabine plus tenofovir alafenamide in the open-label phase. As of data cutoff (Dec 10, 2020), after 15 817 person-years of follow-up, 27 new HIV-1 diagnoses were observed across the total study period, with three occurring during the open-label phase. In participants who were initially assigned to emtricitabine plus tenofovir alafenamide, the incidence was 0·13 per 100 person-years (95% CI 0·061-0·23; ten of 2670). Stored plasma samples were available for 23 of 27 participants, including 22 with incident infection. In four (17%) of 23 participants, retrospective testing detected HIV-1 RNA before serological HIV-1 test positivity; one was a suspected baseline infection. Of the three incident cases, all three were non-adherent to PrEP and none developed drug resistance. Among participants taking emtricitabine plus tenofovir alafenamide for up to 144 weeks, markers of glomerular filtration and proximal renal tubule dysfunction (ß2-microglobulin to creatinine ratio and retinol-binding protein to creatinine ratio) improved or remained stable at 144 weeks compared with baseline, bone mineral density in hip and lumbar spine increased or remained stable from baseline to week 144 (n=191), cholesterol and glucose concentrations remained stable, and median bodyweight increased by less than 1 kg per year. In participants who switched from emtricitabine plus tenofovir disoproxil fumarate during the open-label phase (2115 [79%] of 2693), markers of glomerular filtration and proximal renal tubule dysfunction improved or remained stable, bone mineral density increased, cholesterol concentrations increased, glucose concentrations were similar, and median bodyweight increased more compared with those who remained on emtricitabine and tenofovir alafenamide. INTERPRETATION: Routine HIV-1 RNA testing for follow-up of individuals on daily oral PrEP provides modest additional clinical benefit. Long-term use of emtricitabine and tenofovir alafenamide as daily oral PrEP is safe and well tolerated and can be an especially appropriate choice for people with bone or renal morbidities. FUNDING: Gilead Sciences.

Ecofriendly micellar mediated spectrofluorimetric method for ultrasensitive quantification of the antiparkinsonian drug safinamide in pharmaceutical formulation and spiked human plasma.

A novel, highly sensitive and eco-friendly micellar-mediated spectrofluorimetric method was developed and validated for the determination of the novel antiparkinsonian drug safinamide mesylate in the presence of its related precursor impurity, 4-hydroxybenzaldehyde. The proposed approach relies on increasing the inherent fluorescence emission at 296 nm of safinamide, by forming hydrogen bonds between the mentioned drug and sodium dodecyl sulfate in the micellar system using 0.1 N HCl as a solvent, following excitation at 226 nm. A thorough investigation was conducted into the experimental factors affecting spectrofluorimetric behavior of the studied drug. A linearity plot of safinamide over the concentration range of 10.0-1000.0 ng/mL against the relative fluorescence intensities was established. The proposed method demonstrated excellent sensitivity down to the nano-gram level with detection and quantitation limits of 1.91 and 5.79 ng/mL, respectively. The studied drug was effectively determined in Parkimedine® Tablets. Furthermore, the proposed method allows for ultrasensitive quantification of safinamide in spiked human plasma, with satisfactory percentage recovery (98.97-102.28%). Additionally, the greenness assessment using the advanced green certificate classification approach, the complementary green analytical procedure index (Complex-GAPI), and the analytical GREEness metric approach (AGREE), along with the practicality check using the Blue Applicability Grade Index in addition to the all-inclusive overall whiteness evaluation using the RGB-12 model were carried out. The outcomes demonstrated the effectiveness and whiteness of the proposed technique. Clearly, the suggested approach has the advantages of being simple, requiring no pretreatment steps, and relying solely on direct measuring procedures.

Versatile functionalization of Bifidobacteria-derived extracellular vesicles using amino acid metabolic labeling and click chemistry for immunotherapy.

Extracellular vesicles (EVs) are nanoparticles secreted by various organisms. Methods for modifying EVs functionally have garnered attention for developing EV-based therapeutic systems. However, most technologies used to integrate these functions are limited to mammalian-derived EVs and a promising modification method for bacteria-derived EVs has not yet been developed. In this study, we propose a novel method for the versatile functionalization of immunostimulatory probiotic Bifidobacteria-derived EVs (B-EVs) using amino acid metabolic labeling and azide-alkyne click reaction. Azide D-alanine (ADA), a similar molecule to D-alanine in bacteria cell-wall peptidoglycan, was selected as an azide group-functionalized amino acid. Azide-modified B-EVs were isolated from Bifidobacteria incubated with ADA. The physicochemical and compositional characteristics, as well as adjuvanticity of B-EVs against immune cells were not affected by azide loading, demonstrating that this functionalization approach can retain the endogenous usefulness of B-EVs. By using the fluorescent B-EVs obtained by this method, the intracellular trafficking of B-EVs after uptake by immune cells was successfully observed. Furthermore, this method enabled the formulation of B-EVs for hydrogelation and enhanced adjuvanticity in the host. Our findings will be helpful for further development of EV-based immunotherapy.