Physiological conduction system pacing has attracted attention to overcome the dyssynchrony problems of conventional right ventricular pacing (RVP). Left bundle branch area pacing (LBBAP), which complements short combing of His bundle pacing (HBP), has emerged and has proven its efficiency and safety. In addition, initial experiences of LBBAP were mainly using lumen-less pacing lead, and the feasibility of stylet-driven pacing lead (SDL) was also established. The purpose of this study is to evaluate the learning curve for LBBAP using SDL.  The study enrolled 265 patients who underwent LBBAP or RVP performed by operators without previous LBBAP experience at Yonsei University Severance Hospital in Korea between December 2020 and October 2021. LBBAP was performed using SDL with an extendable helix. The learning curve was evaluated by analyzing fluoroscopy and procedure times. And, before and after reaching the learning curve, we evaluated how much the time required for the LBBAP differed from the time required for the RVP.  LBBAP was successful in 50 of 50 (100.0%) patients left bundle branch pacing was successful in 49 of 50 (98.0%). In 50 patients who underwent LBBAP, mean fluoroscopy and procedural times were 15.1 ± 13.5 minutes and 59.9 ± 24.8 minutes, respectively. The plateau of fluoroscopy time reached in the 25th case and the plateau of procedure time reached in the 24th case.  During the initial experience with LBBAP, fluoroscopy and procedural times improved with increasing operator experience. For operators who were experienced in cardiac pacemaker implantation, the steepest part of the learning curve was over the first 24-25 cases. It is shorter than the previously reported learning curves of HBP. Conventional right ventricular pacing (RVP) can cause electrical and mechanical dyssynchrony and is associated with increased risks of cardiac remodeling, pacemaker-induced cardiomyopathy, heart failure, and mortality , 2. ThesLeft bundle branch area pacing (LBBAP), proposed by Huang et al. in 2017 . In addiLBBAP presents a simpler implantation technique, shorter fluoroscopy duration, and a procedure time with a steeper learning curve compared to HBP. However, most of the learning curve analyzes of LBBAP reported so far have been procedures using LLL , 15. In The purpose of this study is to evaluate the learning curve of one operator of LBBAP using SDL. We confirmed that the procedure was performed stably through the lead and electrocardiogram (ECG) parameters, success rate, and complication rate of the procedure. And, by analyzing fluoroscopy and procedure time, we reported a learning curve for how many cases were needed per operator to get used to the procedure. Furthermore, differences in required radiation dose and procedure time when compared with existing RVP before and after achieving the learning curve of LBBAP were evaluated.n = 200) or LBBAP (n = 65) performed by operators without previous LBBAP experience between December 2020 and October 2021 at Yonsei University Severance Hospital in the Republic of Korea. All patients who required ventricular lead implantation during this period were included. For the learning curve analysis, patients who underwent dual-chamber pacemaker implantation by a single experienced operator were divided into a group that underwent conventional RVP (n = 65) and a group that underwent LBBAP (n = 50)  . This stAll procedures, including conventional RVP and LBBAP, were performed by operators without previous LBBAP experience. LBBAP was performed with a 5.6-Fr sized SDL with an extendable helix  and all the SDL delivered through a preshaped sheath  because it properly targets the pacing site and maintains sufficient backup during lead implantation.The lead was prepared as previously described by De Pooter et al. . BrieflyTwo methods were used to find the optimal site for LBBAP. Initially, tagging the His area using His/right ventricular (RV) catheter, then set that point as a landmark, place the lead tip at 1-2 cm toward the RV apex in the right anterior oblique (RAO) view and perpendicular to the septum in the left anterior oblique view as described in previous studies  activation time in surface V5-V6 ECG of approximately 75–85 ms by programmed stimulation should be noted, which demonstrates the transition from deep ventricular septal pacing to LBBAP  capture was confirmed using published criteria. With the advancement of the lead, QRS configuration with a right bundle branch block (RBBB) observed in surface V1 during the unipolar pacing in addition to one or more of the following findings, the success of LBBP was confirmed: (1) LBB potentials is recorded from the pacing lead, with the potential to ventricle interval of 15–35 ms [V5-V6 is shortens abruptly and measured <75–85 ms. Stim-LVAT is defined as the interval from the pacing stimulus to the peak of the R-wave and is often used to reflect the lateral precordial myocardium depolarization time in leads V5-V6 [RVP was defined as conventional RV apical pacing. LBBAP included deep septal pacing (DSP) and left bundle branch pacing (LBBP). The LBB potentials from the pacing lead were recorded and stimulus to left ventricular activation time was abruptly shortened (Stim-LVAT) which were confirmed as LBB capture. In details, If the RBBB configuration was seen in 15–35 ms –23; (2) ds V5-V6 , 18, 22;ds V5-V6 , 22; andds V5-V6 –23. Evends V5-V6 .t-test and Wilcoxon's rank-sum test. A paired t-test was performed to compare the measured values before and after the procedure. The fluoroscopy and procedure times were modeled as cubic spline functions. The cubic spine curve was mathematically differentiated and the point where the slope gradient converged to zero and forms a plateau was analyzed .Descriptive statistics were used to organize and interpret patients' baseline characteristics and comorbidities. Categorical variables are reported as frequencies (percentages), and continuous variables are reported as mean ± standard deviation or median (interquartile range). Categorical variables were compared using Fisher's exact test or Pearson's chi-square test, whereas continuous variables were compared using Student's A total of 200 patients who underwent conventional RVP  and 50 patients who underwent LBBAP  were included in this study.The distribution of pacing indications was as follows: in the conventional RVP group, sick sinus syndrome (SSS) was the most common indication (54.0%), followed by atrioventricular (AV) block (46.0%): in the LBBAP group, AV block was more frequent (81.5%) than SSS (18.5%). The QRS duration was more likely to be narrow in the conventional RVP group (62.0%) than in the LBBAP group (40.0%). A comparison of baseline characteristics among the groups is summarized in n = 39), and among them, RBBB, LBBB, and bifascicular block accounted for the highest frequency with the same number of 13.8% (n = 9). There was no statistical difference in overall baseline QRS morphology between the SSS and AV block groups (p=0.30). All patients who underwent LBBAP showed reduced QRS duration; the duration decreased from 128.6 ± 30.2 ms at baseline to 120.5 ± 20.1 ms after LBBAP. When the SSS was the pacing indication, the decrease in QRS duration was 7.50  ms, and when the AV block was the pacing indication, the decrease in QRS duration was 8.18  ms. Intraprocedural electrophysiological characteristics of LBBAP are summarized in Among 65 patients who underwent LBBAP, 60% of patients had baseline QRS widening  min. In the late period, the mean procedure time of RVP and LBBAP were 40.4 ± 17.1 and 52.4 ± 10.9 min, respectively, and the difference was 12.0  min. In the early period, the mean fluoroscopy times of RVP and LBBAP was 5.6 ± 3.4 and 20.5 ± 17.7 min, respectively, and the difference was 14.9  min. In the late period, the mean fluoroscopy times of RVP and LBBAP were 5.6 ± 3.0 and 10.0 ± 3.6 min, respectively, and the difference were 4.4  min. The time required for RVP and LBBAP before and after achieving the learning curve for LBBAP was presented as We evaluated how much effort required for the LBBAP procedure was different from that required for the conventional RVP after reaching the learning curve for LBBAP. Based on the 24th case whose procedure time reached the plateau phase, the entire study period was divided into early period and late period (before plateau phase ≤24th case vs. after plateau phase >24th case). Then, the difference between the procedure time for the LBBAP and the procedure time for the conventional RVP at the two periods were analyzed. In the early period, the mean procedure time of RVP and LBBAP were 46.7 ± 11.5 and 68.1 ± 32.4 min, respectively, and the difference was 21.4 , and ventricular pacing impedance of 675.7 ± 97.5 Ω. Baseline QRS duration was 127.7 ± 29.6 ms and paced QRS duration was 117.2 ± 19.0 ms , and LBBP, meaning LBB capture, was successful in all 50 to 49 patients (98.0%). There was 1 case of LBBP fail in the early period. After the procedure, the LBBAP lead parameters were sensed  19.0 ms .There were two cases of complications during the early period, which is LBBAP lead dislodgements occurring 1 day and 3 months after the procedure, respectively. Of the two dislodgement of ventricular lead cases, one patient required ventricular lead revision and LBBAP was achieved again, and one case did not require revision because of stable parameters with a ventricular lead in the RV inferior wall. There were 2 cases of pocket hematoma in the late period, and no procedure-related major complications occurred during that period .The major findings of this study were as follows: (1) the LBB capture threshold and pacing impedance in the LBBAP population were stable, comparable to values in general RVP; (2) after LBBAP, QRS duration was significantly decreased; (3) time required for LBBAP procedures continued to decrease as increasing operator experience and achieved a plateau starting from the 24th case.; and (4) higher success rates and lower complication rates were achieved as experience increased.Although HBP, first introduced by Deshmukh et al. in 2000, has been suggested as the ideal approach for physiological ventricular activation , 26, widIn this study, we evaluated the learning curve of LBBAP using SDL, the acute outcome of the procedure before and after achieving the learning curve, and the difference in time for procedure from the time required for existing RVP. The plateau phase of the procedure time for LBBAP using SDL began from the 24th case, which is shorter than the previously reported learning curves of HBP 30–50 cases) even LBBAP using LLL , 29, 30.–50 casesThere was a significant difference in the procedure time before and after achieving the learning curve; however, there was no statistically significant difference in the outcome of the procedure such as acute success rate, pacing parameter, and paced QRS duration in this study. These results make it possible to expect a stable procedure outcome even if an operator who has no experience of performs the LBBAP using SDL. This is encouraging considering the shortcoming of technical difficulties in identifying the precise location, variable success rates, and potential risk of premature battery depletion and lead revisions due to progressive increases in capture thresholds of HBP, the first CSP , 27.Procedure-related major complications occurred in the early period in all 2 out of 2 total cases. On the other hand, ventricular septal perforation, a known complication of LBBAP, did not occur during the entire duration of this study. This is expected in accordance with our method to prevent complications. We continuously monitored the lead impedance during screwing to avoid ventricular septal perforation and advance was stopped immediately if there was a sudden drop in lead impedance. In the early period, if the lead impedance showed a rapid drop even if it was 500 ohms or more, we did not advance further and fixed the lead in that position. Referring to previous studies , 20 and This study has several limitations. First, this was a retrospective, observational, and nonrandomized study. Second, we purposely analyzed the experience of a single operator with the aim of reducing bias, but we believe that the results are broadly applicable. Comparing the experiences of multiple physicians in multiple centers would have better ensured its generalizability.During the initial experience of LBBAP using SDL, procedural and fluoroscopy times continued to improve with operator experience. For physicians experienced in cardiovascular implantable electronic device implantation and without previous LBBAP experience, the steepest part of the learning curve was over the first 20–25 cases.


Teachers form a large and essential workforce globally. Their wellbeing impacts personal health-related outcomes with flow on effects for the health, and wellbeing of their students. However, food and nutrition (FN) interventions that include teachers, typically neglect the impact of personal FN factors on a teachers’ ability to achieve optimal nutrition-related health and wellbeing, and successfully fulfil their professional FN roles as health promoters, gate keepers, educators’, and role models. The aim of this review was to scope FN constructs that have been studied internationally regarding teacher FN-related health and wellbeing.Six databases were searched, and papers extracted in June/July 2021. Eligibility criteria guided by the population, concept, context mnemonic included studies published after 2000, in English language, with an aspect of personal FN-related health and wellbeing, among in-service (practising) and pre-service (training), primary, and secondary teachers. Screening studies for inclusion was completed by two independent researchers with data extraction piloted with the same reviewers and completed by lead author, along with complete descriptive and thematic analysis.n = 99 papers), with nutrition knowledge and dietary assessment among the most commonly assessed.Ten thousand six hundred seventy-seven unique articles were identified with 368 eligible for full text review and 105 included in final extraction and analysis. Sixty-nine descriptive studies were included, followed by 35 intervention studies, with the main data collection method used to assess both personal and professional FN constructs being questionnaires (FN constructs are used within interventions and studies that include teachers, with diversity in constructs included and how these terms are defined. The evidence from this scoping review can be used to inform data collection and evaluation in future epidemiological and interventional research that addresses teacher FN-related health and wellbeing.The online version contains supplementary material available at 10.1186/s12966-023-01502-5. Teachers’ health and wellbeing, including food and nutrition (FN) practices are influenced by professional workload and school environment , while tIndicators and predictors of food choices and eating behaviours are unique to everyone and acknowledged as complex in the Determinates of Nutrition and Eating (DONE) Framework, which identifies 51 determinate groups across four key domains of individual, interpersonal, environmental and policy , 11. FN Poor diet quality, specifically low intake of vegetables, fruit and wholegrains are well established as risk factors of chronic disease and contributors to global burden of disease . To thisTeachers need support and education on how to optimise their own FN-related health and wellbeing to help them in fulfilling their professional FN roles as healthy role models and advocates for the students in their care , 21. PreTherefore, the current scoping review aims to summarise the range of FN constructs included across research on teacher’s personal FN-related health and wellbeing. The review will map evidence on teacher FN-related health and wellbeing and how this has been evaluated to inform future research.A scoping review methodology was selected to both enable research from a diverse collection of areas across education, and health, while providing a structed yet iterative process that provided a clear review framework with the flexibility to refine parameters as references were collected and information sourced. The term FN-related health and wellbeing is used within the current review to encompass the complexity of individual, interpersonal, environmental and policy related constructs that influence an individuals’ FN decisions and healthy eating behaviours. For the purposes of this review a distinction is made between personal and professional FN constructs, as outlined in Table A scoping review protocol was developed, guided by the Joanna Briggs Institute (JBI) guidelines , the preEarly learning preservice and in-service teacher/educators were not included within this review as earlier scoping reviews exploring this population were identified –26 and fIn June/July 2021 six databases were searched: PsychInfo, ERIC via PROQUEST, CINAHL, Medline, Embase, Scopus. Database specific search strategies were developed in consultation with two senior University of Newcastle librarians’ using the population, concept, context, (PCC) mnemonic . The Medwww.covidence.org. Title and abstract screening, and full text screening were conducted by BF, TJ with conflicts resolved by discussion and by a third independent reviewer when conflicts could not be resolved. Reference lists of excluded full text papers, flagged reviews, and study protocols identified, were also screened for potential papers of interest.Screening of papers was conducted by two independent reviewers  using Covidence systematic review software, Veritas Health Innovation, Melbourne, Australia, available at A data extraction instrument was created by lead author (TJ) guided by the JBI manual for evidence synthesis  and piloPapers were grouped for descriptive analysis based on study type to assess distribution of included study characteristics and data collection methods. FN constructs were initially dichotomised as personal or professional, with constructs allocated to thematically appropriate groups based on the content description and sample questions provided in each eligible research paper. Where a description of the construct was not provided, it was placed in a suitable group based on name only.No formal statistical analysis was conducted to assess trends in different areas of teacher FN-related health and wellbeing such as nutrition knowledge or dietary intake due to the diversity of construct terminology identified across studies. Instead, further descriptive analysis was conducted using the primary study aim to allocate each paper to one of five groups: Teacher Personal FN, Teacher Professional FN, Student FN, Teacher and Student Personal FN, or Other. Final summary tables were transferred from Microsoft Excel spreadsheets into Microsoft Word and simplified for final presentation.n = 93) were the main participants, with the remaining 12 papers utilising pre-service teachers. Thirty-two papers included teachers with other participants such as guidance teachers’, assistant teachers’, administrative staff, transition teachers and in one instance other health fund members (professions not specified), with a complete summary provided in Additional file Figure n = 24) were sent to authors who were provided a month in which to respond. Thirteen papers where authors’ response was not received were excluded under the “Participants ineligible” exclusion reason as participant population was unable to be confirmed.Where some papers specifically mentioned the participant population were teachers, some did not clearly describe them as primary, secondary, or relevant international categories . Where applicable, to clarify this aspect emails (n = 44), Australia (n = 7), Iran (n = 7), Brazil (n = 6), South Africa (n = 5), Canada (n = 4) and Indonesia (n = 4). Country of study origin is provided for all included papers in Additional file Figure n = 69), intervention (n = 35), and measurement (n = 1). Included papers being further grouped using the primary study aim into five main groups for analysis of key characteristics and the types of FN constructs used based on the study purpose , Teacher Professional FN (n = 30), Student FN (n = 11), Teacher and Student Personal FN (n = 6) and Other (n = 21). Papers that focused on student-related aims favoured incorporating professional teaching FN constructs over personal FN constructs with the main personal FN constructs across these being nutrition knowledge, including food safely knowledge and skills, dietary assessment, and food attitudes. Papers within the ‘Other’ category most often included research focused on diet-disease relationships with teachers acting as a convenience sample with the focus solely on ‘personal FN’ or other health-related and wellbeing covariates or constructs.Included papers were grouped by study type as defined by Yoong et al.  which utAcross papers, a wide variety of personal and professional FN constructs were included, with differences noted in how similarly named constructs, such as nutrition knowledge were defined. A summary of all FN constructs identified and thematically grouped based on content descriptions and sample questions is provided in Table n = 31), followed by, nutrition knowledge (n = 12) and food or eating habits and behaviours (n = 8) being the most prominent, (body mass index and waist circumference were not included as a construct in this description). The remaining 38 papers captured two or more constructs, with different combinations represented including knowledge, attitudes [Of the 105 papers, 66 captured one personal FN construct, with dietary assessment  and nutrition knowledge (n = 33) were the two most utilised constructs across papers, followed by food or eating habits and behaviours (n = 23) and nutrition attitudes (n = 15). Additional file Dietary assessment  [n = 1) [n = 1) [n = 1) [n = 3) [n = 18) [n = 6) [n = 2) [n = 3) [n = 3) [n = 1) [n = 1) [n = 2) [Dietary assessment construct terms included dietary intake (e n = 1) , behavio [n = 1) , nutriti [n = 1)  and nutr [n = 1) , with ad [n = 3) –70. Pape[n = 18) –86, frui [n = 6) –92, fat  [n = 2) , 43, die [n = 3) –95, auto [n = 3) , 96, 97, [n = 1) , short f [n = 1)  or brief [n = 2) , 100.n = 8) [n = 7) [n = 4) [n = 5) [n = 4) [n = 5) [Nutrition knowledge was measured across 33 papers  , 101–106 [n = 7) , 107, di [n = 4) , food sa [n = 5) , 106–108 [n = 5) , knowled [n = 5) , healthy [n = 5) , or comb [n = 5) . Where c [n = 5) , 112–115 [n = 5) , 116–119 [n = 5) , 66, 120n = 4) [n = 2) [Within the food or eating habits and behaviours category a range of construct terms were identified across the 23 included papers  , 71, 121 [n = 2) , 122. Si [n = 2)  which ha [n = 2) , 99, 123 [n = 2) , 125. On [n = 2) . Of the  [n = 2) , 127, th [n = 2) , 58, 103n = 7), disordered eating (n = 5), dieting status and weight change behaviours (n = 9), , followed by, nutrition education self-efficacy (n = 11). These were also the most frequently observed construct groups with common tools used across included papers including the Classroom Food Practices construct, the School Food Environment Index [Of 42 papers measuring some aspect of professional FN , alcohol intake (n = 11), and sleep (n = 5). With mental health and wellbeing measured in 11 [Physical activity and/or exercise, including self-regulation of these was the most common covariate or construct included across 43 of the included papers, followed by smoking and/or smoking status and tobacco use  being the predominate data collection method, with participants self-reporting responses in paper-based or digital format. Five papers used researcher assistance to complete questionnaires [n = 51), followed by health-related data, which included blood pressure or fasting blood samples (n = 15). Physical assessment was measured in four papers with three papers, from the same study, using accelerometers with participants [n = 11) or focus groups (n = 4). Validation and reliability testing methods were reviewed for the data collection methods used across included papers with descriptions of psychometric testing often unclear, missing or a reference provided to indicate additional information on tool development and/or testing was reported elsewhere. Of the 105 papers reviewed only 22 papers included a clear description of validation methods used, with 13 providing an explanation of the reliability testing conducted. Further analysis of psychometric testing of data collection methods reviewed was beyond the scope of this current review . Self-reported questionnaires have lower participant burden and can incorporate multiple constructs of interest to address a wide range of study designs. While this method can be prone to participant completion error or bias, when conducting research with teacher participants reducing teacher participation time is a key factor for this population group who are usually time poor. Reducing participant burden as a strategy to increase study participation using questionnaires with close-ended responses may therefore be an effective way to optimise questionnaire completion. Within the current review descriptions of psychometric testing conducted on included data collection methods was limited and often unclear, with complete descriptions of items and scales used within questionnaires rarely included. This lack of validation and reliability descriptions and clear outline of the items or scales included within questionnaires makes comparison between tools challenging and limits other researchers in utilising these tools in future research.Questionnaires were the main method of assessing many different constructs  in the abstract, potentially relevant papers may have been excluded in screening phase. Hence, although many studies are included, results should be interpreted with caution. Additionally, studies that indicated a teacher FN component, but did not provide a clear description within the methods were excluded at full text screening for lack of detail.While thematic analysis provided a broad overview of the common themes and gaps, it is acknowledged that the strength of the current review is the mapping of the definitions, descriptions, and sample questions detailing FN-related constructs provided in the included papers. Therefore, where limited description was provided, these constructs may have been incorrectly placed within a construct category. However, this further highlights the need for more clarity and detailed descriptions of constructs used within teacher interventions and the possible sharing or inclusion of sample questions, to ensure research is represented as intended in future studies and the need for standardisation of construct terminology.This is the first review to map where and how teacher personal FN have been reported across international research including student and school focused papers. While facets of teacher FN have been studied across a wide range of research areas, the lack of validated tools or clearly defined evidence-based FN constructs used in research to date makes comparisons or assessment of teacher population personal FN status challenging. Future research is needed to address these gaps.Additional file 1. PRISMA-Scr Checklist.Additional file 2. Search strategy by Database.Additional file 3. Overview of Data Extraction Table.Additional file 4. Primary Aims Summary Table (All Included Studies).Additional file 5. Personal FN-Summary of Constructs Observed.Additional file 6. Professional FN-Summary of Constructs Observed.Additional file 7. Main Constructs.


PLOS ONE 12(4), e0175583 : null hypothesis significance testing; confidence or credible intervals; meta-analysis of the results of multiple experiments; confidence interval interpretation; effect size interpretation; sample size determination; data exclusion; data availability; materials availability; and preregistered design and analysis plan. The investigation was based on an analysis of all papers published in these journals between 2013 and 2015. The aim of the present study was to follow up changes in both PS and JEPG in subsequent years, from 2016 to 2020, adding code availability as a further open practice. We found improvement in most practices, with some exceptions . Despite these positive changes, our results indicate a need for further improvements in statistical practices and adoption of open practices.Changes in statistical practices and reporting have been documented by Giofrè et al. e0175583 , who invThe online version contains supplementary material available at 10.3758/s13428-022-01993-3. Over the last decade, many published findings have been found not to be fully replicable . The investigation was based on an analysis of all papers published in these journals between 2013 and 2015. Results showed that the introduction of new statistical guidelines, in particular for PS, was followed by an improvement in most statistical practices, for example, reporting confidence or credible intervals, and reporting more information about the sample . Concerning Open Practices, the authors found that the inclusion of new guidelines was beneficial, for example, in increasing the number of papers providing raw data and additional materials. However, the authors also found some limits related to the introduction of these new practices and guidelines.The changes in statistical practices and reporting have been documented by Giofrè et al. , who invp value and not on the interpretation of the effect size or using a different approach, such as Bayesian). And now, although researchers may be reporting more information, they may still be interpreting and basing their conclusions using a traditional NHST approach, which has been criticized on several occasions  Guidelines  of the Publication Manual has recently been released . These guidelines remain very similar to the ones published in 2015, with some parts reworded. For example, in the previous version, PS asked authors to provide a rationale for the sample size; in 2021 they specified where in the manuscript this should go. A blurb was added about replicability being not the only consideration but an important one; authors are also now encouraged to use graphical presentations to show how data are distributed and to include information about sample sizes in the manuscript’s abstract.PS recently updated its guidelines on August 18, 2021, as can be found here: https://www.apa.org/pubs/journals/xge/ (accessed September 2021). Recent changes have included requiring authors to state whether data are publicly available and where they can be accessed; whether or not work has been preregistered. Time will tell how these specific guidelines will affect transparency in the long run, but for the purposes of the present study, in the period from 2016 to 2020, these journal-specific guidelines, or in the case of JEPG, a lack thereof, had not changed markedly.Similarly, JEPG updated its guidelines from July 1, 2021, as can be found here: https://www.cos.io/initiatives/top-guidelines).The aim of the present study is to document any changes in authors’ statistical practices in PS and JEPG from 2016 to 2020. We investigated the previously studied practices, and added one more: the availability of statistical code, or script . Subsequent to the publication of the original report . We hypothesized that improvements in statistical practices and open practices in PS, which changed its journal submission guidelines in 2014, would be larger than the improvements in JEPG, which only referred to third-party guidelines. While we did expect to see an increase in the adoption of the open practices, building on those observed in Giofrè et al. . The scoring procedure and method is identical to the procedure adopted by Giofrè et al.  or relative .NHST. A CI. A confidence interval or credible interval (in the case of Bayesian statistics) was reported for either a standardized or unstandardized measure.MA. Meta-analysis of multiple related results included in the paper was reported. We only included papers with more than one experiment related to the same empirical question.CI_interp. A confidence interval was referred to and used to inform the discussion or interpretation of the results. For example, this would include a paper explicitly mentioning the width or the precision of the CI, a comparison between two or more CIs, or an overlapping between two intervals.d, R2, and η2. Papers were considered which included not only a dichotomous difference vs. no difference approach, but also those referring to the magnitude of the effect  or to the amount of explained variance.ES_interp. An effect size, either standardized or unstandardized, was referred to in the discussion or interpretation of the results. We considered ‘effect size’ in the broad sense, including means, differences between means, percentages, and correlations, as well as Cohen’s Sample_size. The authors had a statistical rationale for how sample size(s) was determined. For example, a prospective power analysis – based on previous research, or on an estimated effect size – had been conducted. We excluded papers where the sample size was left to chance, e.g., “Sample size was determined by signup,” or, “Sample size was determined by participants’ availability."Data_excl. Authors explicitly declare if and which stopping rule was adopted or the criteria used to exclude data/participants.Data. The paper carried the Open Data badge (see below), or stated where the data were available or how they could be obtained. We used a very lenient approach, including all papers mentioning that data were available .Materials. The paper carried the Open Materials badge, or stated where details of the experimental materials and procedure could be obtained. We used a very lenient approach, including all papers mentioning that materials were available .Preregistered. The paper carried the Preregistered badge, or stated where a preregistered plan had been lodged in advance of data collection. Papers in this category typically included information about the number of the preregistration or where the preregistration is available.Code. The paper provided, in the paper, the supplementary materials, or the preregistration, the computer code or syntax  needed to reproduce the analyses. We used a very lenient approach; for example, we did not check if the code was sufficient to successfully replicate all of the analyses presented in the paper.“Published journal papers, and online supplemental material when available, were considered. A single occurrence of a practice anywhere in the paper was sufficient for a coding of 1, indicating that this practice had been used, of 0, if had not been used, and “NA” if not applicable. Papers were examined for the following eleven practices:https://osf.io/tvyxz/wiki/home; accessed September 2021).The three badges  are described in detail by the Center for Open Science . For PS, these badges were certified by an “earned badge” from the Open Science Framework (https://www.comses.net/resources/open-code-badge (accessed October 2022).The Open code badge is described here: For JEPG, badges are not available, but we coded whether or not authors clearly indicated how to obtain the data, materials, and code.”The first level of scoring method and reliability followed exactly the same rules as in the original Giofrè et al.  paper. TPapers were examined for the presence of each of the 11 practices. For each, the score could be 1 if present, 0 if absent, and “NA” if not applicable. The total number of included papers for each journal, from 2016 to 2020, are reported in Table Each paper was independently scored by one of the authors . The authors are experienced researchers with good knowledge of the statistics examined. Second, a random sample of 137 papers, being roughly 10% of the whole set of papers, was scored independently by a second judge to test inter-rater reliability. The overall agreement was 96%, with discrepancies resolved by discussion.Only descriptive statistics are reported, given that they refer to the whole population of papers. For each of the 11 practices analyzed, the number of papers including a practice are expressed as a percentage of the total number of papers. It is also worth mentioning that for ES_Interpr and for CI_Interpr the denominator corresponded the total number of all included papers, not the percentage of papers using an ES or a CI. The checklist for study examination is presented in Table We decided to use percentages rather than proportions. The two are equivalent, but percentages are more commonly reported. As for the Sample_size determination criterion, in which authors clarified how they determined the sample size, on several occasions authors used very vague statements . Therefore, we also decided to check how many papers used a more stringent criterion, that is if authors determined their sample size using a power analysis . Also in this case, we used a very lenient approach, any mention to a power analysis , qualified to achieve this criterion .Table p value , we found that in both journals, percentages were extremely low Table  and S3. The rate of CI_interpretation in both journals is extremely low, and has not improved over time  were calculated on the full set of included papers . As expected, the percentage of papers reporting an a priori power analysis was much lower than the percentage of papers reporting on sample size determination. The pattern was similar in the two journals Table ; Fig. 2.The aim of the present study was to follow up changes in PS and JEPG in subsequent years, from 2016 to 2020, adding code availability as a further open practice.The so-called replicability crisis originated from observations that findings in a variety of areas of science failed to replicate , preregistration requires authors only to answer a series of prompts, such as the following: hypothesis, dependent variable, conditions, analyses, outliers and exclusions, and sample size. Answering only a few basic such questions may result in a lack of completeness in the description of all methodological and statistical details, and so deviations from initial plans might not be identifiable. A full report should permit identification of deviation from what was preregistered. The problem is deviation in details that were not specified in a broad-brush preregistration is difficult to identify.Considering preregistered studies, we found that the percentage of these has considerably increased, having been extremely low in the past


Clinical trials in the pediatric population demonstrated similar results in those with relapsed/refractory ALCL. Incorporation of brentuximab vedotin into upfront therapy for children and adolescents with ALCL showed that this novel combination therapy has clinical advantages in comparison to conventional agents alone. Brentuximab vedotin is well-tolerated in both the pediatric and adult populations, even when used in combination with conventional agents. Brentuximab vedotin is an ideal agent to treat ALCL with excellent targeted activity and limited toxicity. Future studies are needed to identify how brentuximab vedotin should be utilized when combined with immunotherapy or other targeted agents  in both the upfront and relapsed/refractory setting.Anaplastic large cell lymphoma (ALCL) is the most common type of mature T-cell non-Hodgkin lymphoma in children/adolescents. ALCL is characterized by expression of CD30 in the neoplastic lymphoid cells with frequent expression of anaplastic lymphoma kinase (ALK), especially within the pediatric population. Despite multiple efforts to optimize the use of conventional chemotherapy, outcomes in children, adolescents, and adults with ALCL remain suboptimal. Thus, there is a need to improve survival for those with high-risk disease and decrease therapy exposures and toxicities for those with low-risk disease. Targeted therapies, such as the anti-CD30 antibody-drug conjugate, brentuximab vedotin, are new important therapeutic options. Phase I and II studies in adults with relapsed/refractory CD30 Whilse (ALK) . The maj or ALK- . In contK) genes –10.+ malignancies include co-expression of CD30 with epithelial membrane antigen and/or T cell antigens in the presence of anaplastic morphologic features  is a cell membrane protein that is identified in all cases. The features that differentiate ALCL from other types of CD30features . CD30 isfeatures –14. For in vitro and in vivo efficacy in CD30+ lymphoma , a synthetic anti-neoplastic agent that binds tubulin. SGN-30 demonstrated both lymphoma , and thelymphoma . Brentuxlymphoma Figure 1lymphoma –22.in vitro activity against ALCL. Incubation of two CD30-expressing ALCL cell lines with two anti-CD30 monoclonal antibodies (M44 or HeFi-1) resulted in significant growth inhibition and decreased cell viability , preclinical studies showed that the conjugated drug was highly potent and selectively more active against CD300- cells . The antll lines –26.+ ALCL   or CHOP-like regimens were used most frequently. In a systematic review/meta-analysis of anthracycline-based chemotherapy for peripheral T cell lymphoma, the complete remission (CR) rate for ALCL was 65.8% with a 5-year overall survival (OS) of 56.5% . The 2-y P=.008) , 30. Aut P=.008) . Prior t P=.008) , 32, 33.Similar to adults, outcomes for children/adolescents with systemic ALCL were suboptimal with a failure rate of 25 – 30% despite multiple attempts to improve upfront therapy. Adding methotrexate and high-dose cytarabine to APO  chemotherapy, compressing the duration of T-cell lineage therapy, or adding vinblastine to APO produced increased toxicities without improving efficacy –36. In tOutcomes for those with relapsed ALCL, and especially for patients who progress on therapy, are also suboptimal. Brugieres et al. performed a retrospective study within a heterogeneous population of patients with relapsed/refractory ALCL who received vinblastine salvage therapy and found that the 5-year EFS was only 30% . Chemoth+ lymphomas using brentuximab vedotin   while 42 were ALK- (72%), and 29 (50%) had relapsed disease while the remaining 29 had refractory disease. Fifty of the 58 (86%) patients had an objective response with ALK+ and ALK- disease having similar objective response rates . Thirty-eight (66%) of 58 patients achieved a CR, and these patients had a higher 5-year OS (79%) than those who did not (25%). Sixteen of the patients with a CR went on to either autologous (n = 8) or allogeneic (n =8) HSCT, with similar outcomes for each transplant type at five years. At the time of last follow-up, 16 of the 38 patients who achieved a CR remained in remission after receiving a median of 9 cycles of brentuximab vedotin. While 8 of these patients had received an HSCT , 8 remained in remission without additional therapy. Though 33/58 (57%) of patients experienced peripheral neuropathy, 91% of these patients had resolution or improvement of their neuropathy at time of last assessment. This study demonstrated that brentuximab vedotin monotherapy is tolerable and effective in patients with relapsed/refractory ALCL and led to the FDA approval of brentuximab vedotin for adults with relapsed or refractory ALCL in 2011. Additional studies have demonstrated efficacy of brentuximab vedotin retreatment after initial brentuximab vedotin therapy   . Patient+ peripheral T cell lymphoma received either A+CHP or CHOP for 6 – 8 cycles (n=226 in each group) (+ (n= 98) or ALK- (n=218). Overall, both the OS and PFS were superior for the A+CHP arm . The median PFS was 48.2 months (A+CHP) vs. 20.8 months (CHOP), with a 29% reduction in the risk of a PFS event. For those with systemic ALCL, there was a 41% reduction in the risk of a PFS event for those receiving A+CHP. This significant difference in outcomes persisted, with a 5-year PFS of 51.4% with A+CHP vs. 43.0% with CHOP  and a 5-year OS of 70.1% with A+CHP versus 61.0% with CHOP   . The med+ (n=12) or ALK- (n=5), and six had already received a HSCT. This study determined the same recommended phase II dose as adults (1.8 mg/kg). Of the 17 patients with ALCL, 9 (53%) had a response and 7 (41%) patients achieved a CR at this dose. Patients with ALCL received a median of 9 cycles of therapy, and a total of 13 patients subsequently underwent an HSCT. The most common ≥ grade 3 AEs included neutropenia, increased γ-glutamyl transpeptidase, and pyrexia. Overall, 3/36 (8%) patients had a drug-related serious AE. While 12 of 36 (33%) patients experienced peripheral neuropathy, only one had severity ≥ grade 3 and 11 of the 12 cases improved or resolved. This trial was the first to demonstrate meaningful tumor response with a manageable safety profile in children/adolescents with CD30+ lymphoma receiving brentuximab vedotin monotherapy.A multi-center phase I/II study evaluating brentuximab vedotin in pediatric patients with relapsed/refractory classical Hodgkin lymphoma or systemic ALCL enrolled 36 patients with a median age of 14 years (range 7 – 18 years) . Sevente+/CD30+ ALCL  to determine the tolerability, EFS, and OS of adding either brentuximab vedotin or crizotinib to ALCL99 chemotherapy in children and adolescents with systemic ALK30+ ALCL , 52. Six+, including mycosis fungoides (MF), primary cutaneous ALCL (pcALCL), and lymphatoid papulosis (LyP). While these disorders can be more clinically benign and indolent than systemic CD30+ lymphomas, those that progress to a higher stage or systemic disease may have poor outcomes  at doses of 1.8 mg/kg every 21 days revealed an overall response rate of 73% (95% CI: 60 – 86%) and CR rate of 35% (95% CI: 22 – 49%) (+ mycosis fungoides (n=97) or pcALCL (n=31) were treated with either brentuximab vedotin  or physician’s choice  of either oral methotrexate or oral bexarotene. At median follow-up of 22.9 months, those with an objective global response lasting at least 4 months was 56.3% with brentuximab vedotin vs. 12.5% with physician’s choice (P<0.0001) (+ diseases with brentuximab vedotin.Specific subtypes of primary cutaneous T cell lymphomas (PCTL) are CD30outcomes . Importa2 – 49%) . In a ph<0.0001) . Respons+ ALCL tumors demonstrate strong PD-L1 expression, and the successful use of anti-PD1 monoclonal antibodies for ALCL patients has been reported in multiple cases (The scientific advancements surrounding the understanding of ALCL biology and therapy have resulted in better outcomes for children/adolescents and adults with systemic ALCL. Brentuximab vedotin has demonstrated both safety and efficacy in the treatment of pediatric and adult systemic ALCL, and regimens with brentuximab vedotin are now the standard of care in newly diagnosed patients. Optimizing the use of brentuximab vedotin will continue to improve outcomes while hopefully reducing toxicity. Specifically, future studies should evaluate the role of brentuximab vedotin in combination with ALK inhibitors, which have also shown excellent activity in ALCL. Brentuximab vedotin in combination with immunotherapeutic strategies may also demonstrate therapeutic potential. For example, ALKle cases –58. Brenle cases –61. ThesJA and EL: conceived of and designed the manuscript. JA: performed the literature search and drafted the manuscript. JA and EE: designed the tables/figures. JA, EE, EL: edited and revised the manuscript. All authors contributed to the article and approved the submitted version.


The hollow protein capsids from a number of different viruses are being considered for multiple biomedical or nanotechnological applications. In order to improve the applied potential of a given viral capsid as a nanocarrier or nanocontainer, specific conditions must be found for achieving its faithful and efficient assembly in vitro. The small size, adequate physical properties and specialized biological functions of the capsids of parvoviruses such as the minute virus of mice (MVM) make them excellent choices as nanocarriers and nanocontainers. In this study we analyzed the effects of protein concentration, macromolecular crowding, temperature, pH, ionic strength, or a combination of some of those variables on the fidelity and efficiency of self-assembly of the MVM capsid in vitro. The results revealed that the in vitro reassembly of the MVM capsid is an efficient and faithful process. Under some conditions, up to ~40% of the starting virus capsids were reassembled in vitro as free, non aggregated, correctly assembled particles. These results open up the possibility of encapsidating different compounds in VP2-only capsids of MVM during its reassembly in vitro, and encourage the use of virus-like particles of MVM as nanocontainers. The protein capsids of many viruses are able to self-assemble from individual capsid building blocks (CBBs) under appropriate conditions. In vivo, some virus capsids are assembled without the participation of the viral nucleic acid, which is later encapsidated in an independent reaction; other virus capsids are co-assembled with the viral nucleic acid to directly yield virions . HoThe finding of conditions for in vitro assembly of nucleic acid-free (empty) capsids from a number of different viruses has opened up the possibility of using them as nanocarriers or nanocontainers for a plethora of biomedical or nanotechnological applications. A general strategy involves the encapsidation of specific molecules or macromolecules during the in vitro assembly of either natural or engineered viral capsids . ThFor papillomavirus capsids, conditions have been found for the very efficient (~60–100%) self-assembly of structurally correct capsids in vitro ,14. HoweThe capsids of parvoviruses, including the minute virus of mice (MVM) ,26 may bThe authentic MVM capsid is made from two variants (VP2 and VP1) of the same capsid protein, which have an identical sequence and fold but differ in that VP1 has a longer N-terminal extension. VP2 alone is required for self-assembly, as the MVM capsid can be assembled from 60 identical VP2 subunits, both in transfected cells  and in tIn this study we have analyzed the effects of several different conditions on the fidelity and efficiency of self-assembly of the VP2-only MVM capsid in vitro. The results revealed that the in vitro reassembly of the MVM capsid is a remarkably faithful process, irrespective of the conditions used, and that its efficiency can be increased under certain conditions of pH, ionic strength, and/or macromolecular crowding.2HPO4, 1.5 mM KH2PO4, 137 mM NaCl, 2.7 mM KCl, pH 7.6) at 30,000 rpm in an SW40 rotor for 5.5 h at 10 °C. Purified VLP suspensions were extensively dialyzed against PBS, concentrated by ultrafiltration using an Amicon-Ultra4 device of 100 kDa cutoff , and stored at 4 °C. VLP purity, integrity and monodispersity were analyzed by polyacrylamide gel electrophoresis and transmission electron microscopy (TEM). Because of the very large amount of VP2 and VLPs produced in insect cells, the two-step ultracentrifugation approach yielded highly pure preparations. MVM capsid protein concentration was determined by UV spectrophotometry.MVM capsids made of 60 VP2 capsid protein subunits (termed here VLPs) were expressed, assembled in insect cells, and purified as described previously ,38. In eSamples were deposited on ionized Formvar/carbon-coated copper grids , incubated for 3 min and washed 3 times for 30 s with distilled water. Negative staining was carried out using incubation for 45 s in a 2% (weight/volume) uranyl acetate (Electron Microscopy Sciences) solution. The samples were then dried and visualized in a JEM-1010 electron microscope (JEOL) at magnifications ranging from 25,000× to 120,000×. Images were taken with a TemCam-F416 camera (TVIPS). Samples from each experiment were processed strictly in parallel, to allow a direct comparison of the data obtained.Purified native VLPs obtained from cells as described above were disassembled into free capsid building blocks. Purified VLP samples at a total protein concentration of 0.5–0.6 mg/mL were incubated at 25 °C in PBS pH = 7.6 in the presence of 3.5 M guanidinium chloride (GdmHCl) with agitation (350 rpm) in a Thermomixer Compact  for a specified amount of time  until VLP disassembly was nearly complete, as determined by TEM.VLP reassembly from capsid building blocks obtained by the disassembly of purified VLPs as described above was triggered by fast dialysis of 150 µL samples for 12–25 min (unless otherwise noted) in Slide-A-Lyzer cassettes of 0.5 mL capacity and 10000 MWCO . Residual GdmHCl concentration after dialysis was estimated using a spectrophotometry-based approach . DiffereThe relative number of complete VLPs as a function of experimental conditions and time was estimated by counting the particles present in several  EM fields of a defined size (1.23 × 1.23 μm), and averaging the results. Particle counting was usually carried out using a home-made macro developed in Fiji . In ordebN, dN and rN are the average number of complete VLPs counted, respectively, before disassembly was triggered by adding GdmHCl (bN), just after the disassembly reaction was ended  and after the reassembly reaction was ended (rN). dD is the dilution factor due to the addition of a volume of concentrated GdmHCl to trigger VLP disassembly. rD is the dilution factor due to the addition of a volume of a particular solution to determine VLP reassembly under certain specified conditions.The reassembly efficiency was calculated as follows:The chemical stability of the MVM VLPs under different pH and ionic strength conditions in different buffers  was tested by the addition of a GdmHCl solution to a final concentration of 3.75 M and incubation of the sample at 25 °C. Small aliquots were taken at different times and the percent remaining intact VLPs under each condition was estimated by TEM, as described above.Models of MVM CBBs were built using the structure and symmetry matrices corresponding to the PDB entry 1Z14 . Atomic Unless indicated otherwise, in this study the reassembly efficiency of MVM VP2-only capsids (VLPs) was defined in terms of the final yield of free VLPs assembled from capsid building blocks that had been obtained by disassembly of native VLPs. This is a demanding definition: even if 100% of the possible VLPs were correctly reassembled, the reassembly efficiency thus defined would be lower if some of them dissociated or became aggregated before the product was analyzed.To determine the reassembly efficiency under each tested condition we used as a starting point a previously described experimental setup ; see MetA and aftTo start the reassembly reaction, most of the GdmHCl was rapidly removed by dialysis under the conditions chosen for VLP reassembly C. DialysThe total number of free reassembled VLPs and the reassembly efficiency were estimated first as a function of capsid protein (VP2) concentration at 25 °C in PBS pH = 7.6 . ConcentThe main reason behind this decrease in reassembly efficiency  at the higher protein concentrations was traced to the formation of VLP aggregates . These aA propensity for aggregation was also observed for monodisperse native VLPs that had been correctly assembled in cells, and that had not been disassembled and reassembled in vitro. After storage for long enough at relatively high concentrations  at either 25 °C or 4 °C in PBS pH = 7.6, intact, free VLPs formed aggregates similar to those observed after VLP reassembly in the same buffer. VLP aggregation was generally observed, irrespective of the buffer in which the particles were stored or reassembled. Those buffers included phosphate-based buffers from pH 6.0 to 8.0 containing from 0.15 to 2.0 M NaCl, and 75 mM Tris-HCl pH = 6.8, 250 mM NaCl. The stickiness of native MVM VLPs led also to the clustering of latex beads, due to adsorption of VLPs to the beads and to each other . The aboA test was then carried out to see whether the reassembly efficiency could be increased by using a macromolecular crowding agent to increase the capsid protein (VP2) chemical activity (“effective concentration”) without actually increasing the number of protein molecules in the sample volume. Ficoll-70 was chosen as the crowding agent because of its low viscosity, a size comparable to those of many proteins, and its low tendency to interact specifically with proteins and other solutes .The relative reassembly efficiency in the presence or absence of 50 g/L Ficoll-70 at 25 °C in PBS (pH = 7.6) was compared as a function of reaction time . The resNext, tests were carried out to see whether the reassembly efficiency could be increased by using temperatures lower or higher than 25 °C. After the reassembly reaction was essentially complete (the VLP yield reached a plateau) the reassembly efficiency was similar at 25 °C or 37 °C, and somewhat lower at 4 °C or 15 °C .Electrostatic screening of protein–protein interactions by increasing the ionic strength may heavily influence interactions between capsid proteins and/or VLP aggregation. Thus, the effect of ionic strength on VLP reassembly efficiency was tested by increasing the NaCl concentration in the buffer from 0.15 M up to 2 M A. The reWe then asked whether the higher reassembly efficiency found at 0.5 M NaCl correlated with some change in the values and distribution of the surface electrostatic potential on the VLP surface, including the intertrimer interfaces. Simple surface electrostatic calculations  revealedChanges in the protonation state of acidic or basic side chains on the VLP surface or at the intertrimer interfaces could also influence VP2–VP2 interactions and/or VLP aggregation. Thus, we compared the reassembly efficiency within a pH interval from 6 to 8 A. The reThe stability of many viral capsids is pH-dependent. Thus, a higher capsid stability at slightly acidic pHs could contribute to the higher reassembly efficiency determined for MVM VLP at acidic pH, compared to neutral or slightly basic pH. To test the pH dependency of VLP stability, a dissociating agent was added (3.75 M GdmHCl), and the disassembly rate of purified VLPs as a function of pH was compared in two different buffers with a different ionic strength . The VLPAs for ionic strength, we asked whether the higher reassembly efficiency and particle stability at slightly acidic pH correlated with some change in the values and distribution of the surface electrostatic potential on the VLP surface, including the intertrimer interfaces. Surface electrostatic calculations  revealedWe then asked whether the increases in reassembly efficiency at relatively high ionic strength or acidic pH relative to physiologic ionic strength or neutral or basic pH were additive. The reassembly efficiency in PBS was compared at pH = 7.6, 0.15 M NaCl, pH 7.6, 0.5 M NaCl; pH 6.6, 0.15 M NaCl; and pH 6.6, 0.5 M NaCl . As expeVariations in some conditions tested for MVM VLP assembly, including capsid protein concentration, the absence or presence of a macromolecular crowding agent, ionic strength, or pH, led to 2- to 3-fold increases in reassembly efficiency. However, the maximum absolute reassembly efficiencies thus achieved were never higher than 35–40%. From these observations alone it could be thought that, in vitro, the self-assembly reaction that builds the MVM capsid is only moderately efficient, and not fully reversible. In fact, incomplete or aberrant MVM VLPs  were onlDifferent conditions were tested to attempt the disaggregation of the VLP clusters. Sonication did not achieve any substantial effect. Addition of a chaotropic agent (GdmHCl) at moderate concentrations (1.75 M) led to some disaggregation of large clusters into smaller clusters and individual free VLPs visualized by TEM, which again confirmed that the aggregates are made of fully reassembled VLPs. However, the addition of GdmHCl not only led to the release of some VLPs from the aggregates, but also to the dissociation of some of the free VLPs in the sample. Thus, the yields of free VLPs were not significantly increased (results not shown). Monodisperse suspensions of reassembled VLPs of MVM could be achieved only by elimination of the remaining aggregates through centrifugation.Extensive inspection using EM of the products of MVM VLP reassembly in vitro during this study did not reveal significant amounts of incorrectly assembled, aberrant viral particles under any tested condition of protein concentration, macromolecular crowding, temperature, pH, or ionic strength. These results provide evidence for a remarkable fidelity of MVM assembly that is virtually unaffected by changing conditions.The assembly of MVM and other parvoviruses inside infected mammalian cells is a complex process ,50. StabRecombinant VP2-only capsids (VLPs) of MVM and other parvoviruses (canine parvovirus (CPV) and porcine parvovirus) have been assembled in insect cells ,37,58,59Recombinant VLPs of MVM, AAV2 or human parvovirus B19 have been assembled in vitro starting from VP subunits ,60,61. DIn vitro assembly of the MVM capsid along a defined pathway  did not The above observations reinforce the notion that the higher complexity of parvovirus capsid assembly in mammalian cells arose because of the need to achieve during infection a regulated, highly efficient virus morphogenetic process in a compartmentalized, intricate cellular environment. In a highly dilute solution in the test tube under adequate conditions, the isolated, stable VP trimers were able to spontaneously and efficiently recognize each other with the correct orientation. The result is the faithful and efficient self-assembly of the capsid through a defined pathway (, and thiTo sum up, this study has defined conditions for the faithful and efficient in vitro self-assembly of MVM VLPs in the absence of additional factors that may be required for in vivo capsid assembly. These results encourage the use of MVM VLPs as nanocarriers for different compounds. It must be noted that the defined assembly conditions constitute a starting point that might have to be changed somewhat, depending on the influence that the desired cargo molecules could have on the intertrimer interactions. The high thermal, chemical and mechanical stability of these virus-based VLPs and their biological features, such as their nuclear tropism, make them suitable choices for targeted drug delivery or other biomedical or nanotechnological applications.


Signal regulatory protein alpha (SIRPα) is the receptor for cluster of differentiation (CD)47, a potent “don't eat me” signal for macrophages. Disruption of CD47‐SIRPα signaling in the presence of prophagocytic signals can lead to enhanced phagocytosis of tumor cells, resulting in a direct antitumor effect; agents targeting this pathway have shown efficacy in non‐Hodgkin lymphoma (NHL) and other tumor types. GS‐0189 is a novel anti‐SIRPα humanized monoclonal antibody. Here we report: (1) clinical safety, preliminary activity, and pharmacokinetics of GS‐0189 as monotherapy and in combination with rituximab from a phase 1 clinical trial in patients with relapsed/refractory NHL ; (2) in vitro characterization of GS‐0189 binding to SIRPα; and (3) in vitro phagocytic activity. Clinically, GS‐0189 was well tolerated in patients with relapsed/refractory NHL with evidence of clinical activity in combination with rituximab. Receptor occupancy (RO) of GS‐0189 was highly variable in NHL patients; binding affinity studies showed significantly higher affinity for SIRPα variant 1 than variant 2, consistent with RO in patient and healthy donor samples. In vitro phagocytosis induced by GS‐0189 was also SIRPα variant–dependent. Although clinical development of GS‐0189 was discontinued, the CD47‐SIRPα signaling pathway remains a promising therapeutic target and should continue to be explored. SystemiSignal regulatory protein alpha (SIRPα) is a receptor expressed on phagocytic cells—such as macrophages, neutrophils, monocytes, and dendritic cells—that binds to the antiphagocytic signal cluster of differentiation (CD)47 , 14, iniV1/V1 and SIRPαV2/V2) or heterozygotes (SIRPαV1/V2) , MDE1; 30 mg GS‐0189 [n = 1], MDE2; and 100 mg GS‐0189 [n = 4] MDE3) and 3 in the first CDE cohort . After the first CDE cohort, the decision was made to terminate the SRP001 study . Patient characteristics are shown in Table n = 2) and progressive disease (n = 7). Reasons for study discontinuation were death (n = 1), withdrawn consent (n = 2), and study terminated by sponsor (n = 6). Treatment‐emergent AEs (TEAEs) occurred in all but one patient across treatment groups. The most common TEAEs occurring in >1 patient in any group were infusion‐related reactions (IRRs), anemia, and neutropenia. Of the 4grade 3 TEAEs , only neutropenia was related to GS‐0189. There were no DLTs, grade 4 TEAEs, or events leading to death  in CDE1 who had previously relapsed after 2 lines of rituximab‐containing regimens achieved a complete response (CR) on study day 82 with a DOR of 5.19 months and PFS of 7.85 months. The DOR and PFS were censored at the time of last adequate tumor assessment. Study treatment was discontinued due to patient decision, which coincided with the time of sponsor decision to discontinue the study. Two patients had stable disease  with estimated PFS of 5.55 and 3.71 months, respectively; 5 had progressive disease  and patients , homozygous SIRPαV2/V2 , and heterozygous SIRPαV1/V2 . RO binding curves from healthy donors showed GS‐0189 bound to the homozygous SIRPαV1/V1 variant 3‐ and 102‐fold more strongly than to SIRPαV1/V2 and SIRPαV2/V2 variants, respectively  than for SIRPαV2 (KD = 332 nM). KWAR23 demonstrated a narrower range of affinities to SIRPα variants: KD = 6.7 nM for SIRPαV1 and KD = 14.2 nM for SIRPαV2.Binding affinities of GS‐0189 to recombinant SIRPαts Table  and sens3.3.2V1/V1 and SIRPαV2/V2 variants was induced by GS‐0189 combined with rituximab  of GS‐0189 with macrophages from donors expressing SIRPαV1/V1 was 0.02 μg/mL, expressing SIRPαV1/V2 was 13.8 μg/mL, and expressing SIRPαV2/V2 was 9.5 μg/mL. KWAR23 induced phagocytosis across SIRPα variants with EC50 ranging from 0.04 to 0.10 μg/mL  Fc region that was theorized to benefit from the presence of another drug that provides an “eat me” signal, such as rituximab, to induce phagocytic activity. In this clinical study, there was no observed response in patients enrolled in the MDE cohorts. In CDE1, one patient achieved a CR at approximately 12 weeks, which was maintained until study termination. This patient had stage 4 FL, relapsed on 2 prior lines of rituximab‐containing regimens, and was homozygous for the SIRPαV1 variant was nearly 2 orders of magnitude higher than for SIRPαV2, which was evident in the highly variable RO of GS‐0189 to SIRPα in samples from study patients, prompting further exploration using healthy donor samples. Genotyping for SIRPα variants in healthy donors coupled with binding affinity and RO data confirmed that GS‐0189 binding was weaker to SIRPαV2 than SIRPαV1; and the heterogeneity in the study patient RO findings could be explained by the genotyping of study patients. Concordant with SIRPα‐binding affinities, induction of phagocytosis by GS‐0189 as a single agent differed by more than 2 orders of magnitude between macrophages from donors expressing SIRPαV1/V1 and those expressing SIRPαV1/V2 or SIRPαV2/V2 . KWAR23 induced phagocytosis of the DLD‐1 human colorectal cancer cell line across SIRPα variants with equivalent potencies, suggesting that there is no inherent difference in phagocytic capacity of macrophages from patients with different SIRPα variants. Comparable ability of GS‐0189 combined with cetuximab to potentiate phagocytosis by PBMC‐derived macrophages from SIRPαV1/V1‐ and SIRPαV2/V2‐expressing donors was previously demonstrated using HT‐29 cells, the epidermal growth factor receptor–expressing human tumor cell line, as the target cell [V1/V2 and SIRPαV2/V2 genotypes.Affinity of GS‐0189 for the SIRPαGS‐0189 was intended as a low anemia‐risk alternative to magrolimab. With priming‐dose regimen and clinical management, acute anemia observed with magrolimab is no longer expected to limit magrolimab clinical development , 30. TheMN, NLB, SI, LP, AS, JB, VG, TC, and MP conceived of or designed the study. MN, LP, JB, YL, and MP acquired and provided data.All authors analyzed or interpreted the data, drafted, or critically reviewed the manuscript, approved the final version, and agreed to be accountable for all aspects of the work.Mayur Narkhede reports institutional research funding from EUSA Therapeutics, Genmab, and Roche‐Genentech; and attending advisory boards and honoraria from ADC Therapeutics, T.G. Therapeutics, and Seagen. Nancy L. Bartlett reports research funding from ADC Therapeutics, Affimed, Bristol Meyers Squibb, Celgene, Forty‐Seven, Gilead Sciences, Inc., Immune Design, Janssen, Kite Pharma, Merck, Millennium, Pfizer, Pharmacyclics, Roche‐Genentech, and Seattle Genetics. Sami Ibrahimi reports stock in Karyopharm Therapeutics. Leslie Popplewell reports no conflict of interest. Anna Seto, Jamie Bates, Yeonju Lee, Vaishnavi Ganti, Ling Han, and Tianling Chen all report employment with and stock in Gilead Sciences, Inc. Manish Patel reports leadership at ION Pharma; honoraria from Pfizer, Pharmacyclics, Bayer, Janssen Oncology, Genentech, and Adaptive Biotechnologies; consulting or advisory roles for Pharmacyclics/Janssen and Pfizer/EMD Serono; attending speakers’ bureaus for Exelixis, Roche‐Genentech, Taiho Pharmaceutical, and Celgene; and research funding from Acerta Pharma, ADC Therapeutics, Agenus, Aileron Therapeutics, AstraZeneca, BioNTech AG, Boehringer Ingelheim, Celgene, Checkpoint Therapeutics, CicloMed, Clovis Oncology, Cyteir Therapeutics, Daiichi Sankyo, Lilly, EMD Serono, Evelo Therapeutics, FORMA Therapeutics, Roche‐Genentech, Gilead Sciences, Inc., GlaxoSmithKline, H3 Biomedicine, Hengrui Therapeutics, Hutchison MediPharma, Ignyta, Incyte, Jacobio, Janssen, Klus Pharma, Kymab, Loxo, LSK Biopartners, Lycera, Macrogenics, Merck, Millennium, Mirati Therapeutics, Moderna Therapeutics, Pfizer, Placon, Portola Pharmaceuticals, Prelude Therapeutics, Ribon Therapeutics, Seven and Eight Biopharmaceuticals, Syndax, Taiho Pharmaceutical, Takeda, Tesaro, TopAlliance Biosciences Inc., Vigeo, ORIC, Artios, Treadwell, Mabspace, IgM Biosciences, Puretech, BioTheryX, Black Diamond Therapeutics, NGM Biopharmaceuticals, Novartis, Nurix, Relay Therapeutics, Samumed, Silicon Therapeutics, TeneoBio, Zymeworks, Olema, Adagene, Astellas, NGM, Accutar Biotech, TeneoBio, Compugen, Immunogen, and Blueprint Pharmaceuticals.Supporting InformationClick here for additional data file.


Obstructive hydrocephalus in adults can result from various etiologies, including rare cystic lesions such as tectal plate cysts. To depict a unique case of a tectal plate cyst causing hydrocephalus in an adult accompanied by persistent headaches, visual disturbances, and balance problems. In a clinical context, a 43-year-old female patient presented with a 2-week history of persistent headaches, accompanied by symptoms of dizziness, visual disturbances, and impaired balance. These headaches had exhibited a daily aggravation pattern over a year and were associated with concurrent manifestations of nausea, vomiting, and diplopia. Subsequent neuroimaging through a brain computed tomography (CT) scan disclosed the presence of hydrocephalus. Consultation with a neurologist and brain magnetic resonance imaging (MRI) yielded a diagnosis implicating a tectal plate cyst as the causative agent behind the obstructive hydrocephalus. The patient subsequently underwent surgical excision of the cyst. A follow-up assessment postoperation unveiled a marked improvement in the patient's clinical condition, characterized by the resolution of visual and gait impairments, as well as a notable reduction in the frequency and severity of headaches. This case highlights the importance of considering tectal plate cysts as an uncommon cause of hydrocephalus in the differential diagnosis of patients with persistent headaches and neurological symptoms. Early diagnosis and treatment with surgical removal of the cyst can significantly improve the patient's symptoms and prevent further complications such as hydrocephalus. A tectal plate cyst is a rare form of brain cyst that can occur in the tectal plate region The etiology of tectal plate cysts remains enigmatic. While they are widely considered to have a congenital origin, the precise underlying mechanism remains elusive. To gain deeper insights into the pathogenesis and prevalence of tectal plate cysts, further investigative research is imperative. The optimal management strategy for tectal plate lesions lacks definitive consensus. While surgical excision may demonstrate efficacy in ameliorating symptoms and averting complications, such as hydrocephalus, alternative therapeutic modalities may exist. Subsequent studies are warranted to undertake comparative analyses and ascertain the most efficacious and least invasive treatment modalities.A 43-year-old woman presented to the emergency department with 2 weeks of persistent headaches, accompanied by dizziness, visual disturbances, and balance problems. She had been experiencing intermittent headaches for 1-2 hours daily over the past year, which had become continuous over the last month. These headaches worsened in the morning, gradually improving throughout the day, and were associated with nausea and vomiting. The patient noted that bending and coughing exacerbated the headache. Additionally, she reported double vision and balance issues but did not complain of any memory or concentration deficits. There was no significant medical, surgical, or family history.Upon admission, the patient exhibited stable hemodynamics, with a blood pressure of 150/100 mm Hg, a pulse rate of 75 bpm, no fever , and a respiratory rate of 16 breaths per minute. Ophthalmic examination revealed 20/20 visual acuity in both eyes, normal intraocular pressure (12 mm Hg), and an unremarkable anterior eye segment. Fundus examination showed no papilledema.Laboratory tests yielded no abnormalities. A brain computed tomography (CT) scan revealed hydrocephalus and cystic lesion in in midbrain. Consultation with a neurologist was recommended. Brain Magnetic resonance imaging (MRI) indicated a 21 × 18 × 20 mm cyst arising from the midbrain, enveloped by a thin rim of tectal plate tissue and exhibiting cerebrospinal fluid (CSF) signal characteristics across all pulse sequences. The cyst exerted pressure on the quadrigeminal cistern and compressed the cerebral aqueduct, resulting in significant obstructive hydrocephalus in the third, fourth, and lateral ventricles of the brain. The cyst appeared homogeneously hypointense on T1 and hyperintense on T2 sequences, with no diffusion restriction on diffusion-weighted imaging (DWI) or evidence of abnormal enhancement .Fig. 1SaConsidering these findings, the diagnosis was established as a tectal plate cyst causing obstructive hydrocephalus. Surgical excision of the cyst was performed, leading to a marked improvement in the patient's symptoms. Her recovery was uneventful, and she was discharged 1 week after the procedure. At a follow-up appointment 4 weeks postsurgery, the patient reported substantial symptom amelioration, including the resolution of visual and gait disturbances, as well as reduced headache frequency and severity. Physical examination revealed a steady gait and normal neurological findings. A follow-up brain MRI demonstrated the absence of residual cysts or hydrocephalus.The patient was advised to maintain close communication with her neurosurgeon and promptly report any recurrence of symptoms. Furthermore, she received guidance to avoid activities that could elevate intracranial pressure, such as strenuous exertion, heavy lifting, and participation in contact sports.Tectal plate cysts are uncommon developmental anomalies that originate from the tectal plate structure of the midbrain. In the region of the tectal plate, benign cysts are extremely uncommon. By restricting the flow of cerebrospinal fluid (CSF), these cysts can induce obstructive hydrocephalus. The vast majority of tectal plate cysts are asymptomatic and are incidentally discovered during brain imaging investigations. Depending on the size and location of the cysts, when they do produce symptoms, manifestations include hydrocephalus, abnormal gait, urinary incontinence, and deficits in cranial nerves Upon reviewing the manifestations listed, Hydrocephalus may be the predominant presentation. This particular condition can lead to the development of a variety of uncomfortable symptoms, such as headaches, nausea, vomiting, and visual disturbances. The exact incidence and prevalence of these cysts are not well established in the literature, but they are thought to be uncommon. Tectal plate cysts can occur at any age but are diagnosed more frequently in children and adolescents. They are marginally more prevalent in men than women In contrast to previous research, we are presenting a case study involving a middle-aged female who has exhibited neurological symptoms. Upon reviewing the literature, we found only 3 publications representing tectal plate cysts. Vandertop et al. reported the case of a 10-year-old boy who presented with headaches and blurred vision. Although the boy did not have double vision or difficulty moving his eyes, a small tectal plate cyst was the cause of his obstructive hydrocephalus In another study conducted by Kumar et al., 2 cases were reported. One was a 36-year-old male with intermittent headaches for 12-18 months who experienced short-sighted vision and difficulty concentrating at work. He had anisocoria, nystagmus, and eyelid clonus. An MRI showed a cystic lesion from the tectal plate, obstructing the cerebral aqueduct and causing significant lateral and third ventricular enlargement. The other case exhibited a 40-year-old woman with dizzy spells, frequent falls, leg fatigue, and urinary urgency. She had a tonic downward gaze, nystagmus, and normal visual acuity. MRI revealed a tectal plate cyst causing aqueduct stenosis and hydrocephalus. No developmental anomalies were found The noticeable strength of this study was describing a unique case of a tectal plate cyst causing hydrocephalus in an adult with persistent headaches, visual disturbances, and balance problems. Although the patient underwent surgical intervention and improved significantly, A follow-up assessment of the patient's symptoms confirmed that the cyst had been completely removed. However, our findings should be interpreted by considering the following limitations: We did not have access to the patient's post-operative MRI images, so we relied solely on the report confirming the absence of complications and residual lesions.Written informed consent for publication was obtained from the patient.


Microbacterium foliorum NRRL B-24224. KillerTomato is a lytic cluster EE phage with a 17,442-bp genome and 68.6% GC content. Of 25 genes, 20 were assigned putative functions, including a putative tail assembly chaperone protein with a programmed frameshift and an endolysin.Discovered in Pocatello, Idaho, soil near a tomato garden, siphovirus KillerTomato infects  Microbacterium foliorum.The discovery and annotation of bacteriophages are an important step in the development of phage therapy strategies for fighting bacterial infections . Here, wMicrobacterium foliorum NRRL B-24224. After incubation with shaking at 20°C for 48 h, the culture was filtered and the filtrate was plated on PYCa agar with M. foliorum and incubated at 20°C for 48 h. Plaques formed by KillerTomato were purified through three additional rounds of plating . Transmission electron micrographs of KillerTomato revealed a siphovirus with a long tail .KillerTomato was found in garden soil on 1 September 2022 on the Idaho State University campus in Pocatello, Idaho , following established procedures . Soil saUsing a Promega Wizard DNA clean up kit, KillerTomato DNA was extracted from a high-titer lysate and prepared for sequencing using a NEBNext Ultra II FS kit. DNA was sequenced using a shotgun sequencing approach on an Illumina MiSeq (v3 reagents). Sequencing generated 650,649 single-end 150-bp reads with 5,321-fold coverage. The raw reads were trimmed and assembled with Newbler v2.9 using default parameters, yielding a single contig; Consend v29 was used to check for completeness and accuracy and determine phage termini, both using default settings 6. Killehttp://discover.kbrinsgd.org) and DNAmaster v5.23.6 (http://cobamide2.bio.pitt.edu) with manual inspection and confirmation. Genemark v2.5 (http://phages.wustl.edu/starterator/) was applied to compare start sites manually annotated for other EE phages to KillerTomato. HHPred v2.0 (The genome of KillerTomato was auto-annotated with PECAAN (ark v2.5  and Glimark v2.5  were useark v2.5  for clusred v2.0  (searchered v2.0  (searchered v2.0  and tRNAred v2.0 . Using dred v2.0 .Consistent with other EE phages, KillerTomato has a small genome with several fused genes . All but


Objective: To evaluate the cost-effectiveness of brentuximab vedotin in patients with R/R sALCL from a UK NHS perspective.Methods: A partitioned survival model used clinical outcomes for brentuximab vedotin from the pivotal phase-2 single-arm trial of brentuximab vedotin in 58 patients with R/R sALCL , over a lifetime (30-year) time horizon. Comparison with conventional chemotherapy was based on data from the Canadian British Columbia Cancer Agency registry from 40 patients starting salvage chemotherapy after front-line treatment between 1980 and 2012. Survival was extrapolated using parametric distributions, with brentuximab vedotin risk after the trial period assumed equal to conventional chemotherapy. Other modelling assumptions were based on a systematic literature review and clinical expert opinion.Results: Based on statistical extrapolation, brentuximab vedotin was associated with 3.1 years longer duration in the progression-free survival health state and an overall survival improvement of 5.4 years, prior to discounting. In addition, brentuximab vedotin was associated with 2.5 quality-adjusted life years (QALYs) gained at a total incremental cost of £88 556, resulting in an incremental cost-effectiveness ratio (ICER) of approximately £35 400. Sensitivity analyses of alternative model assumptions provided ICERs ranging from approximately £28 100 to £61 900. Comparing only first-line salvage patients reduced the ICER to £26 800 per QALY gained. Conversely, considering only patients with Eastern Corporative Oncology Group performance status of 0 or 1 increased the ICER to approximately £38 200. At a willingness-to-pay threshold of £50 000, the estimated probability that brentuximab vedotin is cost-effective compared with conventional chemotherapy was 86.5%.Conclusion: Compared to conventional chemotherapy, and considering the full survival period, brentuximab vedotin may provide a valuable treatment choice for patients with R/R sALCL, a population with limited therapeutic options.


Scientific Reports 10.1038/s41598-021-81948-4, published online 28 January 2021Correction to: The original version of this Article contained errors in the Discussion, in its interpretation of the effect of vapor pressure (VAP).Consequently, in the Discussion section,“Furthermore, the model also indicated that when drier atmospheric conditions (represented by VAP) were present and extended beyond the dry season into the flowering period, fruit production tended to be reduced.”now reads:“Furthermore, the model also indicated that when wetter atmospheric conditions (represented by VAP) were present and extended beyond the dry season into the flowering period, fruit production tended to be reduced.”“Over the 10-year study, VAP for 2017 production was the lowest ranked (26.27 hPa), and 2016 was the second lowest (25.37 hPa) (SI Table S2), signaling back-to-back years of persistent low atmospheric moisture.”now reads:“Over the 10-year study, VAP for 2017 production was the highest ranked (26.27 hPa), and 2016 was the second highest (25.37 hPa) during the DTF period; however, precipitation totals reveal that these were the 5th and 9th driest production years (SI Table S2), signaling back-to-back years of persistent high atmospheric moisture but low precipitation.”“Model 2 revealed that higher VAP levels (drier air) limited fruit production, but this was mitigated when trees had more sapwood area, particularly in Cachoeira . Sapwood mitigation of drought was also evident in Filipinas in 2013. After 2017, this was the second driest 6-month June–November (DTF) in our data, and in Filipinas, trees with large sapwood areas produced significantly more fruits than those with small. In sum, the more sapwood, the more likely any individual tree could continue producing fruits even in the face of a drier atmosphere.”now reads:“Model 2 revealed that higher VAP levels (more moist air) limited fruit production, but this was mitigated when trees had more sapwood area, particularly in Cachoeira . Sapwood mitigation of drought was also evident in Filipinas in 2013. After 2017, this was the second driest 6-month June–November (DTF) in our data, and in Filipinas, trees with large sapwood areas produced significantly more fruits than those with small. In sum, the more sapwood, the more likely any individual tree could continue producing fruits even in the face of atmospheric conditions.”The original Article has been corrected.


At follow-up, LBBAP seemed suboptimal, with a paced QRS duration of 185 ms with paced left bundle branch block (LBBB) morphology, possibly due to microdislodgment of the lead. We opted for implantation of a new LBBAP lead, with conventional BiV-CRT as bailout strategy. BiV-CRT was considered to be associated with increased risk because of a pseudoaneurysm compressing the coronary sinus as a complication of the previous NeoChord procedure. The new LBBAP lead was implanted according to the method previously described by Heckman et al.A 72-year-old man with ischemic cardiomyopathy and a previous NeoChord  and Tendyne  procedure for severe mitral regurgitation was referred to our device department. After implantation of a DDD pacemaker 2 years before referral with LBBAP because of second-degree AV block, his LV function had declined. Left ventricular ejection fraction (LVEF) decreased from 40% to 15%. The electrocardiogram recorded after initial implantation showed a paced qR morphology in lead VWith the availability of alternative pacing strategies such as LBBAP to conventional BiV-CRT in cardiac resynchronization therapy, patient selection is an important issue, even more so when both strategies can be combined with LOT-CRT.,BiV-CRT has proven to be an effective therapy in patients with heart failure and LBBB.,LOT-CRT has been described as an alternative pacing strategy to classic BiV-CRT and LBBAP when these strategies alone result in suboptimal cardiac resynchronization therapy.In this case report, we showed that stimulus-V6RWPT can be longer than expected in LBBAP with left bundle branch capture due to severe distal conduction system disease, resulting in LBBAP failing to correct dyssynchrony. In patients with nonselective LBBP and severe distal conduction system disease, LV septal myocardium and subsequent LV activation might be faster than LV activation via the (diseased) conduction system, in essence resulting in LV septal pacing only despite capture of the left bundle branch. LOT-CRT can be a good addition to LBBAP when the latter fails to correct electrical dyssynchrony.


In the current study, we addressed the effect of TNF-α (-308G/A) gene polymorphism on circulating TNF-α levels following continuous positive airway pressure (CPAP) therapy. Methods: This study was a secondary analysis of the RICCADSA trial (NCT00519597) conducted in Sweden. CAD patients with OSA (apnea–hypopnea index) of ≥15 events/h and an Epworth Sleepiness Scale (ESS) score of <10 were randomized to CPAP or no-CPAP groups, and OSA patients with an ESS score of ≥10 were offered CPAP treatment. Blood samples were obtained at baseline and 12-month follow-up visits. TNF-α was measured by immunoassay . Genotyping of TNF-α-308G/A (single nucleotide polymorphism Rs1800629) was performed by polymerase chain reaction–restriction fragment length polymorphism. Results: In all, 239 participants  years) with polymorphism data and circulating levels of TNF-α at baseline and 1-year follow-up visits were included. The median circulating TNF-α values fell in both groups between baseline and 12 months with no significant within- or between-group differences. In a multivariate linear regression model, a significant change in circulating TNF-α levels from baseline across the genotypes from GA to GA and GA to AA  −1.82; −0.12; p = 0.025) was observed in the entire cohort. The association was more pronounced among the individuals who were using the device for at least 4 h/night ; p = 0.004)), whereas no significant association was found among the patients who were non-adherent or randomized to no-CPAP. The participants carrying the TNF-α A allele were less responsive to CPAP treatment regarding the decline in circulating TNF-α despite CPAP adherence ; p = 0.005). Conclusions: Our results suggest that TNF-α (-308G/A) gene polymorphism is associated with changes in circulating TNF-α levels in response to CPAP treatment in adults with CAD and OSA.Rationale: We recently demonstrated that patients with coronary artery disease (CAD) and obstructive sleep apnea (OSA) carrying the tumor necrosis factor-alpha  Coronary artery disease (CAD) is associated with high mortality . The traObstructive sleep apnea (OSA) is characterized by intermittent upper airway collapse during sleep, causing sleep fragmentation and intermittent hypoxia . Almost Vascular inflammation plays a key role in the development of atherosclerotic plaques and CAD . It has TNF-α is a pro-inflammatory cytokine that is important for the immune system and plays a notable role in the development of autoimmune and infectious diseases as well as atherosclerosis and CAD [TNF-α also plays a crucial role in sleep regulation [TNF-α [TNF-α has received special attention in this context [TNF-α (position 308G/A); allele A at this position (TNF-α-308A) is suggested to be associated with a higher occurrence of OSA [TNF-α-308G/A (rs1800629) polymorphism with the risk of many diseases, such as allograft rejection [TNF-α-308A allele and obesity has also been reported [TNF-α-308G/A polymorphism and CAD. One study suggested that TNF-α-308G/A polymorphism is associated with ST-elevation myocardial infarction and high plasma levels of biochemical ischemia markers [TNF-α-308G/A and the risk of acute myocardial infarction [ and CAD . TNF-α agulation . Many OSn [TNF-α . Existinn [TNF-α , and TNF context ,20. There of OSA  as well e of OSA ,23,24,25ejection , asthma ejection , chronicejection , ischemiA rs18006 polymorpposition 8G/A; allejection . An assoreported ,21, wher markers , and a mfarction . On the farction .TNF-α A allele had increased circulating TNF-α levels compared with the ones carrying the TNF-α G allele [TNF-α (-308G/A) gene polymorphism on circulating TNF-α levels in response to 12 months of CPAP therapy.We recently demonstrated that patients with CAD and OSA carrying the G allele  in the “G allele . In the TNF-α-308G/A polymorphism study to evaluate the changes in circulating TNF-α levels from baseline to 12 months after CPAP treatment . Written informed consent was obtained from all participants. The main RICCADSA trial was registered with ClinicalTrials.gov (NCT 00519597).® Portable Digital System device  was used for the HSATs [2 exceeding 4% from the immediately preceding baseline was also recorded, and the oxygen desaturation index (ODI) was determined as the number of significant desaturations per hour.The Emblettahe HSATs . Apnea whe HSATs . The totThe ESS  was asse2 were defined as obese, and abdominal obesity was defined as a waist-to-hip ratio (WHR) of ≥0.9 for men and a WHR of ≥0.8 for women [Demographics, smoking habits, and medical history of the study cohort were obtained from the medical records. Individuals with a body mass index (BMI) of ≥30 kg/mor women .TNF-α levels were measured in the plasma samples (undiluted) using commercially available MILLIPLEX MAP (based on Luminex technology) human adipokine assay kits according to the manufacturer’s instructions . The assay sensitivities (minimum detectable levels) for TNF-α were 0.14 pg/mL, and all samples exhibited levels within the standard curve, covering a spectrum of 0 to 10,000 pg/mL. The intra-assay variability ranged from 1.4% to 7.9%, while the inter-assay variability was below 21% for the assessment of TNF-α concentrations. These values were calculated from the mean of the percentage coefficient of variability from multiple reportable results across two different concentrations of the samples in one experiment or from two results each for two different concentrations of samples across several different experiments.All blood samples were collected in the morning (07:00–08.00 am) after overnight fasting using EDTA (ethylenediaminetetraacetic acid) tubes, as previously described . The tubTNF-α promoter −308A/G (Rs1800629) genotyping analysis was performed by polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP), as previously described [As previously described in detail , genomicescribed .TNF-α levels, were tested by the Mann–Whitney test for continuous variables and the Chi-square test for categorical data. Within-group differences in changes from baseline in circulating TNF-α levels were tested by the Wilcoxon signed-rank test. A univariate linear regression analysis was performed to test the association between the change from baseline to the 12-month follow-up in circulating TNF-α levels and age, sex, ESS, BMI, WHR, AHI, ODI, OSA, and comorbidities, as well as TNF-α genotypes  and TNF-α alleles , respectively. Multivariate models included the same significant covariates as the univariate analysis as well as the variables of age, BMI, and sex in order to align with the recent guidelines [p-value of <0.05 was considered significant. Statistical analyses were performed using SPSS® 28.0 for Windows® .For descriptive statistics, variables were reported as medians with interquartile ranges (IQR) for continuous variables and as percentages for categorical variables. The Shapiro–Wilk test was used to test the normality assumption of the current data for all variables. Between-group differences stratified by CPAP allocation and CPAP usage in baseline characteristics, as well as changes from baseline in circulating idelines . All staTNF-α at baseline did not differ significantly between the groups.The study population consisted of 239 participants . As presented in TNF-α-GG was the most prevalent genotype in both groups, whereas TNF-α-AA in the CPAP group and TNF-α-GA in the no-CPAP group were the least frequent ones, respectively.As illustrated in TNF-α decreased from 4.87 (3.43–6.99) pg/mL to 4.62 (3.59–6.59) pg/mL in patients allocated to the CPAP group (p = 0.549) and from 5.15 (3.92–6.54) pg/mL to 4.50 (3.64–7.11) pg/mL in patients allocated to the no-CPAP group (p = 0.665), with no significant between-group differences in the magnitude of change from baseline.The median circulating levels of When analyzing the study population after stratifying by CPAP usage, the baseline characteristics did not differ significantly, except for ESS scores and the proportion of individuals with baseline EDS, which were higher among patients who used the device for at least 4 h/night during the first 12 months .TNF-α-GG was the most prevalent genotype and TNF-α-AA the least frequent one in both CPAP usage groups.As illustrated in TNF-α decreased from 4.84 (3.48–7.53) pg/mL to 4.72 (3.63–7.20) pg/mL in patients who used the device for at least 4 h/night (p = 0.577) and from 5.24 (3.59–6.85) pg/mL to 4.51 (3.50–6.75) pg/mL in patients allocated to the no-CPAP group or who used the device for less than 4 h/night (p = 0.199), with no significant between-group differences in the magnitude of change from baseline.As illustrated in TNF-α levels across the genotypes from GG to GA and GA to AA was observed in the entire cohort  gene polymorphism regarding the change in circulating levels of TNF-𝛼 from baseline. In other words, individuals with CAD and OSA carrying the TNF-α A allele seem to have an increased risk of elevated levels of circulating TNF-α, and the increased inflammatory activity is less likely to normalize despite CPAP use for at least 4 h/night. Whether or not those individuals should use the device even longer in order to reduce the levels of circulating TNF-α levels warrants further research. The clinical implications of our findings may also include that TNF-α-308G/A genotyping together with the analysis of TNF-α levels can be used for the prognostic evaluation of patients with CAD and concomitant OSA. For instance, patients carrying the A allele and higher levels of TNF-α could be included in a tighter follow-up scheme compared with those carrying the G allele.Circulating levels of inflammatory markers predict future cardiovascular events in the general population  as well controls ,48. It hcontrols ,14,15; tPAP arms . This waG allele . The TNFactivity . TNF-α iof TNF-α . Our curTNF-α (-308G/A) (rs1800629) polymorphism and not the TNF-α (-308G/A) (rs3611525) polymorphism. However, TNF-α-308G/A (rs1800629) and TNF-α-238G/A (rs361525) are located very close to each other and are hence tightly linked. In fact, in our sequencing results, we noticed that polymorphism in one 100% mirrored the other, and for the sake of simplicity, we chose to focus on and present only one of these SNPs. Accordingly, we propose that the TNF-α-308G/A (rs1800629) polymorphism analysis results can be extrapolated to TNF-α-238G/A (rs361525) SNP due to their tight linkage.We should acknowledge certain limitations. As also stated in the previous report , the powTNF-α-308G/A gene polymorphism was significantly correlated with the change in the circulating TNF-α levels from baseline in response to 12 months of CPAP treatment in this revascularized Swedish CAD cohort, independent of age, sex, BMI, baseline AHI, ESS, and diabetes. The participants carrying the TNF-α A allele were less responsive to CPAP treatment in terms of the decline in circulating TNF-α levels despite adequate CPAP adherence levels. Further prospective studies in larger cohorts and different geographical locations are warranted in order to better clarify whether the combined gentrifying and protein level analysis can be used as a prognostic biomarker for improved clinical follow-up of patients with CAD and concomitant OSA.We conclude that