Non-coding Transcription Instructs Chromatin Folding and Compartmentalization to Dictate Enhancer-Promoter Communication and T Cell Fate.

It is now established that Bcl11b specifies T cell fate. Here, we show that in developing T cells, the Bcl11b enhancer repositioned from the lamina to the nuclear interior. Our search for factors that relocalized the Bcl11b enhancer identified a non-coding RNA named ThymoD (thymocyte differentiation factor). ThymoD-deficient mice displayed a block at the onset of T cell development and developed lymphoid malignancies. We found that ThymoD transcription promoted demethylation at CTCF bound sites and activated cohesin-dependent looping to reposition the Bcl11b enhancer from the lamina to the nuclear interior and to juxtapose the Bcl11b enhancer and promoter into a single-loop domain. These large-scale changes in nuclear architecture were associated with the deposition of activating epigenetic marks across the loop domain, plausibly facilitating phase separation. These data indicate how, during developmental progression and tumor suppression, non-coding transcription orchestrates chromatin folding and compartmentalization to direct with high precision enhancer-promoter communication.

ARAF mutations confer resistance to the RAF inhibitor belvarafenib in melanoma.

Although RAF monomer inhibitors (type I.5, BRAF(V600)) are clinically approved for the treatment of BRAFV600-mutant melanoma, they are ineffective in non-BRAFV600 mutant cells1-3. Belvarafenib is a potent and selective RAF dimer (type II) inhibitor that exhibits clinical activity in patients with BRAFV600E- and NRAS-mutant melanomas. Here we report the first-in-human phase I study investigating the maximum tolerated dose, and assessing the safety and preliminary efficacy of belvarafenib in BRAFV600E- and RAS-mutated advanced solid tumours (NCT02405065, NCT03118817). By generating belvarafenib-resistant NRAS-mutant melanoma cells and analysing circulating tumour DNA from patients treated with belvarafenib, we identified new recurrent mutations in ARAF within the kinase domain. ARAF mutants conferred resistance to belvarafenib in both a dimer- and a kinase activity-dependent manner. Belvarafenib induced ARAF mutant dimers, and dimers containing mutant ARAF were active in the presence of inhibitor. ARAF mutations may serve as a general resistance mechanism for RAF dimer inhibitors as the mutants exhibit reduced sensitivity to a panel of type II RAF inhibitors. The combination of RAF plus MEK inhibition may be used to delay ARAF-driven resistance and suggests a rational combination for clinical use. Together, our findings reveal specific and compensatory functions for the ARAF isoform and implicate ARAF mutations as a driver of resistance to RAF dimer inhibitors.

CCCTC-Binding Factor Translates Interleukin 2- and α-Ketoglutarate-Sensitive Metabolic Changes in T Cells into Context-Dependent Gene Programs.

Despite considerable research connecting cellular metabolism with differentiation decisions, the underlying mechanisms that translate metabolite-sensitive activities into unique gene programs are still unclear. We found that aspects of the interleukin-2 (IL-2)-sensitive effector gene program in CD4+ and CD8+ T cells in type 1 conditions (Th1) were regulated by glutamine and alpha-ketoglutarate (αKG)-induced events, in part through changes in DNA and histone methylation states. We further identified a mechanism by which IL-2- and αKG-sensitive metabolic changes regulated the association of CCCTC-binding factor (CTCF) with select genomic sites. αKG-sensitive CTCF sites were often associated with loci containing IL-2- and αKG-sensitive genome organization patterns and gene expression in T cells. IL-2- and αKG-sensitive CTCF sites in T cells were also associated with genes from developmental pathways that had αKG-sensitive expression in embryonic stem cells. The data collectively support a mechanism wherein CTCF serves to translate αKG-sensitive metabolic changes into context-dependent differentiation gene programs.

The E-Id Protein Axis Specifies Adaptive Lymphoid Cell Identity and Suppresses Thymic Innate Lymphoid Cell Development.

Innate and adaptive lymphoid development is orchestrated by the activities of E proteins and their antagonist Id proteins, but how these factors regulate early T cell progenitor (ETP) and innate lymphoid cell (ILC) development remains unclear. Using multiple genetic strategies, we demonstrated that E proteins E2A and HEB acted in synergy in the thymus to establish T cell identity and to suppress the aberrant development of ILCs, including ILC2s and lymphoid-tissue-inducer-like cells. E2A and HEB orchestrated T cell fate and suppressed the ILC transcription signature by activating the expression of genes associated with Notch receptors, T cell receptor (TCR) assembly, and TCR-mediated signaling. E2A and HEB acted in ETPs to establish and maintain a T-cell-lineage-specific enhancer repertoire, including regulatory elements associated with the Notch1, Rag1, and Rag2 loci. On the basis of these and previous observations, we propose that the E-Id protein axis specifies innate and adaptive lymphoid cell fate.

Spliceosomal disruption of the non-canonical BAF complex in cancer.

SF3B1 is the most commonly mutated RNA splicing factor in cancer1-4, but the mechanisms by which SF3B1 mutations promote malignancy are poorly understood. Here we integrated pan-cancer splicing analyses with a positive-enrichment CRISPR screen to prioritize splicing alterations that promote tumorigenesis. We report that diverse SF3B1 mutations converge on repression of BRD9, which is a core component of the recently described non-canonical BAF chromatin-remodelling complex that also contains GLTSCR1 and GLTSCR1L5-7. Mutant SF3B1 recognizes an aberrant, deep intronic branchpoint within BRD9 and thereby induces the inclusion of a poison exon that is derived from an endogenous retroviral element and subsequent degradation of BRD9 mRNA. Depletion of BRD9 causes the loss of non-canonical BAF at CTCF-associated loci and promotes melanomagenesis. BRD9 is a potent tumour suppressor in uveal melanoma, such that correcting mis-splicing of BRD9 in SF3B1-mutant cells using antisense oligonucleotides or CRISPR-directed mutagenesis suppresses tumour growth. Our results implicate the disruption of non-canonical BAF in the diverse cancer types that carry SF3B1 mutations and suggest a mechanism-based therapeutic approach for treating these malignancies.

Tailored leadership training in emergency medicine: qualitative exploration of the impact of the EMLeaders programme on consultants and trainees in England.

BACKGROUND: Emergency medicine (EM) consultants are expected to provide leadership to facilitate optimal clinical results, effective teamwork and learning. To foster leadership skills, the Emergency Medicine Leadership Programme (EMLeaders) was launched in 2018 by the Royal College of Emergency Medicine (RCEM), Health Education England and National Health Service England. A mixed-methods evaluation of EMLeaders was commissioned to assess the impact at the strategic, team and individual levels. This paper reports the qualitative evaluation component. METHODS: Qualitative data collected from 2021 to 2022 were drawn from an online survey of RCEM members in England, which included four open questions about leadership training. At the end of the survey, participants were asked to share contact details if willing to undertake an in-depth qualitative interview. Interviews explored perceptions of the programme and impact of curriculum design and delivery. Data were analysed thematically against the Kirkpatrick framework, providing in-depth understanding. RESULTS: There were 417 survey respondents, of whom 177 had participated in EMLeaders. Semistructured interviews were completed with 13 EM consultants, 13 trainees and 1 specialty and associate specialist doctor. EMLeaders was highly valued by EM consultants and trainees, particularly group interaction, expert facilitation and face-to-face practical scenario work. Consultant data yielded the themes: we believe in it; EM relevance is key; on a leadership journey; shaping better leaders; and a broken system. Challenges were identified in building engagement within a pressured workplace system and embedding workplace role modelling. Trainees identified behavioural shift in themselves following the programme but wanted more face-to-face discussions with senior colleagues. Key trainee themes included value in being together, storytelling in leadership, headspace for the leadership lens and survival in a state of collapse. CONCLUSION: The development of leadership skills in EM is considered important. The EMLeaders programme can support leadership learning but further embedding is needed.

Health Equity in Patients Receiving Durvalumab for Unresectable Stage III Non-Small Cell Lung Cancer in the US Veterans Health Administration.

BACKGROUND: Real-world evidence is limited regarding the relationship between race and use of durvalumab, an immunotherapy approved for use in adults with unresectable stage III non-small cell lung cancer (NSCLC) post-chemoradiotherapy (CRT). This study aimed to evaluate if durvalumab treatment patterns differed by race in patients with unresectable stage III NSCLC in a Veterans Health Administration (VHA) population. MATERIALS AND METHODS: This was a retrospective analysis of White and Black adults with unresectable stage III NSCLC treated with durvalumab presenting to any VHA facility in the US from January 1, 2017, to June 30, 2020. Data captured included baseline characteristics and durvalumab treatment patterns, including treatment initiation delay (TID), interruption (TI), and discontinuation (TD); defined as CRT completion to durvalumab initiation greater than 42 days, greater than 28 days between durvalumab infusions, and more than 28 days from the last durvalumab dose with no new durvalumab restarts, respectively. The number of doses, duration of therapy, and adverse events were also collected. RESULTS: A total of 924 patients were included in this study (White = 726; Black = 198). Race was not a significant factor in a multivariate logistic regression model for TID (OR, 1.39; 95% CI, 0.81-2.37), TI (OR, 1.58; 95% CI, 0.90-2.76), or TD (OR, 0.84; 95% CI, 0.50-1.38). There were also no significant differences in median (interquartile range [IQR]) number of doses (White: 15 [7-24], Black: 18 [7-25]; P = .25) or median (IQR) duration of therapy (White: 8.7 months [2.9-11.8], Black: 9.8 months [3.6-12.0]; P = .08), although Black patients were less likely to experience an immune-related adverse event (28% vs. 36%, P = .03) and less likely to experience pneumonitis (7% vs. 14%, P < .01). CONCLUSION: Race was not found to be linked with TID, TI, or TD in this real-world study of patients with unresectable stage III NSCLC treated with durvalumab at the VHA.

Rewriting the script: How COVID-19 affected the relation between intrinsic aspirations and depressive symptoms.

Self-determination theory proposes that intrinsic aspirations protect against negative mental health outcomes by satisfying people's basic psychological needs of autonomy, relatedness, and competence. The present study investigated this relationship using two four-wave prospective longitudinal studies which followed undergraduate students across the Canadian academic calendar (September to May). The first was conducted across 2018-19 and the second across 2019-20. By comparing these two samples, we examined whether baseline levels of intrinsic aspirations moderated the impact of the COVID-19 pandemic on the development of depressive symptoms. Three main findings emerged, the first being that students reported higher levels of depressive symptoms in Spring 2020 than in Spring 2019. Second, students with more intrinsic aspirations in the pre-pandemic sample (2018-19) experienced fewer depressive symptoms from December to May while students with more intrinsic aspirations in the pandemic sample (2019-20) experienced more depressive symptoms during this period. Lastly, the latter relationship was mediated by need frustration, whereby students with higher levels of intrinsic aspirations experienced greater need frustration during the pandemic year. Together, these findings suggest that although intrinsic aspirations typically protect against negative psychological outcomes, the unique need frustrating context of the pandemic made them a risk factor for depression.

Positive Allosteric Modulator of SERCA Pump NDC-1173 Exerts Beneficial Effects in Mouse Model of Alzheimer's Disease.

Alzheimer's disease (AD) is an irreversible neurodegenerative disease that affects millions of people worldwide. AD does not have a cure and most drug development efforts in the AD field have been focused on targeting the amyloid pathway based on the "amyloid cascade hypothesis". However, in addition to the amyloid pathway, substantial evidence also points to dysregulated neuronal calcium (Ca2+) signaling as one of the key pathogenic events in AD, and it has been proposed that pharmacological agents that stabilize neuronal Ca2+ signaling may act as disease-modifying agents in AD. In previous studies, we demonstrated that positive allosteric regulators (PAMs) of the Sarco/endoplasmic reticulum Ca2+ ATPase (SERCA) pump might act as such Ca2+ stabilizing agents. In the present study, we report the development of a novel SERCA PAM agent, compound NDC-1173. To test the effectiveness of this compound, we performed behavioral studies with the APP/PS1 transgenic AD mouse model. We also evaluated effects of this compound on expression of endoplasmic reticulum (ER) stress genes in the hippocampus of APP/PS1 mice. The results of this study support the hypothesis that the SERCA pump is a potential novel therapeutic drug target and that NDC-1173 is a promising lead molecule for developing disease-modifying agents in AD.

Early outcomes from a national Train-the-Trainer Interprofessional Team Development Program.

The imperative need to train health professions faculty (educators and clinicians) to lead interprofessional education efforts and promote interprofessional team-based care is widely recognized. This need stems from a growing body of research that suggests collaboration improves patient safety and health outcomes. This short report provides an overview of a Train-the-Trainer Interprofessional Team Development Program (T3 Program) that equips faculty leaders with the skills to lead interprofessional education and interprofessional collaborative practice across the learning continuum. We also describe the history, approach, and early outcomes of this innovative program.

Direct evidence that the GPCR CysLTR2 mutant causative of uveal melanoma is constitutively active with highly biased signaling.

Uveal melanoma is the most common eye cancer in adults and is clinically and genetically distinct from skin cutaneous melanoma. In a subset of cases, the oncogenic driver is an activating mutation in CYSLTR2, the gene encoding the G protein-coupled receptor cysteinyl-leukotriene receptor 2 (CysLTR2). The mutant CYSLTR2 encodes for the CysLTR2-L129Q receptor, with the substitution of Leu to Gln at position 129 (3.43). The ability of CysLTR2-L129Q to cause malignant transformation has been hypothesized to result from constitutive activity, but how the receptor could escape desensitization is unknown. Here, we characterize the functional properties of CysLTR2-L129Q. We show that CysLTR2-L129Q is a constitutively active mutant that strongly drives Gq/11 signaling pathways. However, CysLTR2-L129Q only poorly recruits ß-arrestin. Using a modified Slack-Hall operational model, we quantified the constitutive activity for both pathways and conclude that CysLTR2-L129Q displays profound signaling bias for Gq/11 signaling pathways while escaping ß-arrestin-mediated downregulation. CYSLTR2 is the first known example of a G protein-coupled receptor driver oncogene that encodes a highly biased constitutively active mutant receptor. These results provide new insights into the mechanism of CysLTR2-L129Q oncoprotein signaling and suggest CYSLTR2 as a promising potential therapeutic target in uveal melanoma.

Rapid Diagnostic Testing for Response to the Monkeypox Outbreak - Laboratory Response Network, United States, May 17-June 30, 2022.

As part of public health preparedness for infectious disease threats, CDC collaborates with other U.S. public health officials to ensure that the Laboratory Response Network (LRN) has diagnostic tools to detect Orthopoxviruses, the genus that includes Variola virus, the causative agent of smallpox. LRN is a network of state and local public health, federal, U.S. Department of Defense (DOD), veterinary, food, and environmental testing laboratories. CDC developed, and the Food and Drug Administration (FDA) granted 510(k) clearance* for the Non-variola Orthopoxvirus Real-time PCR Primer and Probe Set (non-variola Orthopoxvirus [NVO] assay), a polymerase chain reaction (PCR) diagnostic test to detect NVO. On May 17, 2022, CDC was contacted by the Massachusetts Department of Public Health (DPH) regarding a suspected case of monkeypox, a disease caused by the Orthopoxvirus Monkeypox virus. Specimens were collected and tested by the Massachusetts DPH public health laboratory with LRN testing capability using the NVO assay. Nationwide, 68 LRN laboratories had capacity to test approximately 8,000 NVO tests per week during June. During May 17-June 30, LRN laboratories tested 2,009 specimens from suspected monkeypox cases. Among those, 730 (36.3%) specimens from 395 patients were positive for NVO. NVO-positive specimens from 159 persons were confirmed by CDC to be monkeypox; final characterization is pending for 236. Prompt identification of persons with infection allowed rapid response to the outbreak, including isolation and treatment of patients, administration of vaccines, and other public health action. To further facilitate access to testing and increase convenience for providers and patients by using existing provider-laboratory relationships, CDC and LRN are supporting five large commercial laboratories with a national footprint (Aegis Science, LabCorp, Mayo Clinic Laboratories, Quest Diagnostics, and Sonic Healthcare) to establish NVO testing capacity of 10,000 specimens per week per laboratory. On July 6, 2022, the first commercial laboratory began accepting specimens for NVO testing based on clinician orders.

Development of Healthy Eating and Active Lifestyles for Diabetes, a culturally tailored diabetes self-management education and support programme for Black-British adults: A participatory research approach.

AIMS: To develop an evidence-based, culturally tailored, diabetes self-management education and support programme for Black-British adults, called Healthy Eating and Active Lifestyles for Diabetes (HEAL-D), using participatory methods to engage key stakeholders in the intervention design process. METHODS: Black-British adults living with type 2 diabetes, healthcare professionals and community leaders were engaged in an intervention development study. The intervention structure, format, content and delivery were developed through three phases of participatory research: Phase 1, formative research, involved focus groups and interviews; interactive co-development workshops were conducted in Phase 2; and Phase 3 focused on materials development. RESULTS: In Phase 1, focus groups and interviews identified the importance of nurturing collectivism, a reliance on informal sources of information/advice, barriers to attending appointments associated with competing priorities of work, travel and carer commitments, and a preference for directness and simple, clear advice/messages. A priority for healthcare professionals was the intervention embedding within current primary care structures and aligning with incentivised targets/metrics. Phase 2 (workshops) highlighted key requirements: avoidance of medical settings, appropriately trained and culturally knowledgeable educators, flexible appointments, preference for verbal and visual information and avoidance of technical/medical terminology. In Phase 3 (materials development), culturally sensitive videos, short films and information booklets were developed to convey educational messages, and food photography was used to provide culturally relevant dietary advice. CONCLUSIONS: Participatory methods provide a means to understand the needs of specific communities. This approach enables the development of healthcare interventions that are sensitive to the needs of service users and providers.

COVID-19 Vaccine Administration, by Race and Ethnicity - North Carolina, December 14, 2020-April 6, 2021.

COVID-19 has disproportionately affected non-Hispanic Black or African American (Black) and Hispanic persons in the United States (1,2). In North Carolina during January-September 2020, deaths from COVID-19 were 1.6 times higher among Black persons than among non-Hispanic White persons (3), and the rate of COVID-19 cases among Hispanic persons was 2.3 times higher than that among non-Hispanic persons (4). During December 14, 2020-April 6, 2021, the North Carolina Department of Health and Human Services (NCDHHS) monitored the proportion of Black and Hispanic persons* aged ≥16 years who received COVID-19 vaccinations, relative to the population proportions of these groups. On January 14, 2021, NCDHHS implemented a multipronged strategy to prioritize COVID-19 vaccinations among Black and Hispanic persons. This included mapping communities with larger population proportions of persons aged ≥65 years among these groups, increasing vaccine allocations to providers serving these communities, setting expectations that the share of vaccines administered to Black and Hispanic persons matched or exceeded population proportions, and facilitating community partnerships. From December 14, 2020-January 3, 2021 to March 29-April 6, 2021, the proportion of vaccines administered to Black persons increased from 9.2% to 18.7%, and the proportion administered to Hispanic persons increased from 3.9% to 9.9%, approaching the population proportion aged ≥16 years of these groups (22.3% and 8.0%, respectively). Vaccinating communities most affected by COVID-19 is a national priority (5). Public health officials could use U.S. Census tract-level mapping to guide vaccine allocation, promote shared accountability for equitable distribution of COVID-19 vaccines with vaccine providers through data sharing, and facilitate community partnerships to support vaccine access and promote equity in vaccine uptake.

Tandem Homologation-Acylation Chemistry: Single and Double Homologation.

Treatment of ß-dicarbonyls with the Furakawa-variant of the Simmons-Smith reagent results in homologation and production of an intermediate zinc enolate. Treatment of the enolate with various acylating agents generate products with both γ-dicarbonyl functionality and ß-dicarbonyl functionality. In situ exposure of the acylated product to additional zinc carbenoid effects a second regiospecific homologation reaction.

The role of goal-related autonomy: A self-determination theory analysis of perfectionism, poor goal progress, and depressive symptoms.

Adopting a self-determination theory perspective, this 3-wave longitudinal study explores the role of perfectionism in goal pursuit and the experience of depressive symptoms. The findings highlight the role of goal-related autonomy in mediating the opposite effects of self-critical and personal standards perfectionism on goal progress and depressive symptoms over the course of an academic year. The results suggest a way of understanding the pathway to depressive symptoms and poor goal progress in perfectionists. They point to a number of implications for clinical practice when working with self-critical perfectionists. Specifically, they indicate that interventions aimed at promoting autonomous motivation, may not only bolster goal progress but also act as a protective factor against depressive symptoms. Together, the results indicate that autonomous motivation is central to goal progress and suggest that low goal-related autonomy can be linked to negative outcomes. (PsycInfo Database Record (c) 2021 APA, all rights reserved).

A longitudinal investigation of trait-goal concordance on goal progress: The mediating role of autonomous goal motivation.

OBJECTIVES: The present study investigated the benefits of matching personality traits with goal type (i.e., agentic or communal) for goal progress. Autonomous motivation was examined as a mediator. METHODS: A multi-wave prospective longitudinal design was employed to track the progress that 935 university students made in their personal goal pursuits over an academic year. Participants set three personal goals at baseline and completed measures of personality and goal motivation. Participants' goals were coded as being either agentic or communal. Goal progress was assessed mid-year (T2) and at the end of the academic year (T3). Goal motivation was reassessed mid-year (T2). RESULTS: Conscientiousness was significantly related to making better progress on agentic, but not communal, goals. Conversely, Extraversion was related to making communal, but not agentic, goal progress. These trait-goal matching effects on progress were partially mediated by goal-specific motivation, suggesting that the selection of goals that matched one's traits resulted in higher autonomous motivation at the start of the academic year. CONCLUSIONS: The selection of trait concordant personal goals is associated with autonomous goal motivation and greater goal progress. This research integrates Self-Determination Theory with trait theories of personality to enhance our understanding of variations in goal success.

Smartphone applications available to pregnant women in the United Kingdom: An assessment of nutritional information.

The importance of diet during pregnancy is critically important for the short- and long-term health of both mother and child. The number of apps targeting pregnant women is rapidly increasing, yet the nutritional content of these tools remains largely unexplored. This review aimed to evaluate the coverage and content of nutrition information in smartphone apps available to U.K. pregnant women. Keyword searches were conducted in iTunes and Google Play stores in November 2018. Candidate apps were included if they targeted pregnant women, provided pregnancy-specific nutritional information, had a user rating of at least 4+ based on at least 20 ratings, and were available in English. Nutritional content was assessed for accuracy against U.K. recommendations. Behaviour change techniques (BCTs) were also evaluated. Twenty-nine apps were included, seven of which originated in the United Kingdom. There was a large variability in the quality of smartphone app nutritional information. The accuracy of nutrition information varied, and several apps conveyed inappropriate information for pregnancy. On average, 10 BCTs were identified per app (range 2-15). Overall, smartphone apps do not consistently provide accurate and useful nutritional information to pregnant women. This study highlights the need for the integration of evidence-based nutritional information during app development and for increased regulatory oversight. App developers should also make it clear that nutritional content is intended for a specific geographical region or population or modify for the intended audience. These are important considerations for the design of future apps, which are increasingly used to complement existing maternity services.

Mediterranean-style diet in pregnant women with metabolic risk factors (ESTEEM): A pragmatic multicentre randomised trial.

BACKGROUND: Pregnant women with metabolic risk factors are at high risk of complications. We aimed to assess whether a Mediterranean-style diet reduces adverse pregnancy outcomes in high-risk women. METHODS AND FINDINGS: We conducted a multicentre randomised trial in 5 maternity units (4 in London and 1 in Birmingham) between 12 September 2014 and 29 February 2016. We randomised inner-city pregnant women with metabolic risk factors (obesity, chronic hypertension, or hypertriglyceridaemia) to a Mediterranean-style diet with high intake of nuts, extra virgin olive oil, fruits, vegetables, nonrefined grains, and legumes; moderate to high consumption of fish; low to moderate intake of poultry and dairy products; low intake of red and processed meat; and avoidance of sugary drinks, fast food, and food rich in animal fat versus usual care. Participants received individualised dietary advice at 18, 20, and 28 weeks' gestation. The primary endpoints were composite maternal (gestational diabetes or preeclampsia) and composite offspring (stillbirth, small for gestational age, or admission to neonatal care unit) outcomes prioritised by a Delphi survey. We used an intention-to-treat (ITT) analysis with multivariable models and identified the stratification variables and prognostic factors a priori. We screened 7,950 and randomised 1,252 women. Baseline data were available for 593 women in the intervention (93.3% follow-up, 553/593) and 612 in the control (95.6% follow-up, 585/612) groups. Over a quarter of randomised women were primigravida (330/1,205; 27%), 60% (729/1,205) were of Black or Asian ethnicity, and 69% (836/1,205) were obese. Women in the intervention arm consumed more nuts (70.1% versus 22.9%; adjusted odds ratio [aOR] 6.8, 95% confidence interval [CI] 4.3-10.6, p ≤ 0.001) and extra virgin olive oil (93.2% versus 49.0%; aOR 32.2, 95% CI 16.0-64.6, p ≤ 0.001) than controls; increased their intake of fish (p < 0.001), white meat (p < 0.001), and pulses (p = 0.05); and reduced their intake of red meat (p < 0.001), butter, margarine, and cream (p < 0.001). There was no significant reduction in the composite maternal (22.8% versus 28.6%; aOR 0.76, 95% CI 0.56-1.03, p = 0.08) or composite offspring (17.3% versus 20.9%; aOR 0.79, 95% CI 0.58-1.08, p = 0.14) outcomes. There was an apparent reduction in the odds of gestational diabetes by 35% (aOR 0.65, 95% CI 0.47-0.91, p = 0.01) but not in other individual components of the composite outcomes. Mothers gained less gestational weight (mean 6.8 versus 8.3 kg; adjusted difference -1.2 Kg, 95% CI -2.2 to -0.2, p = 0.03) with intervention versus control. There was no difference in any of the other maternal and offspring complications between both groups. When we pooled findings from the Effect of Simple, Targeted Diet in Pregnant Women With Metabolic Risk Factors on Pregnancy Outcomes (ESTEEM) trial with similar trials using random effects meta-analysis, we observed a significant reduction in gestational diabetes (odds ratio [OR] 0.67, 95% CI 0.53-0.84, I2 = 0%), with no heterogeneity (2 trials, 2,397 women). The study's limitations include the use of participant reported tools for adherence to the intervention instead of objective biomarkers. CONCLUSIONS: A simple, individualised, Mediterranean-style diet in pregnancy did not reduce the overall risk of adverse maternal and offspring complications but has the potential to reduce gestational weight gain and the risk of gestational diabetes. TRIAL REGISTRATION: ClinicalTrials.gov NCT02218931.

Meeting the Challenge of Predicting Hepatic Clearance of Compounds Slowly Metabolized by Cytochrome P450 Using a Novel Hepatocyte Model, HepatoPac.

Generating accurate in vitro intrinsic clearance data is an important aspect of predicting in vivo human clearance. Primary hepatocytes in suspension are routinely used to predict in vivo clearance; however, incubation times have typically been limited to 4-6 hours, which is not long enough to accurately evaluate the metabolic stability of slowly metabolized compounds. HepatoPac is a micropatterened hepatocyte-fibroblast coculture system that can be used for continuous incubations of up to 7 days. This study evaluated the ability of human HepatoPac to predict the in vivo clearance (CL) of 17 commercially available compounds with low to intermediate clearance (<12 ml/min/kg). In vitro half-life for disappearance of each compound was converted to hepatic clearance using the well stirred model, with and without correction for plasma protein binding. Hepatic CL, using three individual donors, was accurately predicted for 11 of 17 compounds (59%; predicted clearance within 2-fold of observed human in vivo clearance values). The accuracy of prediction increased to 82% (14 of 17 compounds) with an acceptance criterion defined as within 3-fold. When considering only low clearance compounds (<5 ml/min per kg), which represented 10 of the 17 compounds, the accuracy of prediction was 70% within 2-fold and 100% within 3-fold. In addition, the turnover of three slowly metabolized compounds (alprazolam, meloxicam, and tolbutamide) in HepatoPac was directly compared with turnover in suspended hepatocytes. The turnover of alprazolam and tolbutamide was approximately 2-fold greater using HepatoPac compared with suspended hepatocytes, which was roughly in line with the extrapolated values (correcting for the longer incubation time and lower cell number with HepatoPac). HepatoPac, but not suspended hepatocytes, demonstrated significant turnover of meloxicam. These results demonstrate the utility of HepatoPac for prediction of in vivo hepatic clearance, particularly with low clearance compounds.