Small Molecule Targets TMED9 and Promotes Lysosomal Degradation to Reverse Proteinopathy.

Intracellular accumulation of misfolded proteins causes toxic proteinopathies, diseases without targeted therapies. Mucin 1 kidney disease (MKD) results from a frameshift mutation in the MUC1 gene (MUC1-fs). Here, we show that MKD is a toxic proteinopathy. Intracellular MUC1-fs accumulation activated the ATF6 unfolded protein response (UPR) branch. We identified BRD4780, a small molecule that clears MUC1-fs from patient cells, from kidneys of knockin mice and from patient kidney organoids. MUC1-fs is trapped in TMED9 cargo receptor-containing vesicles of the early secretory pathway. BRD4780 binds TMED9, releases MUC1-fs, and re-routes it for lysosomal degradation, an effect phenocopied by TMED9 deletion. Our findings reveal BRD4780 as a promising lead for the treatment of MKD and other toxic proteinopathies. Generally, we elucidate a novel mechanism for the entrapment of misfolded proteins by cargo receptors and a strategy for their release and anterograde trafficking to the lysosome.

Molecular basis for substrate recruitment to the PRMT5 methylosome.

PRMT5 is an essential arginine methyltransferase and a therapeutic target in MTAP-null cancers. PRMT5 uses adaptor proteins for substrate recruitment through a previously undefined mechanism. Here, we identify an evolutionarily conserved peptide sequence shared among the three known substrate adaptors (CLNS1A, RIOK1, and COPR5) and show that it is necessary and sufficient for interaction with PRMT5. We demonstrate that PRMT5 uses modular adaptor proteins containing a common binding motif for substrate recruitment, comparable with other enzyme classes such as kinases and E3 ligases. We structurally resolve the interface with PRMT5 and show via genetic perturbation that it is required for methylation of adaptor-recruited substrates including the spliceosome, histones, and ribosomal complexes. Furthermore, disruption of this site affects Sm spliceosome activity, leading to intron retention. Genetic disruption of the PRMT5-substrate adaptor interface impairs growth of MTAP-null tumor cells and is thus a site for development of therapeutic inhibitors of PRMT5.

The Risk of Sympathetic Ophthalmia Associated with Open-Globe Injury Management Strategies: A Meta-analysis.

TOPIC: Sympathetic ophthalmia (SO) is a sight-threatening granulomatous panuveitis caused by a sensitizing event. Primary enucleation or primary evisceration, versus primary repair, as a risk management strategy after open-globe injury (OGI) remains controversial. CLINICAL RELEVANCE: This systematic review was conducted to report the incidence of SO after primary repair compared with that of after primary enucleation or primary evisceration. This enabled the reporting of an estimated number needed to treat. METHODS: Five journal databases were searched. This review was registered with International Prospective Register of Systematic Reviews (identifier, CRD42021262616). Searches were carried out on June 29, 2021, and were updated on December 10, 2022. Prospective or retrospective studies that reported outcomes (including SO or lack of SO) in a patient population who underwent either primary repair and primary enucleation or primary evisceration were included. A systematic review and meta-analysis were carried out in accordance with Preferred Reporting Items for Systematic Reviews and Meta-analyses guidelines. Random effects modelling was used to estimate pooled SO rates and absolute risk reduction (ARR). RESULTS: Eight studies reporting SO as an outcome were included in total. The included studies contained 7500 patients and 7635 OGIs. In total, 7620 OGIs met the criteria for inclusion in this analysis; SO developed in 21 patients with OGI. When all included studies were pooled, the estimated SO rate was 0.12% (95% confidence interval [CI], 0.00%-0.25%) after OGI. Of 779 patients who underwent primary enucleation or primary evisceration, no SO cases were reported, resulting in a pooled SO estimate of 0.05% (95% CI, 0.00%-0.21%). For primary repair, the pooled estimate of SO rate was 0.15% (95% CI, 0.00%-0.33%). The ARR using a random effects model was -0.0010 (in favour of eye removal; 95% CI, -0.0031 [in favor of eye removal] to 0.0011 [in favor of primary repair]). Grading of Recommendations, Assessment, Development, and Evaluations analysis highlighted a low certainty of evidence because the included studies were observational, and a risk of bias resulted from missing data. DISCUSSION: Based on the available data, no evidence exists that primary enucleation or primary evisceration reduce the risk of secondary SO. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Should We Stitch-Close the Fetoscopic Percutaneous Access? A Case-Series of Laparotomy to Trans-Amniotic Membrane Suturing for Intrauterine Port Placement in Fetoscopic Surgery for Twins.

INTRODUCTION: Maternal laparotomy-assisted fetoscopic surgery for in-utero myelomeningocele repair has shown that a trans-amniotic membrane suture during fetoscopic port placement can reduce postsurgical complications. Fetoscopic laser photocoagulation (FLP) for complex twins is typically performed percutaneously without a transmembrane stitch. However, in scenarios without a placental-free window, maternal laparotomy may be used for recipient sac access. Here, we present the outcomes of our series of laparotomy-assisted FLP cases, including a trans-amniotic membrane suturing of the fetoscopic port. METHODS: Retrospective series of twin-twin transfusion syndrome or twin anemia-polycythemia sequence (TAPS) cases treated at 2 fetal centers that underwent maternal laparotomy to FLP from September 2017 to January 2023. We recorded preoperative and operative characteristics, as well as pregnancy and neonatal outcomes. RESULTS: During the study period, 9 maternal laparotomy to FLP cases were performed. Two were excluded for prior percutaneous FLP in the pregnancy. The remaining seven utilized a maternal laparotomy to trans-amniotic membrane stitch with confirmation of proper suture placement under ultrasound guidance, and all surgeries were performed with a single 10 F Check-Flo® cannula. Mean gestational age (GA) at surgery was 19.1 weeks (range 16 weeks 4 days-23 weeks 3 days), with delivery occurring at a mean GA of 35.0 weeks (range 32 weeks 0 days-37 weeks 1 day), resulting in a mean latency of 15.8 weeks, significantly longer than what is reported in the literature and our own data (mean latency for percutaneous FLP 10.2, 95% CI 9.9-10.5). Furthermore, all cases underwent iatrogenic delivery before labor onset, with the lone delivery prior to 34 weeks due to concern for post-laser TAPS. CONCLUSION: This case series of laparotomy to FLP with trans-amniotic stitch, demonstrated no cases of spontaneous preterm birth and a longer-than-expected latency from surgery to delivery. Larger studies are warranted to investigate this approach.

Association of amnioinfusion volume at the time of surgery for twin-twin transfusion syndrome and latency to delivery.

OBJECTIVE: To evaluate the impact of amnioinfusion and other peri-operative factors on pregnancy outcomes in the setting of Twin-twin transfusion syndrome (TTTS) treated via fetoscopic laser photocoagulation (FLP). METHODS: Retrospective study of TTTS treated via FLP from 2010 to 2019. Pregnancies were grouped by amnioinfusion volume during FLP (<1 L vs. ≥1 L). The primary outcome was latency from surgery to delivery. An amnioinfusion statistic (AIstat) was created for each surgery based on the volume of fluid infused and removed and the preoperative deepest vertical pocket. Regression analysis was planned to assess the association of AIstat with latency. RESULTS: Patients with amnioinfusion of ≥1 L at the time of FLP had decreased latency from surgery to delivery (61 ± 29.4 vs. 73 ± 28.8 days with amnioinfusion <1 L, p < 0.001) and increased preterm prelabor rupture of membranes (PPROM) <34 weeks (44.7% vs. 33.5%, p = 0.042). Amnioinfusion ≥1 L was associated with an increased risk of delivery <32 weeks (aRR 2.6, 95% CI 1.5-4.5), 30 weeks (aRR 2.4, 95% CI 1.5-3.8), and 28 weeks (aRR 1.9, 95% CI 1.1-2.3). Cox-proportional regression revealed that AIstat was inversely associated with latency (HR 1.1, 95% CI 1.1-1.2). CONCLUSION: Amnioinfusion ≥1 L during FLP was associated with decreased latency after surgery and increased PPROM <34 weeks.

Reconstructing sexual divisions of labor from fingerprints on Ancestral Puebloan pottery.

An understanding of the division of labor in different societies, and especially how it evolved in the human species, is fundamental to most analyses of social, political, and economic systems. The ability to reconstruct how labor was organized, however, especially in ancient societies that left behind few material remains, is challenged by the paucity of direct evidence demonstrating who was involved in production. This is particularly true for identifying divisions of labor along lines of age, sex, and gender, for which archaeological interpretations mostly rely upon inferences derived from modern examples with uncertain applicability to ancient societies. Drawing upon biometric studies of human fingerprints showing statistically distinct ridge breadth measurements for juveniles, males, and females, this study reports a method for collecting fingerprint impressions left on ancient material culture and using them to distinguish the sex of the artifacts' producers. The method is applied to a sample of 985 ceramic sherds from a 1,000-y-old Ancestral Puebloan community in the US Southwest, a period characterized by the rapid emergence of a highly influential religious and political center at Chaco Canyon. The fingerprint evidence demonstrates that both males and females were significantly involved in pottery production and further suggests that the contributions of each sex varied over time and even among different social groups in the same community. The results indicate that despite long-standing assumptions that pottery production in Ancient Puebloan societies was primarily a female activity, labor was not strictly divided and instead was likely quite dynamic.

Influence of Gestational Weight Gain on the Risk of Preterm Birth for Underweight Women Living in Food Deserts.

OBJECTIVE: Preterm birth (PTB) and food insecurity are two of the most significant public health crises in the United States. Effects of being underweight among populations with low food security are not well understood. We assess whether the protective effect of gestational weight gain (GWG) for women with low prepregnancy body mass index (BMI) differs by accessibility to sources of healthy food. STUDY DESIGN: Population-based retrospective cohort study using Ohio birth records analyzing all live births, 2006 to 2015. Analyses were stratified by maternal BMI (underweight, normal, overweight, and obese), Institute of Medicine (IOM) recommended GWG (under vs. met), and whether the U.S. Department of Agriculture (USDA) classified the residential census tract for each birth as a food desert. Food access data were retrieved from the USDA's 2018 Food Access Research Atlas. Covariates were selected using least absolute shrinkage and selection operator regression. Logistic regression models estimated the risk ratio (RR) of PTB for each group based on under or exceeded recommended GWG (reference = met), adjusting for coexisting risk factors. RESULTS: Analysis was performed on 1,124,299 births. PTB risk was highest for underweight women below GWG recommendations (no food desert: 21.3%, RR = 2.15, 95% confidence interval [CI]: 1.81-2.57; food desert: 21.0%, RR = 1.46, 95% CI: 0.96-2.21). Underweight women living in food deserts who exceeded GWG recommendations had lower PTB risk than those who met GWG recommendations (13.5 vs. 14.3%, RR = 0.85, 95% CI: 0.51-1.41). Factors other than GWG significantly associated with PTB included in the adjusted analysis include maternal age and race, education, marital status, interpregnancy interval, and presence of prepregnancy diabetes or hypertension. CONCLUSION: Underweight women who do not meet GWG recommendations are at high risk for PTB. Increasing pregnancy weight gain to a level that exceeds IOM recommendations was not associated with a reduction in PTB risk for underweight women who reside in food deserts compared with women who met GWG recommendations. KEY POINTS: · Women with low prepregnancy BMI are at high risk of PTB.. · Food insecurity increases the risk of PTB for underweight women.. · Excessive GWG for underweight women in food deserts does not reduce PTB risk..

Bacteria-mediated hypoxia functions as a signal for mosquito development.

Mosquitoes host communities of microbes in their digestive tract that consist primarily of bacteria. We previously reported that several mosquito species, including Aedes aegypti, do not develop beyond the first instar when fed a nutritionally complete diet in the absence of a gut microbiota. In contrast, several species of bacteria, including Escherichia coli, rescue development of axenic larvae into adults. The molecular mechanisms underlying bacteria-dependent growth are unknown. Here, we designed a genetic screen around E. coli that identified high-affinity cytochrome bd oxidase as an essential bacterial gene product for mosquito growth. Bioassays showed that bacteria in nonsterile larvae and gnotobiotic larvae inoculated with wild-type E. coli reduced midgut oxygen levels below 5%, whereas larvae inoculated with E. coli mutants defective for cytochrome bd oxidase did not. Experiments further supported that hypoxia leads to growth and ecdysone-induced molting. Altogether, our results identify aerobic respiration by bacteria as a previously unknown but essential process for mosquito development.

Reproductive Outcomes following Laser Surgery for Twin-Twin Transfusion Syndrome.

BACKGROUND: There are limited studies describing future reproductive outcomes in women who have had selective fetoscopic laser photocoagulation (SFLP) for twin-twin transfusion syndrome (TTTS). OBJECTIVE: Our study aims to compare reproductive outcomes following monochorionic multiple gestational pregnancies complicated by TTTS requiring SFLP to those not requiring SFLP. METHODS: This is a retrospective cohort study that analyzed records of patients who were evaluated at the Cincinnati Fetal Center (2007-2014) for monochorionic multiple gestations. A questionnaire regarding reproductive, obstetric, gynecologic, and psychological outcomes following the index pregnancy was administered to consented participants by electronic distribution. The data was compared between pregnancies with prior SFLP versus no prior SFLP. RESULTS: There was a higher response rate in the SFLP group (219/474, 46.2%) versus the referent group (62/187, 33.2%). The median interval between the index pregnancy and survey completion was 74 months and 46 months in the SFLP and referent groups, respectively. Approximately 38 and 37% of the women in the SFLP and referent groups attempted conception after the index pregnancy with a >90% pregnancy success rate in both groups. Use of assisted reproductive technology was highly prevalent in both the index and subsequent pregnancies, with no significant difference between the groups. Over 60% of the women in each group did not attempt future pregnancy. Of those, approximately 1 in 3 cited the outcome of the index pregnancy as the primary reason for not pursuing future conception. There were no significant differences in selected maternal-fetal complications and new-onset gynecologic problems. More than 1 in 4 women in both groups were diagnosed with a mental health disorder following the index pregnancy. CONCLUSION: SFLP does not appear to be associated with adverse reproductive, obstetric, or gynecologic outcomes. The data may help facilitate evidence-based counseling for this patient population.

The influence of interpregnancy interval on infant mortality.

BACKGROUND: In Ohio, the infant mortality rate is above the national average and the black infant mortality rate is more than twice the white infant mortality rate. Having a short interpregnancy interval has been shown to correlate with preterm birth and low birthweight, but the effect of short interpregnancy interval on infant mortality is less well established. OBJECTIVE: We sought to quantify the population impact of interpregnancy interval on the risk of infant mortality. STUDY DESIGN: This was a statewide population-based retrospective cohort study of all births (n = 1,131,070) and infant mortalities (n = 8152) using linked Ohio birth and infant death records from January 2007 through September 2014. For this study we analyzed 5 interpregnancy interval categories: 0-<6, 6-<12, 12-<24, 24-<60, and ≥60 months. The primary outcome for this study was infant mortality. During the study period, 3701 infant mortalities were linked to a live birth certificate with an interpregnancy interval available. We calculated the frequency and relative risk of infant mortality for each interval compared to a referent interval of 12-<24 months. Stratified analyses by maternal race were also performed. Adjusted risks were estimated after accounting for statistically significant and biologically plausible confounding variables. Adjusted relative risk was utilized to calculate the attributable risk percent of short interpregnancy intervals on infant mortality. RESULTS: Short interpregnancy intervals were common in Ohio during the study period. Of all multiparous births, 20.5% followed an interval of <12 months. The overall infant mortality rate during this time was 7.2 per 1000 live births (6.0 for white mothers and 13.1 for black mothers). Infant mortalities occurred more frequently for births following short intervals of 0-<6 months (9.2 per 1000) and 6-<12 months (7.1 per 1000) compared to 12-<24 months (5.6 per 1000) (P < .001 and <.001). The highest risk for infant mortality followed interpregnancy intervals of 0-<6 months (adjusted relative risk, 1.32; 95% confidence interval, 1.17-1.49) followed by interpregnancy intervals of 6-<12 months (adjusted relative risk, 1.16; 95% confidence interval, 1.04-1.30). Analysis stratified by maternal race revealed similar findings. Attributable risk calculation showed that 24.2% of infant mortalities following intervals of 0-<6 months and 14.1% with intervals of 6-<12 months are attributable to the short interpregnancy interval. By avoiding short interpregnancy intervals of ≤12 months we estimate that in the state of Ohio 31 infant mortalities (20 white and 8 black) per year could have been prevented and the infant mortality rate could have been reduced from 7.2-7.0 during this time frame. CONCLUSION: An interpregnancy interval of 12-60 months (1-5 years) between birth and conception of next pregnancy is associated with lowest risk of infant mortality. Public health initiatives and provider counseling to optimize birth spacing has the potential to significantly reduce infant mortality for both white and black mothers.

The clinical significance of an estimated fetal weight below the 10th percentile: a comparison of outcomes of <5th vs 5th-9th percentile.

BACKGROUND: The association between small-for-gestational-age (birthweight <10th percentile for gestational age) and neonatal morbidity is well established. Yet, there is a paucity of data on the relationship between suspected small for gestational age (sonographic-estimated fetal weight <10th percentile) at 2 thresholds and subsequent neonatal morbidity. OBJECTIVE: The objective of this study was to determine the relationship between sonographic-estimated fetal weight <5th percentile vs 5-9th percentile and neonatal morbidity. STUDY DESIGN: This retrospective study involved 5 centers and included nonanomalous, singletons with sonographic-estimated fetal weight <10th percentile for gestational age who delivered from 2009-2012. Composite neonatal morbidity included respiratory distress syndrome, proven sepsis, intraventricular hemorrhage grade III or IV, necrotizing enterocolitis, thrombocytopenia, seizures, or death. Odd ratios were adjusted for center, maternal age, race, body mass index at first visit, smoking status, use of alcohol, use of drugs, and neonatal gender. RESULTS: Of 834 women with suspected small-for-gestational-age fetuses, 513 (62%) had sonographic-estimated fetal weight <5th percentile, and 321 (38%) had sonographic-estimated fetal weight of 5-9th percentile for gestational age. At delivery, 81% of women with a suspected small-for-gestational-age fetus had a confirmed small-for-gestational-age fetus. In the group with a sonographic-estimated fetal weight <5th percentile, 59% of neonates had birthweight <5th percentile; in the group with a sonographic-estimated fetal weight 5-9th percentile, 41% had birthweight <5th percentile, and 36% had birthweight at 5-9th percentile. Neonatal intensive care unit admission differed significantly for those fetuses at <5th percentile (29%) compared with those fetuses at 5-9th percentile (15%; P<.001). The composite neonatal morbidity among the sonographic-estimated fetal weight <5th percentile group was higher than the sonographic-estimated fetal weight of 5-9th percentile group (31% vs 13%; adjusted odds ratio, 2.41; 95% confidence interval, 1.53-3.80). Similar findings were noted when the analysis was limited to sonographic-estimated fetal weight within 28 days of delivery (adjusted odds ratio, 2.22; 95% confidence interval, 1.34-3.67). CONCLUSION: Eight of 10 suspected small-for-gestational-age fetuses had birthweight <10th percentile for gestational age; the prediction of actual birthweight was more accurate in the <5th percentile group. Neonates with sonographic-estimated fetal weight of <5th percentile were more likely to be admitted to the neonatal intensive care unit and have complications than were those neonates with sonographic-estimated fetal weight of 5-9th percentile.

Evaluation of ecdysteroid antisera for a competitive enzyme immunoassay and extraction procedures for the measurement of mosquito ecdysteroids.

Ecdysteroid hormones regulate several aspects of insect development and reproduction. The predominant ecdysteroids produced by insects including mosquitoes are ecdysone (E) and 20-hydroxyecdysone (20E). The ability to measure E and 20E titers is essential for many studies, but few sensitive, low cost options are currently available for doing so. To address this deficiency, we developed a new enzyme-linked immunoassay (EIA). In the first part of the study, we compared the affinity of two new antisera named EAB25 and EAB27 to other available ecdysteroid antisera. EAB25 had a 27-fold higher affinity for 20E than E, while EAB27 had a four-fold higher affinity for 20E. In the second part of the study, EIA protocols were developed for analyzing E and 20E produced by the mosquito Aedes aegypti. Results indicated that pelts from fourth instar larvae and ovaries from blood-fed, adult females produced E and 20E. Methanol extraction in the presence of magnesium from whole body samples altered antibody recognition of E and 20E by EIA. However, extraction with 1-butanol and two organic/water phase separations eliminated this problem and improved assay performance. We conclude the new antisera used in the EIA provide a low-cost, flexible, and sensitive method for measuring E and 20E in insects.

The impact of fetal growth restriction on latency in the setting of expectant management of preeclampsia.

BACKGROUND: Fetal growth restriction is a common complication of preeclampsia. Expectant management for qualifying patients has been found to have acceptable maternal safety while improving neonatal outcomes. Whether fetal growth restriction influences the duration of latency during expectant management of preeclampsia is unknown. OBJECTIVE: The objective of the study was to determine whether fetal growth restriction is associated with a reduced interval to delivery in women with preeclampsia being expectantly managed prior to 34 weeks. STUDY DESIGN: We performed a retrospective cohort of singleton, live-born, nonanomalous deliveries at the University of Cincinnati Medical Center between 2008 and 2013. Patients were included in our analysis if they were diagnosed with preeclampsia prior to 34 completed weeks and if the initial management plan was to pursue expectant management beyond administration of steroids for fetal lung maturity. Two study groups were determined based on the presence or absence of fetal growth restriction. Patients were delivered when they developed persistent neurological symptoms, severe hypertension refractory to medical therapy, renal insufficiency, nonreassuring fetal status, pulmonary edema, or hemolysis elevated liver low platelet syndrome or when they reached 37 weeks if they remained stable without any other indication for delivery. Our primary outcome was the interval from diagnosis of preeclampsia to delivery, measured in days. Secondary outcomes included indications for delivery, rates of induction and cesarean delivery, development of severe morbidities of preeclampsia, and select neonatal outcomes. We performed a multivariate logistic regression analysis comparing those with fetal growth restriction with those with normally grown fetuses to determine whether there is an association between fetal growth restriction and a shortened interval to delivery, neonatal intensive care unit admission, prolonged neonatal stay, and neonatal mortality. RESULTS: A total of 851 patients met the criteria for preeclampsia, of which 199 met inclusion criteria, 139 (69%) with normal growth, and 60 (31%) with fetal growth restriction. Interval to delivery was significantly shorter in women with fetal growth restriction, median (interquartile range) of 3 (1.6) days vs normal growth, 5 (2.12) days, P < .001. The association between fetal growth restriction and latency less than 7 days remained significant, even after post hoc analysis controlling for confounding variables (adjusted odds ratio, 1.66 [95% confidence interval, 1.12-2.47]). There were no differences in the development of severe disease (85.9 vs 91.7%, P = .26), need for intravenous antihypertensive medications (47.1 vs 46.7%, P = .96), and the development of severe complications of preeclampsia (51.1 vs 42.9%, P = .30) in normally grown and growth-restricted fetuses, respectively. Fewer women with fetal growth restriction attained their scheduled delivery date, 3 of 60 (5.0%), compared with normally grown fetuses,12 of 139 (15.7%), P = .03. Admission to the neonatal intensive care unit, neonatal length of stay, and neonatal mortality were higher when there was fetal growth restriction; however, after a logistic regression analysis, these associations were no longer significant. CONCLUSION: Fetal growth restriction is associated with a shortened interval to delivery in women undergoing expectant management of preeclampsia when disease is diagnosed prior to 34 weeks. These data may be helpful in counseling patients regarding the expected duration of pregnancy, guiding decision making regarding administration of steroids and determining the need for maternal transport.

The role of a novel auxiliary pocket in bacterial phenylalanyl-tRNA synthetase druggability.

The antimicrobial activity of phenyl-thiazolylurea-sulfonamides against Staphylococcus aureus PheRS are dependent upon phenylalanine levels in the extracellular fluids. Inhibitor efficacy in animal models of infection is substantially diminished by dietary phenylalanine intake, thereby reducing the perceived clinical utility of this inhibitor class. The search for novel antibacterial compounds against Gram-negative pathogens led to a re-evaluation of this phenomenon, which is shown here to be unique to S. aureus. Inhibition of macromolecular syntheses and characterization of novel resistance mutations in Escherichia coli demonstrate that antimicrobial activity of phenyl-thiazolylurea-sulfonamides is mediated by PheRS inhibition, validating this enzyme as a viable drug discovery target for Gram-negative pathogens. A search for novel inhibitors of PheRS yielded three novel chemical starting points. NMR studies were used to confirm direct target engagement for phenylalanine-competitive hits. The crystallographic structure of Pseudomonas aeruginosa PheRS defined the binding modes of these hits and revealed an auxiliary hydrophobic pocket that is positioned adjacent to the phenylalanine binding site. Three viable inhibitor-resistant mutants were mapped to this pocket, suggesting that this region is a potential liability for drug discovery.

Novel broad-spectrum inhibitors of bacterial methionine aminopeptidase.

With increasing emergence of multi-drug resistant infections, there is a dire need for new classes of compounds that act through unique mechanisms. In this work, we describe the discovery and optimization of a novel series of inhibitors of bacterial methionine aminopeptidase (MAP). Through a high-throughput screening campaign, one azepinone amide hit was found that resembled the native peptide substrate and possessed moderate biochemical potency against three bacterial isozymes. X-ray crystallography was used in combination with substrate-based design to direct the rational optimization of analogs with sub-micromolar potency. The novel compounds presented here represent potent broad-spectrum biochemical inhibitors of bacterial MAP and have the potential to lead to the development of new medicines to combat serious multi-drug resistant infections.

Bioaccumulation of metals in three freshwater mussel species exposed in situ during and after dredging at a coal ash spill site (Tennessee Valley Authority Kingston Fossil Plant).

On December 22, 2008, a dike containing coal fly ash at the Tennessee Valley Authority Kingston Fossil Plant (TN, USA) failed, and within months, dredging operations began to remove ash-contaminated sediments. The purpose of this study was to investigate differences in the bioaccumulation of metals in three mussel species during and after dredging operations. Mussels were caged for approximately 1 year during dredging and after, and then mussel condition index values and As, Cd, Cr, Pb, Ni, Se, Hg, U, Fe, Mg, Al, Sb, Ba, Be, Co, Cu, Mn, Mo, Ag, Sr, Tl, V, and Zn concentrations in soft tissue were determined via inductively coupled plasma-mass spectrometery. Overall, the differences observed in metal bioaccumulation and mussel health suggest that mussels in the immediate downstream area of the dredging site may have been impacted, as evidenced by a significant decrease in mussel condition index values, but that this impact did not result in increased tissue concentrations of metals.

Can first aid training encourage individuals' propensity to act in an emergency situation? A pilot study.

OBJECTIVE: To explore the effect that different activities included in first aid training can have on an individual's propensity to act in a medical emergency. DESIGN: Additional pilot-developed activities were added to a core first aid training session to create six unique groups, including a control group where no activities were added. Participants rated their agreement to pre-identified fears following the course and scored their self-efficacy and willingness to act before, immediately after and 2 months after the course. Change values were compared between groups. SETTING: Three locations in the UK (community halls, schools). PARTICIPANTS: 554 members of the public were recruited using advertising and community groups. A deliberately broad demographic was sought and achieved using targeted approaches where a particular demographic was deficient. INTERVENTION: Each participant attended one British Red Cross first aid course lasting 2 h. MAIN OUTCOME MEASURES: The same questionnaire was completed by all participants before and after each course. Two months later all participants were asked a series of follow-up questions. RESULTS: All courses showed an increase in self-efficacy and willingness to act immediately following the course. The course, which included both factual information relevant to helping in an emergency and 'helper' identity activities, produced significantly more positive responses to pre-identified fears. CONCLUSIONS: Activities which allow the learner to explore and discuss behaviour in an emergency situation can effectively increase the learner's propensity to act. First aid education should be expanded to support the learner to develop both the skill and the will to help.

Identification of Small Molecule Inhibitors of PPM1D Using a Novel Drug Discovery Platform.

Protein phosphatase, Mg2+/Mn2+ dependent 1D (PPM1D), is a serine/threonine phosphatase that is recurrently activated in cancer, regulates the DNA damage response (DDR), and suppresses the activation of p53. Consistent with its oncogenic properties, genetic loss or pharmacologic inhibition of PPM1D impairs tumor growth and sensitizes cancer cells to cytotoxic therapies in a wide range of preclinical models. Given the therapeutic potential of targeting PPM1D specifically and the DDR and p53 pathway more generally, we sought to deepen our biological understanding of PPM1D as a drug target and determine how PPM1D inhibition differs from other therapeutic approaches to activate the DDR. We performed a high throughput screen to identify new allosteric inhibitors of PPM1D, then generated and optimized a suite of enzymatic, cell-based, and in vivo pharmacokinetic and pharmacodynamic assays to drive medicinal chemistry efforts and to further interrogate the biology of PPM1D. Importantly, this drug discovery platform can be readily adapted to broadly study the DDR and p53. We identified compounds distinct from previously reported allosteric inhibitors and showed in vivo on-target activity. Our data suggest that the biological effects of inhibiting PPM1D are distinct from inhibitors of the MDM2-p53 interaction and standard cytotoxic chemotherapies. These differences also highlight the potential therapeutic contexts in which targeting PPM1D would be most valuable. Therefore, our studies have identified a series of new PPM1D inhibitors, generated a suite of in vitro and in vivo assays that can be broadly used to interrogate the DDR, and provided important new insights into PPM1D as a drug target.

Discovery of inhibitors of 4'-phosphopantetheine adenylyltransferase (PPAT) to validate PPAT as a target for antibacterial therapy.

Inhibitors of 4'-phosphopantetheine adenylyltransferase (PPAT) were identified through high-throughput screening of the AstraZeneca compound library. One series, cycloalkyl pyrimidines, showed inhibition of PPAT isozymes from several species, with the most potent inhibition of enzymes from Gram-positive species. Mode-of-inhibition studies with Streptococcus pneumoniae and Staphylococcus aureus PPAT demonstrated representatives of this series to be reversible inhibitors competitive with phosphopantetheine and uncompetitive with ATP, binding to the enzyme-ATP complex. The potency of this series was optimized using structure-based design, and inhibition of cell growth of Gram-positive species was achieved. Mode-of-action studies, using generation of resistant mutants with targeted sequencing as well as constructs that overexpress PPAT, demonstrated that growth suppression was due to inhibition of PPAT. An effect on bacterial burden was demonstrated in mouse lung and thigh infection models, but further optimization of dosing requirements and compound properties is needed before these compounds can be considered for progress into clinical development. These studies validated PPAT as a novel target for antibacterial therapy.

Effect of allograft patch closure on incidence of spinal inclusion cyst formation following open fetal myelomeningocele repair.

OBJECTIVE: The aim of this study was to evaluate the incidence of spinal inclusion cyst (sIC) formation after open fetal myelomeningocele (fMMC) repair and the effect of dural patch closure. METHODS: The authors conducted a retrospective review of patients who underwent open fMMC repair at their institution between March 2011 and June 2020. All patients met the criteria for intervention defined by the Management of Myelomeningocele Study (MOMS). The primary outcomes investigated were development of sIC and need for surgical intervention. Secondary outcomes included need for CSF diversion, extent of reversal of hindbrain herniation, and ambulatory status. RESULTS: Of 56 patients who underwent open fMMC repair, 52 had adequate spinal imaging for review. Twelve of these patients (23%) developed sIC (95% CI 0.11-0.35). Six patients experienced symptoms and required surgical detethering with sIC resection. Six additional patients had evidence of sIC on surveillance MRI but remained asymptomatic. The authors found a statistically significant relationship between the use of a dural allograft patch and sIC formation (p = 0.05). In terms of sIC development, there was no statistically significant difference between patients who underwent primary closure and those who received an allograft at the level of the fascia (p = 0.34) or skin (p = 0.26). The rate of hydrocephalus requiring CSF diversion was 52%. Interestingly, 98% of patients had improvement in extent of hindbrain herniation. Dural patch closure did not have any effect on the rate of progressive hydrocephalus (p = 0.33) or degree of reversal of hindbrain herniation (p > 0.99). CONCLUSIONS: This study suggested that children with prenatally repaired MMC are at higher risk for development of sIC and associated symptoms than those who undergo postnatal repair. The presentation of symptoms was also earlier in these patients than previously reported after postnatal repair. The use of a dural allograft patch appears to have a positive correlation with sIC formation. Future investigations evaluating the incidence of sIC after fetoscopic MMC repair, in which primary dural closure typically cannot be achieved and a dural patch is most often utilized, will be helpful in facilitating prenatal counseling for patients considering fetal intervention.