Stabilizing the Proteomes of Acute Myeloid Leukemia Cells: Implications for Cancer Proteomics.

Previous work has shown that inhibition of abundant myeloid azurophil granule-associated serine proteases (ELANE [neutrophil elastase], PRTN3 [protease 3], and CTSG [Cathepsin G]) is required to stabilize some proteins in myeloid cells. We therefore hypothesized that effective inhibition of these proteases may be necessary for quantitative proteomic analysis of samples containing myeloid cells. To test this hypothesis, we thawed viably preserved acute myeloid leukemia cells from cryovials in the presence or the absence of diisopropyl fluorophosphate (DFP), a cell-permeable and irreversible serine protease inhibitor. Global proteomic analysis was performed, using label-free and isobaric peptide-labeling quantitation. The presence of DFP resulted in an increase of tryptic peptides (14-57%) and proteins (9-31%). In the absence of DFP, 11 to 31% of peptide intensity came from nontryptic peptides; 52 to 75% had cleavage specificity consistent with activities of ELANE-PRTN3. Treatment with DFP reduced the intensity of nontryptic peptides to 4-8% of the total. ELANE inhibition was 95%, based on diisopropyl phosphate modification of active site serine residue. Overall, the relative abundance of 20% of proteins was significantly altered by DFP treatment. These results suggest that active myeloid serine proteases, released during sample processing, can skew quantitative proteomic measurements. Finally, significant ELANE activity was also detected in Clinical Proteomics Tumor Analysis Consortium datasets of solid tumors (many of which have known myeloid infiltration). In the pancreatic cancer dataset, the median percentage of nontryptic intensity detected across patient samples was 34%, with many patient samples having more than half of their detected peptide intensity from nontryptic cleavage events consistent with ELANE-PRTN3 cleavage specificity. Our study suggests that in vitro cleavage of proteins by myeloid serine proteases may be relevant for proteomic studies of any tumor that contains infiltrating myeloid cells.

Direct Proteomic Detection and Prioritization of 19 Onchocerciasis Biomarker Candidates in Humans.

Onchocerca volvulus, the causative agent of onchocerciasis, infects over 20 million people and can cause severe dermatitis and ocular conditions including blindness. Current treatments employed in mass drug administration programs do not kill adult female worms, and common diagnostic tests cannot reliably assess viability of adult worms. There is an urgent need for better diagnostic tests to facilitate monitoring the efficacy of new treatments and disease elimination efforts. Here, eight plasma samples collected from individuals infected with O. volvulus and seven from uninfected individuals were analyzed by MS/MS spectrometry to directly identify O. volvulus proteins present in infected but absent in uninfected control samples. This direct proteomic approach for biomarker discovery had not been previously employed for onchocerciasis. Among all detected proteins, 19 biomarker candidates were supported by two or more unique peptides, identified in the plasma of at least three O. volvulus-infected human samples and absent in all control samples. Comprehensive analysis and ranking of these candidates included detailed functional annotation and a review of RNA-seq gene expression profiles. Isotope-labeled standard peptides were run in parallel and validated MS/MS peptide identifications for 15 peptides from 11 of the 19 proteins, and two infected urine and one uninfected urine sample was used for additional validation. A major antigen/OVOC11613 was identified as the most promising candidate with eight unique peptides across five plasma samples and one urine sample. Additional strong candidates included OVOC1523/ATP synthase, OVOC247/laminin and OVOC11626/PLK5, and along with OVOC11613, and were also detected in urine samples from onchocerciasis patients. This study has identified a promising novel set of proteins that will be carried forward to develop assays that can be used for diagnosis of O. volvulus infections and for monitoring treatment efficacy.

Protein Kinase Signaling Networks Driven by Oncogenic Gq/11 in Uveal Melanoma Identified by Phosphoproteomic and Bioinformatic Analyses.

Metastatic uveal melanoma (UM) patients typically survive only 2 to 3 years because effective therapy does not yet exist. Here, to facilitate the discovery of therapeutic targets in UM, we have identified protein kinase signaling mechanisms elicited by the drivers in 90% of UM tumors: mutant constitutively active G protein α-subunits encoded by GNAQ (Gq) or GNA11 (G11). We used the highly specific Gq/11 inhibitor FR900359 (FR) to elucidate signaling networks that drive proliferation, metabolic reprogramming, and dedifferentiation of UM cells. We determined the effects of FR on the proteome and phosphoproteome of UM cells as indicated by bioinformatic analyses with CausalPath and site-specific gene set enrichment analysis. We found that inhibition of oncogenic Gq/11 caused deactivation of PKC, Erk, and the cyclin-dependent kinases CDK1 and CDK2 that drive proliferation. Inhibition of oncogenic Gq/11 in UM cells with low metastatic risk relieved inhibitory phosphorylation of polycomb-repressive complex subunits that regulate melanocytic redifferentiation. Site-specific gene set enrichment analysis, unsupervised analysis, and functional studies indicated that mTORC1 and 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 2 drive metabolic reprogramming in UM cells. Together, these results identified protein kinase signaling networks driven by oncogenic Gq/11 that regulate critical aspects of UM cell biology and provide targets for therapeutic investigation.

Proteins in Tumor-Derived Plasma Extracellular Vesicles Indicate Tumor Origin.

Cancer-derived extracellular vesicles (EVs) promote tumorigenesis, premetastatic niche formation, and metastasis via their protein cargo. However, the proteins packaged by patient tumors into EVs cannot be determined in vivo because of the presence of EVs derived from other tissues. We therefore developed a cross-species proteomic method to quantify the human tumor-derived proteome of plasma EVs produced by patient-derived xenografts of four cancer types. Proteomic profiling revealed individualized packaging of novel protein cargo, and machine learning accurately classified the type of the underlying tumor.

Proteomic and phosphoproteomic landscapes of acute myeloid leukemia.

We have developed a deep-scale proteome and phosphoproteome database from 44 representative acute myeloid leukemia (AML) patients from the LAML TCGA dataset and 6 healthy bone marrow-derived controls. After confirming data quality, we orthogonally validated several previously undescribed features of AML revealed by the proteomic data. We identified examples of posttranscriptionally regulated proteins both globally (ie, in all AML samples) and also in patients with recurrent AML driver mutations. For example, samples with IDH1/2 mutations displayed elevated levels of the 2-oxoglutarate-dependent histone demethylases KDM4A/B/C, despite no changes in messenger RNA levels for these genes; we confirmed this finding in vitro. In samples with NPMc mutations, we identified several nuclear importins with posttranscriptionally increased protein abundance and showed that they interact with NPMc but not wild-type NPM1. We identified 2 cell surface proteins (CD180 and MRC1/CD206) expressed on AML blasts of many patients (but not healthy CD34+ stem/progenitor cells) that could represent novel targets for immunologic therapies and confirmed these targets via flow cytometry. Finally, we detected nearly 30 000 phosphosites in these samples; globally, AML samples were associated with the abnormal phosphorylation of specific residues in PTPN11, STAT3, AKT1, and PRKCD. FLT3-TKD samples were associated with increased phosphorylation of activating tyrosines on the cytoplasmic Src-family tyrosine kinases FGR and HCK and related signaling proteins. PML-RARA-initiated AML samples displayed a unique phosphorylation signature, and TP53-mutant samples showed abundant phosphorylation of serine-183 on TP53 itself. This publicly available database will serve as a foundation for further investigations of protein dysregulation in AML pathogenesis.

Bariatric surgery alters mitochondrial function in gut mucosa.

BACKGROUND: The obesity pandemic has worsened global disease burden, including type 2 diabetes, cardiovascular disease, and cancer. Metabolic/bariatric surgery (MBS) is the most effective and durable obesity treatment, but the mechanisms underlying its long-term weight loss efficacy remain unclear. MBS drives substrate oxidation that has been linked to improvements in metabolic function and improved glycemic control that are potentially mediated by mitochondria-a primary site of energy production. As such, augmentation of intestinal mitochondrial function may drive processes underlying the systemic metabolic benefits of MBS. Herein, we applied a highly sensitive technique to evaluate intestinal mitochondrial function ex vivo in a mouse model of MBS. METHODS: Mice were randomized to surgery, sham, or non-operative control. A simplified model of MBS, ileal interposition, was performed by interposition of a 2-cm segment of terminal ileum into the proximal bowel 5 mm from the ligament of Treitz. After a four-week recovery period, intestinal mucosa of duodenum, jejunum, ileum, and interposed ileum were assayed for determination of mitochondrial respiratory function. Citrate synthase activity was measured as a marker of mitochondrial content. RESULTS: Ileal interposition was well tolerated and associated with modest body weight loss and transient hypophagia relative to controls. Mitochondrial capacity declined in the native duodenum and jejunum of animals following ileal interposition relative to controls, although respiration remained unchanged in these segments. Similarly, ileal interposition lowered citrate synthase activity in the duodenum and jejunum following relative to controls but ileal function remained constant across all groups. CONCLUSION: Ileal interposition decreases mitochondrial volume in the proximal intestinal mucosa of mice. This change in concentration with preserved respiration suggests a global mucosal response to segment specific nutrition signals in the distal bowel. Future studies are required to understand the causes underlying these mitochondrial changes.

External validation of prognostic models to predict stillbirth using International Prediction of Pregnancy Complications (IPPIC) Network database: individual participant data meta-analysis.

OBJECTIVE: Stillbirth is a potentially preventable complication of pregnancy. Identifying women at high risk of stillbirth can guide decisions on the need for closer surveillance and timing of delivery in order to prevent fetal death. Prognostic models have been developed to predict the risk of stillbirth, but none has yet been validated externally. In this study, we externally validated published prediction models for stillbirth using individual participant data (IPD) meta-analysis to assess their predictive performance. METHODS: MEDLINE, EMBASE, DH-DATA and AMED databases were searched from inception to December 2020 to identify studies reporting stillbirth prediction models. Studies that developed or updated prediction models for stillbirth for use at any time during pregnancy were included. IPD from cohorts within the International Prediction of Pregnancy Complications (IPPIC) Network were used to validate externally the identified prediction models whose individual variables were available in the IPD. The risk of bias of the models and cohorts was assessed using the Prediction study Risk Of Bias ASsessment Tool (PROBAST). The discriminative performance of the models was evaluated using the C-statistic, and calibration was assessed using calibration plots, calibration slope and calibration-in-the-large. Performance measures were estimated separately in each cohort, as well as summarized across cohorts using random-effects meta-analysis. Clinical utility was assessed using net benefit. RESULTS: Seventeen studies reporting the development of 40 prognostic models for stillbirth were identified. None of the models had been previously validated externally, and the full model equation was reported for only one-fifth (20%, 8/40) of the models. External validation was possible for three of these models, using IPD from 19 cohorts (491 201 pregnant women) within the IPPIC Network database. Based on evaluation of the model development studies, all three models had an overall high risk of bias, according to PROBAST. In the IPD meta-analysis, the models had summary C-statistics ranging from 0.53 to 0.65 and summary calibration slopes ranging from 0.40 to 0.88, with risk predictions that were generally too extreme compared with the observed risks. The models had little to no clinical utility, as assessed by net benefit. However, there remained uncertainty in the performance of some models due to small available sample sizes. CONCLUSIONS: The three validated stillbirth prediction models showed generally poor and uncertain predictive performance in new data, with limited evidence to support their clinical application. The findings suggest methodological shortcomings in their development, including overfitting. Further research is needed to further validate these and other models, identify stronger prognostic factors and develop more robust prediction models. © 2021 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology.

Proteogenomics connects somatic mutations to signalling in breast cancer.

Somatic mutations have been extensively characterized in breast cancer, but the effects of these genetic alterations on the proteomic landscape remain poorly understood. Here we describe quantitative mass-spectrometry-based proteomic and phosphoproteomic analyses of 105 genomically annotated breast cancers, of which 77 provided high-quality data. Integrated analyses provided insights into the somatic cancer genome including the consequences of chromosomal loss, such as the 5q deletion characteristic of basal-like breast cancer. Interrogation of the 5q trans-effects against the Library of Integrated Network-based Cellular Signatures, connected loss of CETN3 and SKP1 to elevated expression of epidermal growth factor receptor (EGFR), and SKP1 loss also to increased SRC tyrosine kinase. Global proteomic data confirmed a stromal-enriched group of proteins in addition to basal and luminal clusters, and pathway analysis of the phosphoproteome identified a G-protein-coupled receptor cluster that was not readily identified at the mRNA level. In addition to ERBB2, other amplicon-associated highly phosphorylated kinases were identified, including CDK12, PAK1, PTK2, RIPK2 and TLK2. We demonstrate that proteogenomic analysis of breast cancer elucidates the functional consequences of somatic mutations, narrows candidate nominations for driver genes within large deletions and amplified regions, and identifies therapeutic targets.

Can risk prediction models help us individualise stillbirth prevention? A systematic review and critical appraisal of published risk models.

BACKGROUND: Stillbirth prevention is an international priority - risk prediction models could individualise care and reduce unnecessary intervention, but their use requires evaluation. OBJECTIVES: To identify risk prediction models for stillbirth, and assess their potential accuracy and clinical benefit in practice. SEARCH STRATEGY: MEDLINE, Embase, DH-DATA and AMED databases were searched from inception to June 2019 using terms relevant to stillbirth, perinatal mortality and prediction models. The search was compliant with Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines. SELECTION CRITERIA: Studies developing and/or validating prediction models for risk of stillbirth developed for application during pregnancy. DATA COLLECTION AND ANALYSIS: Study screening and data extraction were conducted in duplicate, using the CHARMS checklist. Risk of bias was appraised using the PROBAST tool. RESULTS: The search identified 2751 citations. Fourteen studies reporting development of 69 models were included. Variables consistently included were: ethnicity, body mass index, uterine artery Doppler, pregnancy-associated plasma protein and placental growth factor. For almost all models there were significant concerns about risk of bias. Apparent model performance (i.e. in the development dataset) was highest in models developed for use later in pregnancy and including maternal characteristics, and ultrasound and biochemical variables, but few were internally validated and none were externally validated. CONCLUSIONS: Almost all models identified were at high risk of bias. There are first-trimester models of possible clinical benefit in early risk stratification; these require validation and clinical evaluation. There were few later pregnancy models but, if validated, these could be most relevant to individualised discussions around timing of birth. TWEETABLE ABSTRACT: Prediction models using maternal factors, blood tests and ultrasound could individualise stillbirth prevention, but existing models are at high risk of bias.

Prediction of stillbirth: an umbrella review of evaluation of prognostic variables.

BACKGROUND: Stillbirth accounts for over 2 million deaths a year worldwide and rates remains stubbornly high. Multivariable prediction models may be key to individualised monitoring, intervention or early birth in pregnancy to prevent stillbirth. OBJECTIVES: To collate and evaluate systematic reviews of factors associated with stillbirth in order to identify variables relevant to prediction model development. SEARCH STRATEGY: MEDLINE, Embase, DARE and Cochrane Library databases and reference lists were searched up to November 2019. SELECTION CRITERIA: We included systematic reviews of association of individual variables with stillbirth without language restriction. DATA COLLECTION AND ANALYSIS: Abstract screening and data extraction were conducted in duplicate. Methodological quality was assessed using AMSTAR and QUIPS criteria. The evidence supporting association with each variable was graded. RESULTS: The search identified 1198 citations. Sixty-nine systematic reviews reporting 64 variables were included. The most frequently reported were maternal age (n = 5), body mass index (n = 6) and maternal diabetes (n = 5). Uterine artery Doppler appeared to have the best performance of any single test for stillbirth. The strongest evidence of association was for nulliparity and pre-existing hypertension. CONCLUSION: We have identified variables relevant to the development of prediction models for stillbirth. Age, parity and prior adverse pregnancy outcomes had a more convincing association than the best performing tests, which were PAPP-A, PlGF and UtAD. The evidence was limited by high heterogeneity and lack of data on intervention bias. TWEETABLE ABSTRACT: Review shows key predictors for use in developing models predicting stillbirth include age, prior pregnancy outcome and PAPP-A, PLGF and Uterine artery Doppler.

Regulated Phosphosignaling Associated with Breast Cancer Subtypes and Druggability.

Aberrant phospho-signaling is a hallmark of cancer. We investigated kinase-substrate regulation of 33,239 phosphorylation sites (phosphosites) in 77 breast tumors and 24 breast cancer xenografts. Our search discovered 2134 quantitatively correlated kinase-phosphosite pairs, enriching for and extending experimental or binding-motif predictions. Among the 91 kinases with auto-phosphorylation, elevated EGFR, ERBB2, PRKG1, and WNK1 phosphosignaling were enriched in basal, HER2-E, Luminal A, and Luminal B breast cancers, respectively, revealing subtype-specific regulation. CDKs, MAPKs, and ataxia-telangiectasia proteins were dominant, master regulators of substrate-phosphorylation, whose activities are not captured by genomic evidence. We unveiled phospho-signaling and druggable targets from 113 kinase-substrate pairs and cascades downstream of kinases, including AKT1, BRAF and EGFR. We further identified kinase-substrate-pairs associated with clinical or immune signatures and experimentally validated activated phosphosites of ERBB2, EIF4EBP1, and EGFR. Overall, kinase-substrate regulation revealed by the largest unbiased global phosphorylation data to date connects driver events to their signaling effects.

Core outcome set for studies investigating management of selective fetal growth restriction in twins.

OBJECTIVE: Selective fetal growth restriction (sFGR) occurs in monochorionic twin pregnancies when unequal placental sharing leads to restriction in the growth of just one twin. Management options include laser separation of the fetal circulations, selective reduction or expectant management, but what constitutes the best treatment is not yet known. New trials in this area are urgently needed but, in this rare and complex group, maximizing the relevance and utility of clinical research design and outputs is paramount. A core outcome set ensures standardized outcome collection and reporting in future research. The objective of this study was to develop a core outcome set for studies evaluating treatments for sFGR in monochorionic twins. METHODS: An international steering group of clinicians, researchers and patients with experience of sFGR was established to oversee the process of development of a core outcome set for studies investigating the management of sFGR. Outcomes reported in the literature were identified through a systematic review and informed the design of a three-round Delphi survey. Clinicians, researchers, and patients and family representatives participated in the survey. Outcomes were scored on a Likert scale from 1 (limited importance for making a decision) to 9 (critical for making a decision). Consensus was defined a priori as a Likert score of ≥ 8 in the third round of the Delphi survey. Participants were then invited to take part in an international meeting of stakeholders in which the modified nominal group technique was used to consider the consensus outcomes and agree on a final core outcome set. RESULTS: Ninety-six outcomes were identified from 39 studies in the systematic review. One hundred and three participants from 23 countries completed the first round of the Delphi survey, of whom 88 completed all three rounds. Twenty-nine outcomes met the a priori criteria for consensus and, along with six additional outcomes, were prioritized in a consensus development meeting, using the modified nominal group technique. Twenty-five stakeholders participated in this meeting, including researchers (n = 3), fetal medicine specialists (n = 3), obstetricians (n = 2), neonatologists (n = 3), midwives (n = 4), parents and family members (n = 6), patient group representatives (n = 3), and a sonographer. Eleven core outcomes were agreed upon. These were live birth, gestational age at birth, birth weight, intertwin birth-weight discordance, death of surviving twin after death of cotwin, loss during pregnancy or before final hospital discharge, parental stress, procedure-related adverse maternal outcome, length of neonatal stay in hospital, neurological abnormality on postnatal imaging and childhood disability. CONCLUSIONS: This core outcome set for studies investigating the management of sFGR represents the consensus of a large and diverse group of international collaborators. Use of these outcomes in future trials should help to increase the clinical relevance of research on this condition. Consensus agreement on core outcome definitions and measures is now required. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.

Proteogenomic characterization of human colon and rectal cancer.

Extensive genomic characterization of human cancers presents the problem of inference from genomic abnormalities to cancer phenotypes. To address this problem, we analysed proteomes of colon and rectal tumours characterized previously by The Cancer Genome Atlas (TCGA) and perform integrated proteogenomic analyses. Somatic variants displayed reduced protein abundance compared to germline variants. Messenger RNA transcript abundance did not reliably predict protein abundance differences between tumours. Proteomics identified five proteomic subtypes in the TCGA cohort, two of which overlapped with the TCGA 'microsatellite instability/CpG island methylation phenotype' transcriptomic subtype, but had distinct mutation, methylation and protein expression patterns associated with different clinical outcomes. Although copy number alterations showed strong cis- and trans-effects on mRNA abundance, relatively few of these extend to the protein level. Thus, proteomics data enabled prioritization of candidate driver genes. The chromosome 20q amplicon was associated with the largest global changes at both mRNA and protein levels; proteomics data highlighted potential 20q candidates, including HNF4A (hepatocyte nuclear factor 4, alpha), TOMM34 (translocase of outer mitochondrial membrane 34) and SRC (SRC proto-oncogene, non-receptor tyrosine kinase). Integrated proteogenomic analysis provides functional context to interpret genomic abnormalities and affords a new paradigm for understanding cancer biology.

Unlike calorie restriction, Roux-en-Y gastric bypass surgery does not increase hypothalamic AgRP and NPY in mice on a high-fat diet.

OBJECTIVES: Dieting often fails because weight loss triggers strong counter-regulatory biological responses such as increased hunger and hypometabolism that are thought to be critically dependent on the master fuel sensor in the mediobasal hypothalamus (MBH). Because prolonged starvation has been shown to increase AgRP and NPY, the expression level of these two orexigenic genes has been taken as an experimental readout for the presence or absence of hunger. Roux-en-Y gastric bypass (RYGB) surgery leads to a significant weight loss without inducing the associated hunger, indicating possible changes in hypothalamic neuropeptides and/or signaling. Our goal was to assess key genes in the MBH involved in regulating body weight, appetite, and inflammation/oxidative stress after RYGB surgery in mice. METHODS: Obese mice on a high-fat diet were subjected to either sham or RYGB surgery, or caloric restriction to match the weight of RYGB group. Chow-fed mice without surgery served as an additional control group. After 2 or 12 weeks post-surgery, hypothalamic genes were analyzed by real-time qPCR. RESULTS: During the rapid weight loss phase at 2 weeks after RYGB surgery, hypothalamic AgRP and NPY gene expression was not increased compared to mice with sham surgery, indicating that the mice are not hungry. In contrast, the same weight loss induced by caloric restriction promptly triggered increased AgRP and NPY expression. This differential effect of RYGB and caloric restriction was no longer observed during the weight-maintenance phase at 12 weeks after surgery. A similar differential effect was observed for ObRb, but not for POMC and CART expression. Furthermore, RAGE and IBA-1, two markers for inflammation/oxidative stress, were significantly suppressed after RYGB compared to caloric restriction at 2 weeks post-surgery. CONCLUSIONS: These findings suggest that RYGB prevents the biologically adaptive hunger response triggered by undernutrition and weight loss, and suppresses weight loss-induced hypothalamic inflammation markers.

Proteomic analysis of synovial fluid identifies periostin as a biomarker for anterior cruciate ligament injury.

OBJECTIVE: Emerging evidence suggests that injury to the anterior cruciate ligament (ACL) typically initiates biological changes that contribute to the development of osteoarthritis (OA). The molecular biomarkers or mediators of these biological events remain unknown. The goal of this exploratory study was to identify novel synovial fluid biomarkers associated with early biological changes following ACL injury distinct from findings in end-stage OA. METHODS: Synovial fluid was aspirated from patients with acute (≤30 days) and subacute (31-90 days) ACL tears and from patients with advanced OA and probed via tandem mass spectrometry for biomarkers to distinguish OA from ACL injury. Periostin (POSTN) was identified as a potential candidate. Further analyses of POSTN were performed in synovial fluid, OA cartilage, torn ACL remnants, and cultured cells and media by Western blot, PCR, immunostaining and ELISA. RESULTS: Synovial fluid analysis revealed that POSTN exhibited higher expression in subacute ACL injury than OA. POSTN expression was relatively low in cartilage/chondrocytes suggesting it is also produced by other intra-articular tissues. Conversely, high and time-dependent expression of POSTN in ACL tear remnants and isolated cells was consistent with the synovial fluid results. CONCLUSIONS: Elevated POSTN may provide a synovial fluid biomarker of subacute ACL injury setting separate from OA. Increased expression of POSTN in ACL suggests that the injured ACL may play a pivotal role in POSTN production, which is sensitive to time from injury. Previous studies have shown potential catabolic effects of POSTN, raising the possibility that POSTN contributes to the initiation of joint degeneration and may offer a window of opportunity to intervene in the early stages of post-traumatic OA.

Perinatal outcome of monochorionic twin pregnancy complicated by selective fetal growth restriction according to management: systematic review and meta-analysis.

OBJECTIVE: To explore the impact of severity and management (expectant, laser treatment or selective reduction) on perinatal outcome of monochorionic twin pregnancies complicated by selective fetal growth restriction (sFGR). METHODS: MEDLINE, EMBASE, CINAHL, ClinicalTrials.gov and The Cochrane Library databases were searched for studies on outcome following expectant management, laser treatment or selective reduction in monochorionic twin pregnancies complicated by sFGR. Only pregnancies affected by sFGR and categorized according to the Gratacós classification (Type I, II or III) were included. The primary outcome was mortality, including single and double intrauterine (IUD), neonatal (NND) and perinatal deaths. Secondary outcomes were neonatal morbidity, abnormal postnatal brain imaging, intraventricular hemorrhage, periventricular leukomalacia, respiratory distress syndrome, admission to neonatal intensive care unit and survival free from neurological complications (intact survival). Meta-analyses of proportions were used to analyze the extracted data according to management, severity of sFGR and fetal size (smaller vs larger twin). RESULTS: Sixteen observational studies (786 monochorionic twin pregnancies) were included. In pregnancies complicated by Type-I sFGR managed expectantly, IUD occurred in 3.1% (95% CI, 1.1-5.9%) of fetuses and 97.9% (95% CI, 93.6-99.9%) of twins had intact survival. In pregnancies complicated by Type-I sFGR treated using laser therapy, IUD occurred in 16.7% (95% CI, 0.4-64.1%) of fetuses and, in those treated using selective reduction, IUD occurred in 0% (95% CI, 0-34.9%) of cotwins, with no evidence of neurological complications in the survivors. In pregnancies complicated by Type-II sFGR managed expectantly, IUD occurred in 16.6% (95% CI, 6.9-29.5%) and NND in 6.4% (95% CI, 0.2-28.2%) of fetuses, and 89.3% (95% CI, 71.8-97.7%) of twins survived without neurological compromise. In Type-II sFGR pregnancies treated using laser therapy, IUD occurred in 44.3% (95% CI, 22.2-67.7%) of fetuses, while none of the affected cases experienced morbidity and survivors were free of neurological complications. Of pregnancies undergoing selective reduction, IUD of the cotwin occurred in 5.0% (95% CI, 0.03-20.5%) and NND in 3.7% (95% CI, 0.2-11.1%), and 90.6% (95% CI, 42.3-94.3%) of surviving cotwins were free from neurological complications. In pregnancies complicated by Type-III sFGR managed expectantly, IUD occurred in 13.2% (95% CI, 7.2-20.5%) and NND in 6.8% (95% CI, 0.7-18.6%) of fetuses, and 61.9% (95% CI, 38.4-81.9%) of twins had intact survival. In pregnancies complicated by Type-III sFGR treated with laser therapy, IUD occurred in 32.9% (95% CI, 20.9-46.2%) of fetuses and all surviving twins were without neurological complications. Finally, in pregnancies with Type-III sFGR treated with selective reduction, NND occurred in 5.2% (95% CI, 0.8-12.8%) of cotwins and 98.8% (95% CI, 93.9-99.9%) had intact survival. CONCLUSION: Type-I sFGR is characterized by good perinatal outcome when managed expectantly, which represents the most reasonable management strategy for the large majority of affected cases. Pregnancies complicated by Type-II or -III sFGR treated with fetoscopic laser ablation have a higher rate of mortality but lower rate of morbidity compared with those managed expectantly, supporting the use of fetal therapy at gestations remote from neonatal viability. Data on outcome following selective reduction are scarce. In view of the lack of evidence from randomized controlled trials, prenatal management of sFGR should be individualized according to gestational age at diagnosis, severity of growth discordance and magnitude of Doppler anomalies. Copyright © 2018 ISUOG. Published by John Wiley & Sons Ltd.

Variation in outcome reporting in randomized controlled trials of interventions for prevention and treatment of fetal growth restriction.

OBJECTIVE: Although fetal growth restriction (FGR) is well known to be associated with adverse outcomes for the mother and offspring, effective interventions for the management of FGR are yet to be established. Trials reporting interventions for the prevention and treatment of FGR may be limited by heterogeneity in the underlying pathophysiology. The aim of this study was to conduct a systematic review of outcomes reported in randomized controlled trials (RCTs) assessing interventions for the prevention or treatment of FGR, in order to identify and categorize the variation in outcome reporting. METHODS: MEDLINE, EMBASE and The Cochrane Library were searched from inception until August 2018 for RCTs investigating therapies for the prevention and treatment of FGR. Studies were assessed systematically and data on outcomes that were reported in the included studies were extracted and categorized. The methodological quality of the included studies was assessed using the Jadad score. RESULTS: The search identified 2609 citations, of which 153 were selected for full-text review and 72 studies (68 trials) were included in the final analysis. There were 44 trials relating to the prevention of FGR and 24 trials investigating interventions for the treatment of FGR. The mean Jadad score of all studies was 3.07, and only nine of them received a score of 5. We identified 238 outcomes across the included studies. The most commonly reported were birth weight (88.2%), gestational age at birth (72.1%) and small-for-gestational age (67.6%). Few studies reported on any measure of neonatal morbidity (27.9%), while adverse effects of the interventions were reported in only 17.6% of trials. CONCLUSIONS: There is significant variation in outcome reporting across RCTs of therapies for the prevention and treatment of FGR. The clinical applicability of future research would be enhanced by the development of a core outcome set for use in future trials. Copyright © 2018 ISUOG. Published by John Wiley & Sons Ltd.

Prediction of pre-eclampsia: review of reviews.

OBJECTIVE: Primary studies and systematic reviews provide estimates of varying accuracy for different factors in the prediction of pre-eclampsia. The aim of this study was to review published systematic reviews to collate evidence on the ability of available tests to predict pre-eclampsia, to identify high-value avenues for future research and to minimize future research waste in this field. METHODS: MEDLINE, EMBASE and The Cochrane Library including DARE (Database of Abstracts of Reviews of Effects) databases, from database inception to March 2017, and bibliographies of relevant articles were searched, without language restrictions, for systematic reviews and meta-analyses on the prediction of pre-eclampsia. The quality of the included reviews was assessed using the AMSTAR tool and a modified version of the QUIPS tool. We evaluated the comprehensiveness of search, sample size, tests and outcomes evaluated, data synthesis methods, predictive ability estimates, risk of bias related to the population studied, measurement of predictors and outcomes, study attrition and adjustment for confounding. RESULTS: From 2444 citations identified, 126 reviews were included, reporting on over 90 predictors and 52 prediction models for pre-eclampsia. Around a third (n = 37 (29.4%)) of all reviews investigated solely biochemical markers for predicting pre-eclampsia, 31 (24.6%) investigated genetic associations with pre-eclampsia, 46 (36.5%) reported on clinical characteristics, four (3.2%) evaluated only ultrasound markers and six (4.8%) studied a combination of tests; two (1.6%) additional reviews evaluated primary studies investigating any screening test for pre-eclampsia. Reviews included between two and 265 primary studies, including up to 25 356 688 women in the largest review. Only approximately half (n = 67 (53.2%)) of the reviews assessed the quality of the included studies. There was a high risk of bias in many of the included reviews, particularly in relation to population representativeness and study attrition. Over 80% (n = 106 (84.1%)) summarized the findings using meta-analysis. Thirty-two (25.4%) studies lacked a formal statement on funding. The predictors with the best test performance were body mass index (BMI) > 35 kg/m2 , with a specificity of 92% (95% CI, 89-95%) and a sensitivity of 21% (95% CI, 12-31%); BMI > 25 kg/m2 , with a specificity of 73% (95% CI, 64-83%) and a sensitivity of 47% (95% CI, 33-61%); first-trimester uterine artery pulsatility index or resistance index > 90th centile (specificity 93% (95% CI, 90-96%) and sensitivity 26% (95% CI, 23-31%)); placental growth factor (specificity 89% (95% CI, 89-89%) and sensitivity 65% (95% CI, 63-67%)); and placental protein 13 (specificity 88% (95% CI, 87-89%) and sensitivity 37% (95% CI, 33-41%)). No single marker had a test performance suitable for routine clinical use. Models combining markers showed promise, but none had undergone external validation. CONCLUSIONS: This review of reviews calls into question the need for further aggregate meta-analysis in this area given the large number of published reviews subject to the common limitations of primary predictive studies. Prospective, well-designed studies of predictive markers, preferably randomized intervention studies, and combined through individual-patient data meta-analysis are needed to develop and validate new prediction models to facilitate the prediction of pre-eclampsia and minimize further research waste in this field. Copyright © 2018 ISUOG. Published by John Wiley & Sons Ltd.

Predicción de la preeclampsia: revisión de revisiones OBJETIVO: Los estudios primarios y las revisiones sistemáticas proporcionan estimaciones de precisión variable para diferentes factores en la predicción de la preeclampsia. El objetivo de este estudio fue revisar las revisiones sistemáticas publicadas para recopilar evidencia sobre la capacidad de las pruebas disponibles para predecir la preeclampsia, identificar avenidas de investigación futura valiosas y minimizar el desperdicio futuro de investigación en este campo. MÉTODOS: Se realizaron búsquedas de artículos relevantes en bibliografías sobre el tema y en las bases de datos MEDLINE, EMBASE y The Cochrane Library, incluida DARE (Database of Abstracts of Reviews of Effects), desde el inicio de cada base de datos hasta marzo de 2017, sin restricciones de idioma, para obtener revisiones sistemáticas y metaanálisis sobre la predicción de la preeclampsia. La calidad de las revisiones incluidas se evaluó utilizando la herramienta AMSTAR y una versión modificada de la herramienta QUIPS. Se evaluó la amplitud de la búsqueda, el tamaño de la muestra, las pruebas y los resultados evaluados, los métodos de síntesis de datos, las estimaciones de la capacidad de predicción, el riesgo de sesgo relacionado con la población estudiada, la medición de los predictores y los resultados, la deserción del estudio y el ajuste por confusión. RESULTADOS: De las 2444 citas identificadas, se incluyeron 126 revisiones, que informaron sobre más de 90 predictores y 52 modelos de predicción para la preeclampsia. Alrededor de un tercio (n=37 (29,4%)) de todas las revisiones investigaron únicamente marcadores bioquímicos para predecir la preeclampsia, 31 (24,6%) investigaron asociaciones genéticas con la preeclampsia, 46 (36,5%) informaron sobre las características clínicas, cuatro (3,2%) evaluaron sólo marcadores ecográficos y seis (4,8%) estudiaron una combinación de pruebas; dos (1,6%) revisiones adicionales evaluaron los estudios primarios que investigaron cualquier prueba de diagnóstico de la preeclampsia. Las revisiones incluyeron entre dos y 265 estudios primarios, que incluyeron hasta 25 356 688 mujeres en la revisión más grande. Sólo aproximadamente la mitad (n=67 (53,2%)) de las revisiones evaluaron la calidad de los estudios incluidos. En muchas de las revisiones incluidas hubo un alto riesgo de sesgo, particularmente en relación con la representatividad de la población y la deserción de los estudios. Más del 80% (n=106 (84,1%)) resumió los hallazgos utilizando el metaanálisis. Treinta y dos (25,4%) estudios carecían de una declaración formal sobre la financiación. Los predictores con el mejor rendimiento de la prueba fueron el índice de masa corporal (IMC) >35 kg.m-2 , con una especificidad del 92% (IC 95%, 89-95%) y una sensibilidad del 21% (IC 95%, 12-31%); IMC >25 kg.m-2 , con una especificidad del 73% (IC 95%: 64-83%) y una sensibilidad del 47% (IC 95%: 33-61%); índice de pulsatilidad de la arteria uterina en el primer trimestre o índice de resistencia >90° percentil (especificidad del 93% (IC 95%: 90-96%) y sensibilidad del 26% (IC 95%: 23-31%)); factor de crecimiento placentario (especificidad 89% (IC 95%, 89-89%) y sensibilidad 65% (IC 95%, 63-67%)); y proteína placentaria 13 (especificidad 88% (IC 95%, 87-89%) y sensibilidad 37% (IC 95%, 33-41%)). Ningún marcador por sí solo tuvo un rendimiento de la prueba adecuado para el uso clínico rutinario. Los modelos que combinan marcadores son prometedores, pero ninguno fue sometido a una validación externa. CONCLUSIONES: Esta revisión de revisiones ha puesto en duda la necesidad de un metaanálisis agregado adicional en esta área, dado el gran número de revisiones publicadas sujetas a las limitaciones comunes de los estudios predictivos primarios. Se necesitan estudios prospectivos bien diseñados de marcadores predictivos, preferiblemente en estudios de intervención aleatorios, y combinados mediante el metaanálisis de datos de pacientes individuales, para desarrollar y validar nuevos modelos predictivos que faciliten la predicción de la preeclampsia y minimicen el desperdicio de investigación adicional en este campo.

Proteome Profiling Outperforms Transcriptome Profiling for Coexpression Based Gene Function Prediction.

Coexpression of mRNAs under multiple conditions is commonly used to infer cofunctionality of their gene products despite well-known limitations of this "guilt-by-association" (GBA) approach. Recent advancements in mass spectrometry-based proteomic technologies have enabled global expression profiling at the protein level; however, whether proteome profiling data can outperform transcriptome profiling data for coexpression based gene function prediction has not been systematically investigated. Here, we address this question by constructing and analyzing mRNA and protein coexpression networks for three cancer types with matched mRNA and protein profiling data from The Cancer Genome Atlas (TCGA) and the Clinical Proteomic Tumor Analysis Consortium (CPTAC). Our analyses revealed a marked difference in wiring between the mRNA and protein coexpression networks. Whereas protein coexpression was driven primarily by functional similarity between coexpressed genes, mRNA coexpression was driven by both cofunction and chromosomal colocalization of the genes. Functionally coherent mRNA modules were more likely to have their edges preserved in corresponding protein networks than functionally incoherent mRNA modules. Proteomic data strengthened the link between gene expression and function for at least 75% of Gene Ontology (GO) biological processes and 90% of KEGG pathways. A web application Gene2Net (http://cptac.gene2net.org) developed based on the three protein coexpression networks revealed novel gene-function relationships, such as linking ERBB2 (HER2) to lipid biosynthetic process in breast cancer, identifying PLG as a new gene involved in complement activation, and identifying AEBP1 as a new epithelial-mesenchymal transition (EMT) marker. Our results demonstrate that proteome profiling outperforms transcriptome profiling for coexpression based gene function prediction. Proteomics should be integrated if not preferred in gene function and human disease studies.

Human Experimental Challenge With Enterotoxigenic Escherichia coli Elicits Immune Responses to Canonical and Novel Antigens Relevant to Vaccine Development.

Background: Enterotoxigenic Escherichia coli (ETEC) is a major cause of diarrheal illness in the developing world. Enterotoxigenic E coli vaccinology has been challenged by genetic diversity and heterogeneity of canonical antigens. Examination of the antigenic breadth of immune responses associated with protective immunity could afford new avenues for vaccine development. Methods: Antibody lymphocyte supernatants (ALS) and sera from 20 naive human volunteers challenged with ETEC strain H10407 and from 10 volunteers rechallenged 4-6 weeks later with the same strain (9 of whom were completely protected on rechallenge) were tested against ETEC proteome microarrays containing 957 antigens. Results: Enterotoxigenic E coli challenge stimulated robust serum and mucosal (ALS) responses to canonical vaccine antigens (CFA/I, and the B subunit of LT) as well as a small number of antigens not presently targeted in ETEC vaccines. These included pathovar-specific secreted proteins (EtpA, EatA) as well as highly conserved E coli antigens including YghJ, flagellin, and pertactin-like autotransporter proteins, all of which have previously afforded protection against ETEC infection in preclinical studies. Conclusions: Taken together, studies reported here suggest that immune responses after ETEC infection involve traditional vaccine targets as well as a select number of more recently identified protein antigens that could offer additional avenues for vaccine development for these pathogens.