Risk of Neonatal Hypoglycemia in Infants of Mothers With Gestational Glucose Intolerance.

OBJECTIVE: To examine the relationship between gestational glucose intolerance (GGI) and neonatal hypoglycemia. RESEARCH DESIGN AND METHODS: This was a secondary analysis of 8,262 mother-infant dyads, with delivery at two hospitals between 2014 and 2023. We categorized maternal glycemic status as normal glucose tolerance (NGT), GGI, or gestational diabetes mellitus (GDM). We defined NGT according to a normal glucose load test result, GGI according to an abnormal glucose load test result with zero (GGI-0) or one (GGI-1) abnormal value on the 100-g oral glucose tolerance test, and GDM according to an abnormal glucose load test result with two or more abnormal values on the glucose tolerance test. Neonatal hypoglycemia was defined according to blood glucose <45 mg/dL or ICD-9 or ICD-10 diagnosis of neonatal hypoglycemia. We used logistic regression analysis to determine associations between maternal glucose tolerance category and neonatal hypoglycemia and conducted a sensitivity analysis using Δ-adjusted multiple imputation, assuming for unscreened infants a rate of neonatal hypoglycemia as high as 33%. RESULTS: Of infants, 12% had neonatal hypoglycemia. In adjusted models, infants born to mothers with GGI-0 had 1.28 (95% 1.12, 1.65), GGI-1 1.58 (95% CI 1.11, 2.25), and GDM 4.90 (95% CI 3.81, 6.29) times higher odds of neonatal hypoglycemia in comparison with infants born to mothers with NGT. Associations in sensitivity analyses were consistent with the primary analysis. CONCLUSIONS: GGI is associated with increased risk of neonatal hypoglycemia. Future research should include examination of these associations in a cohort with more complete neonatal blood glucose ascertainment and determination of the clinical significance of these findings on long-term child health.

The utility of plasma glycated CD59 in predicting postpartum glucose intolerance: A prospective study of women diagnosed with GDM during a period of universal GDM screening.

AIMS: Gestational diabetes (GDM) is associated with the development of postpartum (PP) glucose intolerance. Plasma glycated CD59 (pGCD59) is an emerging biomarker for the detection of hyperglycaemia. The aim of this study was to assess the ability of PP pGCD59 to predict the development of PP GI as defined by the 2 h 75 g OGTT using the ADA criteria, in a cohort of women diagnosed with prior GDM in the index pregnancy using the 2 h 75 g OGTT at 24-28 weeks of gestation according to the World Health Organization (WHO) 2013 criteria. METHODS: Of the 2017 pregnant women recruited prospectively 140 women with gestational diabetes had samples for pGCD59 taken PP at the time of the OGTT. The ability of pGCD59 to predict the results of the PP OGTT was assessed using nonparametric receiver operating characteristic (ROC) curves. RESULTS: Women with PP glucose intolerance had significantly higher PP pGCD59 levels compared to women with normal glucose tolerance PP (3.8 vs. 2.7 SPU). PP pGCD59 identified women who developed glucose intolerance PP with an AUC of 0.80 (95% CI: 0.70-0.91). A PP pGCD59 cut-off value of 1.9 SPU generated a sensitivity of 100% (95% CI: 83.9-100), specificity of 16.9% (95% CI: 9.8-26.3), positive predictive value of 22.1% (95% CI: 21.0-22.6), and negative predictive value of 100% (95% CI: 87.4-100). PP fasting plasma glucose generated an AUC of 0.96 (95% CI: 0.89-0.99) for the identification of PP glucose intolerance. CONCLUSION: Our study found that PP pGCD9 may be a promising biomarker to identify women not requiring PP glucose intolerance screening using the traditional OGTT. While the diagnostic accuracy of pGCD59 is good, fasting plasma glucose remains a better test for the identification of PP glucose intolerance.

Role of symmetry in 3,3-diphenyl-1,3-dihydroindol-2-one derivatives as inhibitors of translation initiation.

The ternary complex (eIF2·GTP·Met-tRNAiMet) and the eIF4F complex assembly are two major regulatory steps in the eukaryotic translation initiation. Inhibition of the ternary complex assembly is therefore a promising target for the development of novel anti-cancer therapeutics. Building on the finding that clotrimazole (CLT), a molecular probe that depletes intracellular Ca2+ stores and subsequently induce eIF2α phosphorylation, inhibit translation initiation, and reduce preferentially the expression of oncoproteins over "housekeeping" ones,1-3 we undertook structure activity relationship (SAR) studies that identified 3,3-diarylindoline-2-one #1181 as an interesting scaffold. Compound #1181 also induce phosphorylation of eIF2α thereby reducing the availability of the ternary complex, which leads to inhibition of translation initiation.4 Our subsequent efforts focused on understanding SAR iterative lead optimization to enhance potency and improve bioavailability. Herein, we report a complementing study focusing on heavily substituted symmetric and asymmetric 3,3-(o,m-disubstituted)diarylindoline-2-ones. These compounds were evaluated by the dual luciferase reporter ternary complex assay that recapitualates phosphorylation of eIF2α in a quantitative manner. We also evaluated all compounds by sulforhodamine B assay, which measures the overall effect of compounds on cell proliferations and/or viability.

The ability of pGCD59 to predict adverse pregnancy outcomes: a prospective study of non-diabetic pregnant women in Ireland.

AIM: Even though most pregnancies are uneventful, occasionally complications do occur. Gestational diabetes is linked to an increased risk of adverse pregnancy outcomes. Early identification of women at risk of experiencing adverse outcomes, ideally through a single blood test, would facilitate early intervention. Plasma glycated CD59 (pGCD59) is an emerging biomarker which has shown promise in identifying hyperglycaemia during pregnancy and has been associated with the risk of delivering an LGA infant. The aim of this study was to explore the ability of the first- and second-trimester pGCD59 to predict adverse pregnancy outcomes. METHODS: This was a prospective study of 378 pregnant women. Samples for pGCD59 were taken at the first antenatal visit and at the time of the 2 h 75 g OGTT (24-28 weeks of gestation). Adjusted receiver operating characteristic curves were used to evaluate the ability of pGCD59 to predict maternal and neonatal outcomes. RESULTS: First-trimester pGCD59 levels were higher in women with gestational diabetes who delivered a macrosomic infant (4.2 ± 0.7 vs. 3.5 ± 1.0 SPU, p < 0.01) or an LGA infant (4.3 ± 0.3 vs. 3.6 ± 1.0 SPU, p = 0.01) compared to women with GDM that did not experience these outcomes. Second-trimester pGCD59 levels were higher in women that developed polyhydramnios (2.9 ± 0.4 vs. 2.5 ± 1.1 SPU, p = 0.03). First- and second-trimester pGCD59 predicted pregnancy-induced hypertension with good accuracy (AUC:0.85, 95%CI:0.78-0.91; AUC: 0.80, 95%CI: 0.73-0.88, respectively) and neonatal hypoglycaemia with fair to good accuracy (AUC:0.77, 95%CI: 0.54-0.99, AUC:0.81, 95%CI:0.62-0.99). CONCLUSIONS: This study has shown that pGCD59 has the potential to predict adverse pregnancy outcomes. Prospective studies with a larger number of cases are necessary to fully explore and validate the potential of this emerging biomarker in predicting adverse pregnancy outcomes.

The Role of Early Pregnancy Maternal pGCD59 Levels in Predicting Neonatal Hypoglycemia-Subanalysis of the DALI Study.

CONTEXT: Neonatal hypoglycaemia (NH) is the most common metabolic problem in infants born of mothers with gestational diabetes. Plasma glycated CD59 (pGCD59) is an emerging biomarker that has shown potential in identifying women at risk of developing gestational diabetes. The aim of this study was to assess the association between early maternal levels of pGCD59 and NH. OBJECTIVE: The aim of this study was to assess the association between early pregnancy maternal levels of plasma glycated CD59 (pGCD59) and neonatal hypoglycemia (NH). METHODS: This is an observational study of pregnant women with a prepregnancy body mass index (BMI) greater than or equal to 29 screened for eligibility to participate in the Vitamin D and Lifestyle Intervention for Gestational Diabetes (DALI) trial. This analysis included 399 pregnancies. Levels of pGCD59 were measured in fasting maternal samples taken at the time of a 75-g, 2-hour oral glucose tolerance test performed in early pregnancy (< 20 weeks). NH, the study outcome, was defined as a heel-prick capillary glucose level of less than 2.6 mmol/L within 48 hours of delivery. RESULTS: We identified 30 infants with NH. Maternal levels of pGCD59 in early pregnancy were positively associated with the prevalence of NH (one-way analysis of variance, P < .001). The odds of NH were higher in infants from mothers in tertile 3 of pGCD59 levels compared to those from mothers in tertile 1 (odds ratio [OR]: 2.41; 95% CI, 1.03-5.63). However, this was attenuated when adjusted for maternal BMI (OR: 2.28; 95% CI, 0.96-5.43). The cross-validated area under the curve (AUC) was 0.64 (95% CI, 0.54-0.74), and adjusted for maternal BMI, age, and ethnicity, the AUC was 0.70 (95% CI, 0.56-0.78). CONCLUSION: Although pGCD59 levels in early pregnancy in women with BMI greater than or equal to 29 are associated with NH, our results indicate that this biomarker by itself is only a fair predictor of NH.

The Diagnostic Accuracy of Second Trimester Plasma Glycated CD59 (pGCD59) to Identify Women with Gestational Diabetes Mellitus Based on the 75 g OGTT Using the WHO Criteria: A Prospective Study of Non-Diabetic Pregnant Women in Ireland.

The aim of this study was to evaluate the ability of second trimester plasma glycated CD59 (pGCD59), a novel biomarker, to predict the results of the 2 h 75 g oral glucose tolerance test at 24−28 weeks of gestation, employing the 2013 World Health Organisation criteria. This was a prospective study of 378 pregnant women. The ability of pGCD59 to predict gestational diabetes (GDM) was assessed using adjusted ROC curves for maternal age, BMI, maternal ethnicity, parity, previous GDM, and family history of diabetes. The pGCD59 levels were significantly higher in women with GDM compared to women with normal glucose tolerance (p = 0.003). The pGCD59 generated an adjusted AUC for identifying GDM cases of 0.65 (95%CI: 0.58−0.71, p < 0.001). The pGCD59 predicted GDM status diagnosed by a fasting glucose value of 5.1 mmol/L with an adjusted AUC of 0.74 (95%CI: 0.65−0.81, p < 0.001). Analysis of BMI subgroups determined that pGCD59 generated the highest AUC in the 35 kg/m2 ≤ BMI < 40 kg/m2 (AUC: 0.84 95%CI: 0.69−0.98) and BMI ≥ 40 kg/m2 (AUC: 0.96 95%CI: 0.86−0.99) categories. This study found that second trimester pGCD59 is a fair predictor of GDM status diagnosed by elevated fasting glucose independent of BMI and an excellent predictor of GDM in subjects with a very high BMI.

The utility of first trimester plasma glycated CD59 (pGCD59) in predicting gestational diabetes mellitus: A prospective study of non-diabetic pregnant women in Ireland.

AIMS: To evaluate the ability of first trimester plasma glycated CD59 (pGCD59) to predict gestational diabetes mellitus (GDM) at 24-28 weeks of gestation. METHODS: Prospectively, in 378 pregnant women, GDM was diagnosed using the one step 2 h 75 g oral glucose tolerance test adjudicated by the World Health Organisation (WHO) 2013 criteria. The ability of pGCD59 to predict GDM was assessed using receiver operating characteristic (ROC) curves adjusted for maternal age, body mass index (BMI), maternal ethnicity, parity, previous GDM, family history of diabetes mellitus and week of gestation at time of pGCD59 sampling. RESULTS: pGCD59 generated an adjusted area under the curve (AUC) of (a) 0.63 (95 %CI:0.56-0.70, p < 0.001) for predicting GDM, and (b) 0.71 (95 %CI:0.62-0.79, p < 0.001 for GDM diagnosed with a fasting plasma glucose (FPG) ≥ 5.1 mmol/L. Sensitivity analysis of BMI subgroups showed that pGCD59 generated the highest AUC in the 35 kg/m2 ≤ BMI < 40 kg/m2 (AUC:0.85, 95 %CI:0.70-0.98) and BMI ≥ 40 kg/m2 (AUC:0.88, 95 %CI:0.63-0.99) categories. CONCLUSIONS: Early in pregnancy, pGCD59 may be a good predictor of GDM in women with a high BMI and a fair predictor of GDM diagnosed by an elevated FPG independent of BMI.

Plasma-glycated CD59 as an early biomarker for gestational diabetes mellitus: prospective cohort study protocol.

INTRODUCTION: The significant maternal and neonatal outcomes of gestational diabetes mellitus (GDM) make it a major public health concern. Mothers with GDM are at greater risk of pregnancy complications and their offspring are at higher risk of diabetes and obesity. Currently, GDM is diagnosed with glucose load methods which are time-consuming and inconvenient to administer more than once during pregnancy; for this reason, there is a recognised need for a more accurate and simpler test for GDM. Previous studies indicate that plasma-glycated CD59 (pGCD59) is a novel biomarker for GDM. We present here the protocol of a prospective cohort study designed to (1) determine the accuracy of pGCD59 as an early, first trimester predictor of GDM and gestational impaired glucose tolerance and (2) assess the associations between pGCD59 levels and adverse maternal and neonatal outcomes. METHODS AND ANALYSIS: We will obtain discarded plasma samples from pregnant women at two time points: first prenatal visit (usually <14 weeks gestation) and gestational weeks 24-28. A study-specific medical record abstraction tool will be used to obtain relevant maternal and neonatal clinical data from the EPIC clinical database. The prevalence of GDM will be determined using standard of care glucose load test results. We will determine the sensitivity and specificity of pGCD59 to predict the diagnosis of GDM and gestational impaired glucose tolerance, as well as the associations between levels of pGCD59 and the prevalence of maternal and neonatal outcomes. ETHICS AND DISSEMINATION: This study has been approved by the Mass General Brigham Institutional Review Board (protocol 2011P002254). The results of this study will be presented at international meetings and disseminated in peer-reviewed journals.

Plasma glycated CD59 predicts postpartum glucose intolerance after gestational diabetes.

AIM: To assess whether in women with gestational diabetes mellitus (GDM), postpartum plasma glycated CD59 (pGCD59) levels predict conversion to glucose intolerance diagnosed with an oral glucose tolerance test (OGTT). METHODS: Blood levels of pGCD59 were measured in a case-control study of 105 women with GDM who underwent a 75 g OGTT 3 months postpartum. The 35 postpartum glucose intolerant cases were individually matched for age, BMI, ethnic origin, and parity with 70 women with GDM but normal postpartum OGTT (controls). The GDM cohort (105) was also matched with 105 normal glucose tolerant women during pregnancy. pGCD59 was measured by ELISA in standard peptide units (SPU). RESULTS: Mean pGCD59 postpartum was significantly higher in cases than in controls (1.5 ± 0.6 SPU vs 1.0 ± 0.6 SPU, P < 0.001). The area under the receiving operating characteristic curve (AUC) in cases vs controls was 0.72 (95% CI: 0.62-0.83) for postpartum pGCD59 and 0.50 (95% CI: 0.36-0.61) for postpartum HbA1c. A 0.5-unit increase in postpartum pGCD59 was associated with an odds ratio (OR) of 3.3 (95% CI: 1.82-6.16, P < 0.001) for glucose intolerance postpartum. A pGCD59 cut-off postpartum of 0.9 SPU had a sensitivity of 85.7% (95% CI: 69.7-95.2%), specificity of 47.8% (95% CI: 35.6-60.2%), positive predictive value of 45.4% (95% CI: 33.1-58.2%), and negative predictive value of 86.8% (95% CI: 71.9-95.6%). pGCD59 in pregnancy was a poor predictor for glucose intolerance postpartum (AUC of 0.61 (95% CI: 0.50-0.72)). CONCLUSION: pGCD59 might identify women at low risk for glucose intolerance postpartum and could help to avoid an OGTT.

Emerging Protein Biomarkers for the Diagnosis or Prediction of Gestational Diabetes-A Scoping Review.

Introduction: Gestational diabetes (GDM), defined as hyperglycemia with onset or initial recognition during pregnancy, has a rising prevalence paralleling the rise in type 2 diabetes (T2DM) and obesity. GDM is associated with short-term and long-term consequences for both mother and child. Therefore, it is crucial we efficiently identify all cases and initiate early treatment, reducing fetal exposure to hyperglycemia and reducing GDM-related adverse pregnancy outcomes. For this reason, GDM screening is recommended as part of routine pregnancy care. The current screening method, the oral glucose tolerance test (OGTT), is a lengthy, cumbersome and inconvenient test with poor reproducibility. Newer biomarkers that do not necessitate a fasting sample are needed for the prompt diagnosis of GDM. The aim of this scoping review is to highlight and describe emerging protein biomarkers that fulfill these requirements for the diagnosis of GDM. Materials and Methods: This scoping review was conducted according to preferred reporting items for systematic reviews and meta-analyses (PRISMA) guidelines for scoping reviews using Cochrane Central Register of Controlled Trials (CENTRAL), the Cumulative Index to Nursing & Allied Health Literature (CINAHL), PubMed, Embase and Web of Science with a double screening and extraction process. The search included all articles published in the literature to July 2020. Results: Of the 3519 original database citations identified, 385 were eligible for full-text review. Of these, 332 (86.2%) were included in the scoping review providing a total of 589 biomarkers studied in relation to GDM diagnosis. Given the high number of biomarkers identified, three post hoc criteria were introduced to reduce the items set for discussion: we chose only protein biomarkers with at least five citations in the articles identified by our search and published in the years 2017-2020. When applied, these criteria identified a total of 15 biomarkers, which went forward for review and discussion. Conclusions: This review details protein biomarkers that have been studied to find a suitable test for GDM diagnosis with the potential to replace the OGTT used in current GDM screening protocols. Ongoing research efforts will continue to identify more accurate and practical biomarkers to take GDM screening and diagnosis into the 21st century.

Novel Biochemical Markers of Glycemia to Predict Pregnancy Outcomes in Women With Type 1 Diabetes.

OBJECTIVE: The optimal method of monitoring glycemia in pregnant women with type 1 diabetes remains controversial. This study aimed to assess the predictive performance of HbA1c, continuous glucose monitoring (CGM) metrics, and alternative biochemical markers of glycemia to predict obstetric and neonatal outcomes. RESEARCH DESIGN AND METHODS: One hundred fifty-seven women from the Continuous Glucose Monitoring in Women With Type 1 Diabetes in Pregnancy Trial (CONCEPTT) were included in this prespecified secondary analysis. HbA1c, CGM data, and alternative biochemical markers (glycated CD59, 1,5-anhydroglucitol, fructosamine, glycated albumin) were compared at ∼12, 24, and 34 weeks' gestation using logistic regression and receiver operating characteristic (ROC) curves to predict pregnancy complications (preeclampsia, preterm delivery, large for gestational age, neonatal hypoglycemia, admission to neonatal intensive care unit). RESULTS: HbA1c, CGM metrics, and alternative laboratory markers were all significantly associated with obstetric and neonatal outcomes at 24 weeks' gestation. More outcomes were associated with CGM metrics during the first trimester and with laboratory markers (area under the ROC curve generally <0.7) during the third trimester. Time in range (TIR) (63-140 mg/dL [3.5-7.8 mmol/L]) and time above range (TAR) (>140 mg/dL [>7.8 mmol/L]) were the most consistently predictive CGM metrics. HbA1c was also a consistent predictor of suboptimal pregnancy outcomes. Some alternative laboratory markers showed promise, but overall, they had lower predictive ability than HbA1c. CONCLUSIONS: HbA1c is still an important biomarker for obstetric and neonatal outcomes in type 1 diabetes pregnancy. Alternative biochemical markers of glycemia and other CGM metrics did not substantially increase the prediction of pregnancy outcomes compared with widely available HbA1c and increasingly available CGM metrics (TIR and TAR).

Plasma Glycated CD59 Predicts Early Gestational Diabetes and Large for Gestational Age Newborns.

CONTEXT: Gestational diabetes mellitus (GDM) diagnosed in early pregnancy is a health care challenge because it increases the risk of adverse outcomes. Plasma-glycated CD59 (pGCD59) is an emerging biomarker for diabetes and GDM. The aim of this study was to assess the performance of pGCD59 as a biomarker of early GDM and its association with delivering a large for gestational age (LGA) infant. OBJECTIVES: To assess the performance of pGCD59 to identify women with GDM in early pregnancy (GDM < 20) and assess the association of pGCD59 with LGA and potentially others adverse neonatal outcomes linked to GDM. METHODS: Blood levels of pGCD59 were measured in samples from 693 obese women (body mass index > 29) undergoing a 75-g, 2-hour oral glucose tolerance test (OGTT) at <20 weeks' gestation in the Vitamin D and Lifestyle Intervention study: the main analyses included 486 subjects who had normal glucose tolerance throughout the pregnancy, 207 who met criteria for GDM at <20 weeks, and 77 diagnosed with GDM at pregnancy weeks 24 through 28. Reference tests were 75-g, 2-hour OGTT adjudicated based on International Association of Diabetes and Pregnancy Study Group criteria. The index test was a pGCD59 ELISA. RESULTS: Mean pGCD59 levels were significantly higher (P < 0.001) in women with GDM < 20 (3.9 ±â€…1.1 standard peptide units [SPU]) than in those without (2.7 ±â€…0.7 SPU). pGCD59 accurately identified GDM in early pregnancy with an area under the curve receiver operating characteristic curves of 0.86 (95% confidence interval [CI], 0.83-0.90). One-unit increase in maternal pGCD59 level was associated with 36% increased odds of delivering an LGA infant (odds ratio for LGA vs non-LGA infant: 1.4; 95% CI, 1.1-1.8; P = 0.016). CONCLUSION: Our results indicate that pGCD59 is a simple and accurate biomarker for detection of GDM in early pregnancy and risk assessment of LGA.

New activators of eIF2α Kinase Heme-Regulated Inhibitor (HRI) with improved biophysical properties.

Heme-regulated inhibitor (HRI), a eukaryotic translation initiation factor 2 alpha (eIF2α) kinase, is critically important for coupling protein synthesis to heme availability in reticulocytes and adaptation to various environmental stressors in all cells. HRI modifies the severity of several hemoglobin misfolding disorders including ß-thalassemia. Small molecule activators of HRI are essential for studying normal- and patho-biology of this kinase as well as for the treatment of various human disorders for which activation of HRI or phosphorylation of eIF2α may be beneficial. We previously reported development of 1-((1,4-trans)-4-aryloxycyclohexyl)-3-arylureas (cHAUs) as specific HRI activators and demonstrated their potential as molecular probes for studying HRI biology and as lead compounds for treatment of various human disorders. To develop more druglike cHAUs for in vivo studies and drug development and to expand the chemical space, we undertook bioassay guided structure-activity relationship studies replacing cyclohexyl ring with various 4-6-membered rings and explored further substitutions on the N-phenyl ring. We tested all analogs in the surrogate eIF2α phosphorylation and cell proliferation assays, and a subset of analogs in secondary mechanistic assays that included endogenous eIF2α phosphorylation and expression of C/EBP homologous protein (CHOP), a downstream effector. Finally, we determined specificity of these compounds for HRI by testing their anti-proliferative activity in cells transfected with siRNA targeting HRI or mock. These compounds have significantly improved cLogPs with no loss of potencies, making them excellent candidates for lead optimization for development of investigational new drugs that potently and specifically activate HRI.

A distinctive histidine residue is essential for in vivo glycation-inactivation of human CD59 transgenically expressed in mice erythrocytes: Implications for human diabetes complications.

Clinical and experimental evidences support a link between the complement system and the pathogenesis of diabetes complications. CD59, an extracellular cell membrane-anchored protein, inhibits formation of the membrane attack complex (MAC), the main effector of complement-mediated tissue damage. This complement regulatory activity of human CD59 (hCD59) is inhibited by hyperglycemia-induced ɛ-amino glycation of Lys41 . Biochemical and structural analyses of glycated proteins with known three-dimensional structure revealed that glycation of ɛ-amino lysyl residues occurs predominantly at "glycation motives" that include lysyl/lysyl pairs or proximity of a histidyl residue, in which the imidazolyl moiety is ≈ 5Å from the ɛ-amino group. hCD59 contains a distinctive Lys41 /His44 putative glycation motif within its active site. In a model of transgenic diabetic mice expressing in erythrocytes either the wild type or a H44Q mutant form of hCD59, we demonstrate in vivo that the His44 is required for Lys41 glycation and consequent functional inactivation of hCD59, as evidenced using a mouse erythrocytes hemolytic assay. Since (1) the His44 residue is not present in CD59 from other animal species and (2) humans are particularly prone to develop complications of diabetes, our results indicate that the Lys41 /His44 glycation motif in human CD59 may confer humans a higher risk of developing vascular disease in response to hyperglycemia.

Development of 1-((1,4-trans)-4-Aryloxycyclohexyl)-3-arylurea Activators of Heme-Regulated Inhibitor as Selective Activators of the Eukaryotic Initiation Factor 2 Alpha (eIF2α) Phosphorylation Arm of the Integrated Endoplasmic Reticulum Stress Response.

Heme-regulated inhibitor (HRI), an eukaryotic translation initiation factor 2 alpha (eIF2α) kinase, plays critical roles in cell proliferation, differentiation, adaptation to stress, and hemoglobin disorders. HRI phosphorylates eIF2α, which couples cellular signals, including endoplasmic reticulum (ER) stress, to translation. We previously identified 1,3-diarylureas and 1-((1,4-trans)-4-aryloxycyclohexyl)-3-arylureas (cHAUs) as specific activators of HRI that trigger the eIF2α phosphorylation arm of ER stress response as molecular probes for studying HRI biology and its potential as a druggable target. To develop drug-like cHAUs needed for in vivo studies, we undertook bioassay-guided structure-activity relationship studies and tested them in the surrogate eIF2α phosphorylation and cell proliferation assays. We further evaluated some of these cHAUs in endogenous eIF2α phosphorylation and in the expression of the transcription factor C/EBP homologous protein (CHOP) and its mRNA, demonstrating significantly improved solubility and/or potencies. These cHAUs are excellent candidates for lead optimization for development of investigational new drugs that potently and specifically activate HRI.

Plasma Glycated CD59, a Novel Biomarker for Detection of Pregnancy-Induced Glucose Intolerance.

OBJECTIVE: Plasma glycated CD59 (pGCD59) is an emerging biomarker in diabetes. We assessed whether pGCD59 could predict the following: the results of the glucose challenge test (GCT) for screening of gestational diabetes mellitus (GDM) (primary analysis); and the diagnosis of GDM and prevalence of large for gestational age (LGA) newborns (secondary analyses). RESEARCH DESIGN AND METHODS: Case-control study of 1,000 plasma samples from women receiving standard prenatal care, 500 women having a normal GCT (control subjects) and 500 women with a failed GCT and a subsequent oral glucose tolerance test (case patients). RESULTS: Compared with control subjects, the median (interquartile range) pGCD59 value was 8.5-fold higher in case patients and 10-fold higher in GDM patients, as follows: control subjects 0.33 (0.19); case patients 2.79 (1.4); GDM patients 3.23 (1.43) (P < 0.001); area under the receiver operating characteristic curve 0.92. LGA prevalence was 4.3% in the lowest quartile and 13.5% in the highest quartile of pGCD59. CONCLUSIONS: One pGCD59 measurement during weeks 24-28 identifies pregnancy-induced glucose intolerance with high sensitivity and specificity and can potentially identify the risk for LGA.

Deficiency of the complement regulatory protein CD59 accelerates the development of diabetes-induced atherosclerosis in mice.

AIMS: Clinical and experimental evidence supports a strong link between the complement system, complement regulatory proteins and the pathogenesis of diabetes vascular complications. We previously reported that the complement regulatory protein CD59 is inactivated by glycation in humans with diabetes. Our objective for this study is to assess experimentally how the deficiency of CD59 impacts the development of diabetic atherosclerosis in vivo. METHODS: We crossed mCD59 sufficient and deficient mice into the ApoE-/- background to generate mCd59ab+/+/ApoE-/- and mCd59ab-/-/ApoE-/- mice, and induced diabetes by multiple low dose injections of streptozotocin. Atherosclerosis was detected by hematoxylin and eosin (H&E) and oil red-O staining. Membrane attack complex (MAC) deposition and macrophage infiltration were detected by immunostaining. RESULTS: Diabetic mCD59 deficient (mCD59ab-/-/ApoE-/-) mice developed nearly 100% larger atherosclerotic lesion areas in the aorta (7.5%±0.6 vs 3.6%±0.7; p<0.005) and in the aortic roots (H&E: 26.2%±1.9 vs. 14.3%±1.1; p<0.005), in both cases associated with increased lipid (Oil red-O: 14.9%±1.1 vs. 7.8%±1.1; p<0.05) and MAC deposition (6.8%±0.8 vs. 3.0%±0.7; p<0.005) and macrophage infiltration (31.5%±3.7 vs. 16.4%±3.0; p<0.05) in the aortic roots as compared to their diabetic mCD59 sufficient (mCD59ab+/+/ApoE-/-) counterpart. CONCLUSIONS: The deficiency of CD59 accelerates the development of diabetic atherosclerosis.

Role of complement and complement regulatory proteins in the complications of diabetes.

It is well established that the organ damage that complicates human diabetes is caused by prolonged hyperglycemia, but the cellular and molecular mechanisms by which high levels of glucose cause tissue damage in humans are still not fully understood. The prevalent hypothesis explaining the mechanisms that may underlie the pathogenesis of diabetes complications includes overproduction of reactive oxygen species, increased flux through the polyol pathway, overactivity of the hexosamine pathway causing intracellular formation of advanced glycation end products, and activation of protein kinase C isoforms. In addition, experimental and clinical evidence reported in past decades supports a strong link between the complement system, complement regulatory proteins, and the pathogenesis of diabetes complications. In this article, we summarize the body of evidence that supports a role for the complement system and complement regulatory proteins in the pathogenesis of diabetic vascular complications, with specific emphasis on the role of the membrane attack complex (MAC) and of CD59, an extracellular cell membrane-anchored inhibitor of MAC formation that is inactivated by nonenzymatic glycation. We discuss a pathogenic model of human diabetic complications in which a combination of CD59 inactivation by glycation and hyperglycemia-induced complement activation increases MAC deposition, activates pathways of intracellular signaling, and induces the release of proinflammatory, prothrombotic cytokines and growth factors. Combined, complement-dependent and complement-independent mechanisms induced by high glucose promote inflammation, proliferation, and thrombosis as characteristically seen in the target organs of diabetes complications.

Depletion of eIF2·GTP·Met-tRNAi translation initiation complex up-regulates BRCA1 expression in vitro and in vivo.

Most sporadic breast and ovarian cancers express low levels of the breast cancer susceptibility gene, BRCA1. The BRCA1 gene produces two transcripts, mRNAa and mRNAb. mRNAb, present in breast cancer but not in normal mammary epithelial cells, contains three upstream open reading frames (uORFs) in its 5'UTR and is translationally repressed. Comparable tandem uORFs are characteristically seen in mRNAs whose translational efficiency paradoxically increases when the overall translation rate is decreased due to phosphorylation of eukaryotic translation initiation factor 2 α (eIF2α). Here we show fish oil derived eicosopanthenoic acid (EPA) that induces eIF2α phosphorylation translationally up-regulates the expression of BRCA1 in human breast cancer cells. We demonstrate further that a diet rich in EPA strongly induces expression of BRCA1 in human breast cancer xenografts.

Structure of the eukaryotic translation initiation factor eIF4E in complex with 4EGI-1 reveals an allosteric mechanism for dissociating eIF4G.

The interaction of the eukaryotic translation initiation factor eIF4E with the initiation factor eIF4G recruits the 40S ribosomal particle to the 5' end of mRNAs, facilitates scanning to the AUG start codon, and is crucial for eukaryotic translation of nearly all genes. Efficient recruitment of the 40S particle is particularly important for translation of mRNAs encoding oncoproteins and growth-promoting factors, which often harbor complex 5' UTRs and require efficient initiation. Thus, inhibiting the eIF4E/eIF4G interaction has emerged as a previously unpursued route for developing anticancer agents. Indeed, we discovered small-molecule inhibitors of this eIF4E/eIF4G interaction (4EGIs) that inhibit translation initiation both in vitro and in vivo and were used successfully in numerous cancer-biology and neurobiology studies. However, their detailed molecular mechanism of action has remained elusive. Here, we show that the eIF4E/eIF4G inhibitor 4EGI-1 acts allosterically by binding to a site on eIF4E distant from the eIF4G binding epitope. Data from NMR mapping and high-resolution crystal structures are congruent with this mechanism, where 4EGI-1 attaches to a hydrophobic pocket of eIF4E between ß-sheet2 (L60-T68) and α-helix1 (E69-N77), causing localized conformational changes mainly in the H78-L85 region. It acts by unfolding a short 310-helix (S82-L85) while extending α-helix1 by one turn (H78-S82). This unusual helix rearrangement has not been seen in any previous eIF4E structure and reveals elements of an allosteric inhibition mechanism leading to the dislocation of eIF4G from eIF4E.