Bladder cancer therapy using a conformationally fluid tumoricidal peptide complex.

Partially unfolded alpha-lactalbumin forms the oleic acid complex HAMLET, with potent tumoricidal activity. Here we define a peptide-based molecular approach for targeting and killing tumor cells, and evidence of its clinical potential (ClinicalTrials.gov NCT03560479). A 39-residue alpha-helical peptide from alpha-lactalbumin is shown to gain lethality for tumor cells by forming oleic acid complexes (alpha1-oleate). Nuclear magnetic resonance measurements and computational simulations reveal a lipid core surrounded by conformationally fluid, alpha-helical peptide motifs. In a single center, placebo controlled, double blinded Phase I/II interventional clinical trial of non-muscle invasive bladder cancer, all primary end points of safety and efficacy of alpha1-oleate treatment are reached, as evaluated in an interim analysis. Intra-vesical instillations of alpha1-oleate triggers massive shedding of tumor cells and the tumor size is reduced but no drug-related side effects are detected (primary endpoints). Shed cells contain alpha1-oleate, treated tumors show evidence of apoptosis and the expression of cancer-related genes is inhibited (secondary endpoints). The results are especially encouraging for bladder cancer, where therapeutic failures and high recurrence rates create a great, unmet medical need.

General Guidelines for Sample Preparation Strategies in HR-µMAS NMR-based Metabolomics of Microscopic Specimens.

The study of the metabolome within tissues, organisms, cells or biofluids can be carried out by several bioanalytical techniques. Among them, nuclear magnetic resonance (NMR) is one of the principal spectroscopic methods. This is due to a sample rotation technique, high-resolution magic angle spinning (HR-MAS), which targets the analysis of heterogeneous specimens with a bulk sample mass from 5 to 10 mg. Recently, a new approach, high-resolution micro-magic angle spinning (HR-µMAS), has been introduced. It opens, for the first time, the possibility of investigating microscopic specimens (<500 µg) with NMR spectroscopy, strengthening the concept of homogeneous sampling in a heterogeneous specimen. As in all bioanalytical approaches, a clean and reliable sample preparation strategy is a significant component in designing metabolomics (or -omics, in general) studies. The sample preparation for HR-µMAS is consequentially complicated by the µg-scale specimen and has yet to be addressed. This report details the strategies for three specimen types: biofluids, fluid matrices and tissues. It also provides the basis for designing future µMAS NMR studies of microscopic specimens.

Size Determination of Protein Oligomers/Aggregates Using Diffusion NMR Spectroscopy.

Diffusion-ordered spectroscopy (DOSY) is a widely used NMR technique for the identification of different chemical moieties/compounds contained in mixtures and has been successfully employed for the separation of small molecules based on hydrodynamic radii. Herein we show that DOSY can also be applied for the size determination of larger biomolecules such as proteins and protein oligomers/aggregates. Proof-of-principle is first shown with a cross-linked oligomeric protein mixture where the hydrodynamic volumes of each component are estimated and subsequently verified with size-exclusion HPLC and SDS polyacrylamide gel electrophoresis. We then determine the sizes of protein oligomers contained in a protein solution subjected under amyloid fibrillogenesis conditions. These studies aim to provide insight into the kinetics behind protein aggregation involved in amyloidosis as well as to determine the hydrodynamic radii of proteins within the mixture.

Fetal heart deceleration patterns in relation to asphyxia and weight-gestational age percentile of the fetus.

The relation between fetal heart rate deceleration patterns and fetal asphyxia and the weight-gestational age percentile characteristics of the fetus has been reviewed from data obtained from 533 patients monitored during the intrapartum period. There is a significant relation between the frequency of total and late decelerations and fetal asphyxia, again confirming this as a useful criterion in the identification of fetal asphyxia. There is a significant relation between decreasing weight-gestational age percentile of the fetus and increasing frequency of total and late decelerations, identifying this fetal characteristic as another variable which will influence fetal heart rate deceleration patterns. The appreciation of the effect of this variable is useful in the interpretation of the fetal heart rate record during the intrapartum period.

Maternal and neonatal implications of congenital complete heart block in the fetus.

The detection of congenital complete heart block (CCHB) in a fetus should alert the obstetrician that the welfare of both the mother and the newborn infant may be in jeopardy. An awareness of this uncommon cause of fetal bradycardia and judicious intrapartum monitoring can avert hasty and unnecessary cesarean section for suspected fetal asphyxia. Neonatal consequences may range from no effect to life-threatening congestive heart failure. The apparently healthy mothers of these infants may be at increased risk for the subsequent development of collagen vascular disease. Three recent case reports demonstrate the spectrum of neonatal and maternal disease that may accompany CCHB. The significance of abnormal serology suggesting a propensity for collagen vascular disease in an otherwise healthy parturient is discussed, and a program for follow-up is proposed.

Caspofungin for invasive fungal infections: combination treatment with liposomal amphotericin B in children undergoing hemopoietic stem cell transplantation.

Invasive fungal infections often prove difficult to eradicate especially in the stem cell transplant setting. Amphotericin has been the mainstay of treatment for years but has significant toxicity. Newer antifungal agents, such as caspofungin, have shown promising results in adults, particularly when used in combination with amphotericin as both drugs differ in their mode of action. However, there are few data from children and no previous published information about the use of Caspofungin after paediatric stem cell transplantation. We report our experience in children with proven invasive fungal infections after stem cell transplantation. This combination was non-toxic, and two of three patients survived their infections.

Intrapartum fetal heart rate profiles with and without fetal asphyxia.

Fetal heart rate profiles for periods up to 12 hours prior to delivery have been reviewed in 515 patients with a fetus at risk. Mechanisms other than fetal asphyxia will cause fetal heart rate decelerations, and fetal asphyxia may in some instances develop in the absence of total or late decelerations. However, an increasing incidence of total decelerations and late decelerations and particularly a marked pattern of total decelerations and late decelerations are of value in the prediction of fetal asphyxia. Fetal heart rate deceleration patterns can predict the probability of fetal asphyxia at the time of initial intervention, while a progression of fetal heart rate deceleration patterns in the individual fetus can be of assistance in the subsequent scheduling of serial acid-base assessments during labor.

Comparative study of sodium nitroprusside induced vasodilation of human placental veins from premature and full-term normotensive and preeclamptic pregnancy.

The objective of this study was to determine whether nitrovasodilator (nitric oxide donor) drug induced relaxation is different between preeclamptic and normotensive pregnancy. Human placental veins were collected at the time of delivery from normotensive (n = 6 premature, n = 6 full term) and preeclamptic (n = 6 premature, n = 3 full term) women and cut into rings. Cumulative concentration-response curves were performed on 5-hydroxytryptamine-contracted rings for the nitrovasodilator agent, sodium nitroprusside (SNP). The EC50 values were determined from the SNP concentration-response curves of the individual subjects, and were compared across gestational age and between preeclamptic and normotensive pregnancy by two-way ANOVA. There was no difference in the SNP EC50 values for normotensive and preeclamptic human placental veins in either the premature or full-term study groups. However, for both normotensive and preeclamptic pregnancy, the SNP EC50 value for full-term placental vein was greater than that for premature tissue. The sensitivity of human placental veins to SNP is similar between preeclamptic and normotensive pregnancy at similar gestational ages, but is increased in premature compared with full-term pregnancy in both preeclamptic and normotensive pregnant women.

Intrapartum fetal asphyxia: clinical characteristics, diagnosis, and significance in relation to pattern of development.

The clinical and fetal heart rates and acid-base characteristics and their sequelae have been reviewed in 587 patients. The relevant clinical factors in the asphyxia group were the preterm fetus, the intrauterine growth retarded fetus, maternal toxemia, and midforceps delivery. The duration of the developing metabolic acidosis in the asphyxia group ranged from terminal to the last two hours of labor. Marked patterns of total decelerations and moderate and marked patterns of late decelerations are of predictive value in the diagnosis of intrapartum fetal asphyxia with a trend to an increased incidence in the longer duration categories, between four and two hours prior to delivery, and a significant increase in all categories during the last two hours of labor. The significance of intrapartum fetal asphyxia to the newborn infant is evident from the low Apgar scores, increased incidence of moderate and severe respiratory distress syndrome, and central nervous system complications in the asphixia group in relation to the normal group.

The incidence of fetal asphyxia in six hundred high-risk monitored pregnancies.

Six hundred high-risk monitored obstetric patients were reviewed for evidence of fetal asphyxia at delivery. The over-all incidence was 20 per cent, i.e., 8 times the incidence in a normal obstetric population. Highly significant indicators of risk for asphyxia were severe toxemia (79 per cent), prematurity with further medical or obstetric complications (36 per cent), and clinical fetal distress, particularly meconium staining with fetal heart rate abnormality (33 per cent). All obstetric, medical, or gestational complications in this review were associated with an increased risk for fetal asphyxia when compared to that in a normal obstetric population.

The acid-base and biochemical characteristics of intrapartum fetal asphyxia.

The maternal and fetal acid-base, lactate, and pyruvate characteristics during the course of labor and at delivery were studied in 124 patients delivered of an infant with evidence of metabolic acidosis at delivery. This metabolic acidosis is principally caused by hyperlactatemia resulting from the tissue oxygen debt accompanying fetal asphyxia. Hypoxemia was one mechanism contributing to this fetal asphyxia and tissue oxygen debt. This evidence of fetal asphyxia developed during the last half and principally during the last two hours of the intrapartum period. Acid-base assessment of fetal blood with identification of a metabolic acidosis will provide an accurate objective diagnosis of intrapartum fetal asphyxia.

Clinical characteristics of pregnancies complicated by intrapartum fetal asphyxia.

The clinical characteristics of 124 pregnancies complicated by intrapartum fetal asphyxia have been reviewed. The evidence of fetal asphyxia tends to appear earlier in patients with maternal medical and obstetric complications than in those with labor complications. Evidence of clinical fetal distress was present in 36 per cent and was not related to the severity of the asphyxia. Low Apgar scores occurred in 40 per cent of infants with moderate asphyxia and in 80 per cent of infants with severe asphyxia at delivery. In the newborn infants, clinical evidence of cerebral abnormality was observed in 3 per cent, and evidence of the respiratory distress syndrome was seen in 3 per cent of the study group.

The probability of fetal metabolic acidosis during labor in a population at risk as determined by clinical factors.

The clinical data derived from 2,772 pregnancies managed in an intrapartum intensive care unit have been analyzed to establish which criteria will indicate in a more definite manner the probability that fetal metabolic acidosis will occur during labor and delivery. All antepartum and intrapartum clinical factors indicate a pregnancy and fetus with an increased probability of fetal metabolic acidosis. However, there is a remarkably consistent relationship between decreasing fetal weight in each week of gestational age and in increasing probability of fetal metabolic acidosis that will permit the magnitude of the risk to be determined with greater precision. The following clinical guidelines are proposed: (1) Current antepartum and intrapartum risk factors are appropriate for the selection of patients for intrapartum intensive care. (2) An accurate gestational age and an estimate of fetal weight within 200 gm will provide an indication of the probability of fetal metabolic acidosis in the individual fetus ranging from 15% to 50%. (3) The presence of meconium in the amniotic fluid increases the probability of metabolic acidosis as defined by fetal weight in relation to gestational age.

The prediction of intrapartum fetal metabolic acidosis by fetal heart rate monitoring.

Fetal heart rate characteristics during the 8 hours prior to delivery have been studied in 200 patients in whom the fetus had evidence of a metabolic acidosis at delivery, and compared to those in 200 patients in whom the fetus had a normal acid-base at delivery. Baseline fetal heart rate moderate bradycardia and tachycardia, decreased baseline variability, and decreased fetal heart rate accelerations are predictors of intrapartum fetal hypoxia with metabolic acidosis. Marked patterns of total decelerations and late decelerations are predictive of intrapartum fetal hypoxia with metabolic acidosis. The probability of fetal metabolic acidosis in the presence of a marked pattern of total decelerations is 25%, and with late decelerations it is 48%, in a population of high-risk pregnancies.

The effect of maternal, labor, and fetal factors upon fetal heart rate during the intrapartum period.

The effect of maternal labor, and fetal characteristics upon fetal heart rate behavior during the intrapartum period has been studied in 400 patients. Abnormal labor in comparison to normal labor has a higher baseline fetal heart rate with an increased incidence of baseline tachycardia and an increased incidence of absent or decreased baseline variability. A decreasing fetal weight gestational age percentile is associated with an increased incidence of variable decelerations. Segmental epidural and Demerol analgesia carefully administered has little effect upon fetal heart rate behavior.