State-wide utilization and performance of traditional and cell-free DNA-based prenatal testing pathways: the Victorian Perinatal Record Linkage (PeRL) study.

OBJECTIVES: To perform individual record linkage of women undergoing screening with cell-free DNA (cfDNA), combined first-trimester screening (CFTS), second-trimester serum screening (STSS), and/or prenatal and postnatal cytogenetic testing with the aim to (1) obtain population-based estimates of utilization of prenatal screening and invasive diagnosis, (2) analyze the performance of different prenatal screening strategies, and (3) report the residual risk of any major chromosomal abnormality following a low-risk aneuploidy screening result. METHODS: This was a retrospective study of women residing in the state of Victoria, Australia, who underwent prenatal screening or invasive prenatal diagnosis in 2015. Patient-funded cfDNA referrals from multiple providers were merged with state-wide results for government-subsidized CFTS, STSS and invasive diagnostic procedures. Postnatal cytogenetic results from products of conception and infants up to 12 months of age were obtained to ascertain cases of false-negative screening results and atypical chromosomal abnormalities. Individual record linkage was performed using LinkageWizTM . RESULTS: During the study period, there were 79 140 births and 66 166 (83.6%) women underwent at least one form of aneuploidy screening. Linkage data were complete for 93.5% (n = 61 877) of women who underwent screening, and of these, 73.2% (n = 45 275) had CFTS alone, 20.2% (n = 12 486) had cfDNA alone; 5.3% (n = 3268) had STSS alone, 1.3% (n = 813) had both CFTS and cfDNA, and < 0.1% (n = 35) had both STSS and cfDNA. CFTS had a combined sensitivity for trisomies 21 (T21), 18 (T18) and 13 (T13) of 89.57% (95% CI, 82.64-93.93%) for a screen-positive rate (SPR) of 2.94%. There were 12 false-negative results in the CFTS pathway, comprising 10 cases of T21, one of T18 and one of T13. cfDNA had a combined sensitivity for T21, T18 and T13 of 100% (95% CI, 95.00-100%) for a SPR of 1.21%. When high-risk cfDNA results for any chromosome (including the sex chromosomes) and failed cfDNA tests were treated as screen positives, the SPR for cfDNA increased to 2.42%. The risk of any major chromosomal abnormality (including atypical abnormalities) detected on prenatal or postnatal diagnostic testing after a low-risk screening result was 1 in 1188 for CFTS (n = 37) and 1 in 762 for cfDNA (n = 16) (P = 0.13). The range of chromosomal abnormalities detected after a low-risk cfDNA result included pathogenic copy-number variants (n = 6), triploidy (n = 3), rare autosomal trisomies (n = 3) and monosomy X (n = 2). CONCLUSIONS: Our state-wide record-linkage analysis delineated the utilization and clinical performance of the multitude of prenatal screening pathways available to pregnant women. The sensitivity of cfDNA for T21, T18 and T13 was clearly superior to that of CFTS. While there was no statistically significant difference in the residual risk of any major chromosomal abnormality after a low-risk CFTS or cfDNA result, there were fewer live infants diagnosed with a major chromosomal abnormality in the cfDNA cohort. These data provide valuable population-based evidence to inform practice recommendations and health policies. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.

A comparison of drug substance predicted chemical stability with ICH compliant stability studies.

OBJECTIVE: The aim of this study is to demonstrate the applicability of predictive stability studies to the degradation of drug substances. SIGNIFICANCE: The use of predicted stability studies during pharmaceutical development and in regulatory submissions is increasing, particularly in early phase to support an initial retest period/shelf life claim in the absence of standard stability data. These studies offer an alternative to standard stability testing and can facilitate clinical trials to be started earlier and medicines to reach patients faster. They involve a short-term stressed stability study, typically designed to degrade a drug substance or product to the specification level of the shelf life limiting attribute. The results are used to predict degradation under long-term storage conditions and enable stability understanding to be gained over a short time frame, using limited amounts of material. METHODS: In this work, Accelerated Stability Assessment Program (ASAP) studies were performed for 10 different drug substances and the predictions obtained for chemical degradation were compared to ICH compliant stability data. RESULTS: Across the studies good agreement was achieved, with the initial retest period predictions from the ASAP studies being conservative by design. When minimal degradation was observed during an ASAP study, it was demonstrated that at least a 12-month initial retest period could be supported. CONCLUSION: This comparison of ASAP predictions and ICH compliant stability data has demonstrated the ability of well-designed ASAP studies to predict the long-term chemical stability of drug substances.

Population-based trends in invasive prenatal diagnosis for ultrasound-based indications: two decades of change from 1994 to 2016.

OBJECTIVE: To assess trends in ultrasound-indicated prenatal diagnostic testing performed over the past two decades in the Australian state of Victoria, in the context of rapidly changing practices in aneuploidy screening and chromosome analysis. METHODS: This was a retrospective analysis of all ultrasound-indicated prenatal diagnostic testing (amniocentesis and chorionic villus sampling) performed in the state of Victoria between 1994 and 2016. Ultrasound indications for testing included: fetal structural abnormality, fetal death, fetal growth restriction, abnormal amniotic fluid volume, genetic 'soft marker' and unspecified ultrasound abnormality. Maternal age, indication for testing, type of diagnostic procedure, gestational age, type of chromosome analysis (G-banded karyotyping or chromosomal microarray (CMA)) and test results were obtained. Diagnostic yield (i.e. percentage of tests yielding a major abnormality) was analyzed by year, maternal age and gestational age. Statistical analysis was performed using the χ2 tests for trend or difference in proportions, as appropriate. RESULTS: During the 23-year study period, 1 533 317 births were recorded and 16 152 diagnostic procedures were performed for the primary indication of ultrasound abnormality. In recent years, ultrasound abnormality became the most common indication for prenatal invasive testing (29.4% of diagnostic tests between 2013 and 2016) due to a steep decline in testing for other indications such as positive result on combined first-trimester screening or advanced maternal age alone. In 2016, over 95% of ultrasound-indicated procedures were performed with CMA; among these, pathogenic copy number variant (CNV) was the most common (3.5%) abnormality detected, followed by trisomy 21 (2.8%). The diagnostic yield of ultrasound-indicated tests performed < 16 weeks was significantly higher than that of tests performed after 20 weeks (31.5% vs 9.0%). CONCLUSIONS: Ultrasound-indicated invasive testing is contributing to prenatal diagnosis in new ways in the genomic era. A pathogenic CNV is now the most likely diagnosis after ultrasound-indicated testing, rather than trisomy 21 or other whole-chromosome aneuploidy. Despite steady improvements in first-trimester screening for aneuploidy, the diagnostic yield of ultrasound-indicated tests > 20 weeks has remained stable due to increased utilization of CMA. Procedures performed for structural abnormalities < 16 weeks continue to have the highest diagnostic yield, supporting the benefits of early fetal structural assessment at 11-13 weeks. Copyright © 2018 ISUOG. Published by John Wiley & Sons Ltd.

Prenatal diagnostic testing and atypical chromosome abnormalities following combined first-trimester screening: implications for contingent models of non-invasive prenatal testing.

OBJECTIVES: To investigate by means of a population-based analysis of a cohort of women who underwent combined first-trimester screening (CFTS), changes in uptake of invasive prenatal diagnosis according to risk of trisomy 21 (T21) on CFTS, and prevalence and methods for ascertainment of atypical chromosome abnormalities. METHODS: This was a retrospective cohort study using state-wide prenatal datasets from Victoria, Australia. A three-step approach was taken to analyze the data: (1) linkage of records between serum screening and diagnostic results; (2) comparison of rates of diagnostic testing according to CFTS T21 risk result category in a 2014-2015 cohort with those of a historical 2002-2004 cohort; (3) detailed analysis of atypical abnormalities in the 2014-2015 group according to CFTS T21 risk result, individual serum analyte level and other indications for invasive diagnostic testing. RESULTS: In 2014-2015, there were 100 418 CFTS results issued for 146 776 births (68.4%). The overall prevalence of atypical chromosome abnormalities in the entire CFTS cohort was 0.10% and was highest in those with CFTS T21 risk > 1 in 10 (4.6%), or serum analyte levels < 0.2 multiples of the median (MoM) (6.9% for pregnancy-associated plasma protein-A (PAPP-A) and 5.2% for beta-human chorionic gonadotropin (ß-hCG)). Almost half (49.2%) of women with PAPP-A < 0.2 MoM had a risk for T21 on CFTS of less than 1 in 100. The majority (55%) of atypical abnormalities occurred in women with CFTS T21 risk below 1 in 300, and were most commonly detected on ultrasound examination (47.1%). CONCLUSION: Concerns regarding missed diagnoses of atypical chromosome abnormalities when non-invasive prenatal testing is offered after a result of high risk on CFTS can be mitigated if invasive diagnostic testing is offered to those women with CFTS T21 risk of > 1 in 100, serum PAPP-A or ß-hCG < 0.2 MoM, or ultrasound-detected abnormality. This has implications for contingent models of screening. Copyright © 2017 ISUOG. Published by John Wiley & Sons Ltd.

Growth on stimulant medication; clarifying the confusion: a review.

AIMS: To get an overview of the studies of growth in height in children with attention deficit hyperactivity disorder (ADHD) treated with stimulant medication, to establish the consistencies and to try to resolve the discrepancies. METHODS: Twenty nine studies were reviewed following a Medline search: 22 related to children, six to late adolescents or adults, and one to children and adults. RESULTS: Children: Eleven studies gave results consistent with height attenuation on stimulant medication: eight were longitudinal, one was cross-sectional, and two showed growth rebound on ceasing medication. Studies with negative findings were inadequately powered (n = 3), lacked controls or statistical analysis (n = 3), measured height velocity without reference to treatment duration (n = 2), or used inappropriate growth parameters (n = 1), controls (n = 1), or normative data (n = 1). Late adolescents/adults treated with stimulant medication in childhood: Two studies associated childhood gastrointestinal side effects with attenuated late adolescent or adult height; all six cross-sectional studies had negative findings. The methodologies varied widely but there was some consistency in the degree of attenuation shown in studies with positive findings. The most sensitive methods analysed the changes in z-scores (standard deviation scores) or calculated the height deficits from paired measurements taken before and after an initial period of treatment with stimulant medication. The height deficit amounted to approximately 1 cm/year during the first 1-3 years of treatment. CONCLUSIONS: Further research is needed into the causal mechanisms, the rate of physical maturation, and the long term implications for final stature.

Slowing of growth in height and weight on stimulants: a characteristic pattern.

OBJECTIVE: The aims of the present study were to describe the growth pattern of children starting stimulant medication and to analyse the changes over time in height, weight and height velocity in a cohort of treated patients. METHODS: Retrospective review of growth data from files of all newly treated patients with attention-deficit/hyperactivity disorder in one paediatric practice. Forty-four boys and seven girls were treated for 6-42 months with either dexam-phetamine (n = 32) or methylphenidate (n = 19). RESULTS: During the first 6 months on stimulant medication 44 children (86%) had a height velocity below the age-corrected mean and there was weight loss in 39 (76%). The height and weight standard deviation score (SDS) showed a progressive decline that was statistically significant after 6 and 18 months (P < 0.001, paired t-test). The height velocity was significantly attenuated for the first 30 months (P < 0.01), being lowest during the first 6 months. The mean height deficit during the first 2 years was approximately 1 cm/year. The change in weight SDS was 2.4 times the change in height SDS after 30 months on treatment with a significant correlation (Pearson's correlation coefficient r = 0.88, P < 0.001). CONCLUSIONS: Stimulant medication is associated with a decrease in height and weight SDS during the first 6-30 months with a characteristic pattern on the growth chart.

A wireless telemetry system for training users of upper limb prostheses.

Telemetry from prosthetic systems is useful in aiding the users and clinical team in setting up and training and monitoring in the use of a prosthesis. The different forms are discussed and a new radio frequency based telemetry system is described. This system uses a computer to display the data and a video mixer to place the information on the same screen as the image of the user so that their actions and control signals can be simultaneously recorded for later study.

Open spina bifida: a complete cohort reviewed 25 years after closure.

This study summarises the long-term results of non-selective treatment of open spina bifida. Every member of a consecutive series of 117 cases was ascertained 22 to 28 years after closure of the back. 56 of the cohort had died. The condition of the 61 survivors ranged from normal to severe disability. Only 33 of the survivors were capable of living independently, 11 required supervision and some help, and 17 needed daily care which was generally provided by the parents. The two main determinants of disability and dependency were the extent of the neurological deficit and IQ. Improvements in the management of patients with open spina bifida have greatly reduced mortality, but they are less likely to influence long-term disability since that is dependent on the severity of the neurological deficit.

Retrospective study of children with renal scarring associated with reflux and urinary infection.

OBJECTIVE: To review the histories of children with bilateral renal scarring and severe vesicoureteric reflux to determine whether an improvement in early management might reduce the risk of scarring. DESIGN: Retrospective study of medical records and discussion with parents. SETTING: Outpatient departments of two teaching hospitals. PATIENTS: 52 children aged 1-12 years participating in a randomised comparison of medical and surgical management. All had a history of symptomatic urinary tract infection. Two thirds presented with fever and two with hypertension or renal failure. In only one out of 32 children examined by antenatal ultrasonography was an abnormality suspected. RESULTS: There was delay in diagnosis or appropriate imaging or effective treatment of urinary infection in 50 of the 52 children. In 41 there was delay in diagnosis; there was delay in treating a confirmed infection in 45; no antibacterial prophylaxis was prescribed before imaging in 28; and investigation of the urinary tract was delayed in 33. The severity of scarring was significantly related to delay in diagnosis (chi 2 for trend 7.43, P = 0.01). Four children of mothers known to have reflux nephropathy were not investigated until they developed urinary tract infection. CONCLUSIONS: Efforts to reduce the incidence and severity of renal scarring should be directed towards rapid diagnosis and effective early management of urinary tract infection in infancy and childhood. Siblings and offspring of known patients with severe reflux nephropathy should be investigated for reflux.

Anthelmintic resistance in Australian sheep nematode populations.

The resistance status of gastro-intestinal nematodes to anthelmintics was evaluated on 881 sheep farms throughout Australia during 1991-92. Resistance was shown to be widespread. Overall, 85% of farms had sheep infected with nematodes resistant to benzimidazole, 65% to levamisole and 34% to combination (benzimidazole+levamisole) products. Resistance to ivermectin was not detected. On only 9% of farms did all anthelmintic groups reduce egg counts by greater than or equal to 95%. The culture of faeces from untreated sheep showed Telodorsagia circumcincta, Trichostrongylus spp, Chabertia ovina and Haemonchus contortus to be the principal species. The nature and prevalence of resistance was not significantly correlated with stocking rate. However, resistance to combination products was almost twice as prevalent on farms in areas with an average annual rainfall of greater than 500 mm.

Synthesis and biological activity of a series of diaryl-substituted alpha-cyano-beta-hydroxypropenamides, a new class of anthelmintic agents.

A series of alpha-cyano-beta-hydroxypropenamides was prepared and tested for anthelmintic activity. alpha-Cyano-beta-hydroxy-N-[4- (trifluoromethyl)phenyl]-3-[4-(trifluoromethyl)phenyl]propenamide (1) showed good activity against the nematode Nematospirodes dubius in a mixed parasite infection in mice; several of the analogues were also effective against the cestode Hymenolepis nana. In sheep trials, 1 caused 100% reduction of the hematophagous nematode Haemonchus contortus after a single dose of 20 mg/kg but did not show satisfactory control of Trichostrongylus colubriformis or Ostertagia circumcincta. Against the liver fluke Fasciola hepatica, 1 suppressed egg production but only temporarily, suggesting that the adult flukes were not eliminated. Mechanism of action studies on 1 using Ascaris mitochondria showed it to be an uncoupler of oxidative phosphorylation.

Synthesis and anthelmintic activity of 3'-benzoylurea derivatives of 6-phenyl-2,3,5,6-tetrahydroimidazo[2,1-b]thiazole.

Reaction of 3-amino derivatives of the nematocides tetramisole and levamisole with variously substituted benzoylisocyanates gave a series of benzoylureas I which were tested for activity against helminths and ectoparasites. Compounds bearing 2,6-difluoro and 4-trifluoromethyl substituents had potent nematocidal activity in both mice and sheep. No antiectoparasitic activity was observed.

Plasma prolactin concentrations in pinealectomized ewes receiving melatonin treatment and in pineal intact ewes maintained under a non-24-hour photoperiod.

We have investigated the profiles of prolactin secretion in relation to onset of breeding activity in Suffolk-Cross ewes with artificially modified melatonin rhythms. With treatments commencing in midsummer, groups of ewes were 1) subjected to a 8L:16D photoperiod, 2) maintained under a "short" (6L:16D) photoperiod repeated over a 22-h cycle that induces an elevation in plasma melatonin that does not endure for the entire dark phase, 3) pinealectomized (pnx) to abolish plasma melatonin levels, and 4) pinealectomized and treated with a melatonin implant (subcutaneous) to provide a constant (no 24-h rhythm) elevation in plasma melatonin. The onset of breeding activity was significantly advanced in both the 8L:16D and pnx/implant groups compared to the 6L:16D and untreated pnx ewes. The two latter groups displayed a normal timing in seasonal breeding activity. Low and high plasma prolactin levels corresponded with short and long photoperiods during both the 24 and 22-h cycles. There was no clearcut "seasonal" rhythm in plasma prolactin in either of the pnx groups. A clear differentiation was seen between reproductive response and prolactin response, particularly in the case of ewes monitored on 22-h cycles of short photoperiod.

Pineal and ovarian response to 22- and 24-h days in the ewe.

Melatonin secretion in ewes was entrained by 22-h light-dark cycles whether of long (16L:6D) or short (6L:16D) photoperiod. In photoperiods of 6L:16D, a phase-delay of melatonin secretion was evident, leading to a dark-phase duration shorter than that found in 8L:16D. Early onset of estrus was induced in anestrous ewes kept in 8L:16D, but not 6L:16D, from 22 July compared to controls in natural light. In photoperiods of 16L:6D, the melatonin profile corresponded precisely to the dark phase. Early offset of estrus was induced in estrous ewes kept in both 18L:6D and 16L:6D from 18 December compared to controls in natural light. Thus, when the duration of melatonin secretion was appropriate to the long photoperiod (16L:6D), but with a constantly changing phase position, a long-day reproductive response was found. Activity-rest cycles were not entrained by 16L:6D; thus the synchronization of melatonin and activity-rest cycles does not appear to be essential for the induction of a long-day reproductive response. These results support the hypothesis that the duration, not the circadian-phase position, of melatonin is critical to the induction of photoperiodic effects.

Use of an intraruminal soluble glass bolus containing melatonin for early lamb production.

The use of melatonin to advance the onset of seasonal oestrus is potentially useful for early lamb production. A number of delivery systems for melatonin have been developed and this study evaluates an intraruminal soluble glass bolus under field conditions. Anoestrous, non-lactating mule and Dorset cross mule ewes were treated in late June with two types of bolus of different solubilities (melatonin 1, 'slow release' and melatonin 2, 'extra slow release'). Rams were introduced in mid-August for a target lambing pattern in mid-January. The percentage pregnancy/lambing rates were 80/78, 97/92 and 100/100 in the control, melatonin 1 and melatonin 2 groups, respectively. The number of lambs born per ewe put to the ram was significantly increased by treatment with melatonin 2 (P less than 0.05) from 1.17 to 1.64 in the mule ewes and 1.64 to 1.72 in the Dorset cross mule ewes. The melatonin containing soluble glass bolus is a novel and convenient method of reducing the number of barren ewes in an early lambing flock.

How does melatonin control seasonal reproductive cycles?

The pineal gland is essential for the perception of photoperiod change in many species. Information about photoperiod length is conveyed through pineal secretion of the methoxyindole melatonin. Melatonin, suitably administered in physiological quantities is equipotent with artificial photoperiod in the induction of photoperiodic responses. Most experimental work suggests that it is the duration of high night time melatonin secretion (positively correlated with the length of the natural or artificial dark phase) which conveys the photoperiodic signal. Continuous release implants induce short day effects in ewes, entirely comparable to daily feeding of melatonin or short photoperiod. A minimum duration of secretion rather than a specific duration is therefore probably critical to short day effects. There appears to be a seasonal variation in sensitivity to short day melatonin effects (induction of early oestrus) which can be shifted to an earlier time of year following one oestrus advance the previous year. Short duration melatonin is read as a long day even secreted with 22 hour periodicity, suggesting a lack of circadian variation in sensitivity to melatonin.

The proposed use of melatonin in controlled sheep breeding.

The regulation by melatonin of hypothalamic-pituitary events in the ewe to advance seasonal oestrous activity, with no undesirable effects upon fertility, and its induction of those seasonal responses associated with short days indicates an essential role for melatonin in controlled-breeding programs in major sheep-producing countries. The development of suitable controlled-release systems to provide a choice of practical methods of melatonin delivery under field conditions is discussed as also are geographical and breed factors in controlled breeding with melatonin.