Paediatrics (RCH) - Research Publications
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ItemYY1 Haploinsufficiency Causes an Intellectual Disability Syndrome Featuring Transcriptional and Chromatin Dysfunction.(Elsevier BV, 2017-06-01)Yin and yang 1 (YY1) is a well-known zinc-finger transcription factor with crucial roles in normal development and malignancy. YY1 acts both as a repressor and as an activator of gene expression. We have identified 23 individuals with de novo mutations or deletions of YY1 and phenotypic features that define a syndrome of cognitive impairment, behavioral alterations, intrauterine growth restriction, feeding problems, and various congenital malformations. Our combined clinical and molecular data define "YY1 syndrome" as a haploinsufficiency syndrome. Through immunoprecipitation of YY1-bound chromatin from affected individuals' cells with antibodies recognizing both ends of the protein, we show that YY1 deletions and missense mutations lead to a global loss of YY1 binding with a preferential retention at high-occupancy sites. Finally, we uncover a widespread loss of H3K27 acetylation in particular on the YY1-bound enhancers, underscoring a crucial role for YY1 in enhancer regulation. Collectively, these results define a clinical syndrome caused by haploinsufficiency of YY1 through dysregulation of key transcriptional regulators.
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ItemNo Preview AvailableStreamlined genetic education is effective in preparing women newly diagnosed with breast cancer for decision making about treatment-focused genetic testing: a randomized controlled noninferiority trial(NATURE PUBLISHING GROUP, 2017-04)PURPOSE: Increasingly, women newly diagnosed with breast cancer are being offered treatment-focused genetic testing (TFGT). As the demand for TFGT increases, streamlined methods of genetic education are needed. METHODS: In this noninferiority trial, women aged <50 years with either a strong family history (FH+) or other features suggestive of a germ-line mutation (FH-) were randomized before definitive breast cancer surgery to receive TFGT education either as brief written materials (intervention group (IG)) or during a genetic counseling session at a familial cancer clinic (usual-care group (UCG)). Women completed self-report questionnaires at four time points over 12 months. RESULTS: A total of 135 women were included in the analysis, all of whom opted for TFGT. Decisional conflict about TFGT choice (primary outcome) was not inferior in the IG compared with the UCG (noninferiority margin of -10; mean difference = 2.45; 95% confidence interval -2.87-7.76; P = 0.36). Costs per woman counseled in the IG were significantly lower (AUD$89) compared with the UCG (AUD$173; t(115) = 6.02; P < 0.001). CONCLUSION: A streamlined model of educating women newly diagnosed with breast cancer about TFGT seems to be a cost-effective way of delivering education while ensuring that women feel informed and supported in their decision making, thus freeing resources for other women to access TFGT.Genet Med 19 4, 448-456.
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ItemChildhood interstitial lung diseases in immunocompetent children in Australia and New Zealand: a decade's experience.(Springer Science and Business Media LLC, 2017-07-25)BACKGROUND: Childhood interstitial lung disease (chILD) represents a rare heterogeneous group of respiratory disorders. In the absence of randomized controlled clinical trials, global collaborations have utilized case series with an aim to standardising approaches to diagnosis and management. Australasian data are lacking. The aim of this study was to calculate prevalence and report the experience of chILD in Australasia over a decade. METHODS: Paediatric pulmonologists in Australia and New Zealand involved in the care of patients aged 0-18 years with chILD completed a questionnaire on demographics, clinical features and outcomes, over a 10 year period. These data, together with data from the 2 reference genetics laboratories, were used to calculate prevalence. RESULTS: One hundred fifteen cases were identified equating to a period prevalence (range) of 1.5 (0.8-2.1) cases/million for children aged 0-18years. Clinical data were provided on 106 patients: the <2 year group comprised 66 children, median age (range) 0.50 years (0.01-1.92); the ≥2 year group comprised 40 children, median age 8.2 years (2.0-18.0). Management approach was heterogeneous. Overall, 79% of patients had a good clinical outcome. Mortality rate was 7% in the study population. CONCLUSION: chILD is rare in Australasia. This study demonstrates variation in the investigations and management of chILD cases across Australasia, however the general outcome is favorable. Further international collaboration will help finesse the understanding of these disorders.
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ItemAssociation of blood pressure in the supine position with target organ damage in subjects over 60 years old.(SAGE Publications, 2017-02)Objective Toexplore the correlation between blood pressure in the supine position and target organ damage in subjects over 60 years of age. Methods In 2444 individuals, we investigated the association of systolic blood pressure (SBP) in the supine position with the target organ damage indices microalbuminuria (ALBU), brachial-ankle pulse wave velocity (baPWV), and carotid intima-media thickness (IMT). Supine hypertension (SH) is defined as systolic blood pressure ≥ 140 mmHg and/or diastolic blood pressure ≥ 90 mmHg. Subjects were assigned to either the SH group (1275 cases) or the non-SH group (1169 cases). Results The levels of ALBU, baPWV, and IMT, as well as the percentage of participants with ALBU > 30 mg/L, baPWV ≥ 1400 cm/s, and IMT ≥ 1 mm, were significantly higher in the SH group than in the non-SH group. Multivariate logistic regression analysis showed that SH was an independent risk factor for baPWV and IMT, but the relationship with ALBU was not statistically significant after correction for confounding factors. Conclusions SH is a risk factor for target organ damage, as expressed by the indices baPWV and IMT. The association of SH with kidney damage requires further study.
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ItemIncidence, temporal trend and factors associated with ventilator-associated pneumonia in mainland China: a systematic review and meta-analysis(BMC, 2017-07-04)BACKGROUND: Data to date is far from sufficient to describe the recent epidemiology of ventilator-associated pneumonia (VAP) in mainland China. This study aimed to estimate the overall incidence of VAP, with a special focus on its temporal trend and associated factors. METHODS: Meta-analyses of 195 studies published from 2010 to 2015 were conducted, followed by subgroup analyses by methodological quality, pre-defined setting characteristics and attributes of populations. RESULTS: The overall cumulative VAP incidence in mainland China was 23.8% (95% confidence interval (CI) 20.6-27.2%), with the results showing high heterogeneity. The pooled incidence densities were 24.14 (95% CI 21.19-27.51) episodes and 22.83 (95% CI 19.88-26.23) patients per 1000 ventilator-days. A decline in the cumulative incidence was observed from 2006 (49.5%, 95% CI 40.0-59.0%) to 2014 (19.6%, 95% CI 10.4-31.0%); differences in the incidence rates were also documented according to Chinese provinces and diagnostic criteria (p < 0.001). Older age (≥60 years), coma, re-intubation, tracheotomy and prolonged ventilation were the factors significantly associated with the occurrence of VAP. CONCLUSIONS: The incidence of VAP remains high in mainland China but has decreased since 2006. The reported rates vary considerably across individual studies, probably due to variations in diagnosis and geographical region. More studies using standard definitions and cut-off points are needed to better clarify the epidemiology of VAP across the country.
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ItemAdverse Events Associated with Treatment of Multidrug-Resistant Tuberculosis in China: An Ambispective Cohort Study(INT SCIENTIFIC INFORMATION, INC, 2017-05-18)BACKGROUND Adverse events are under-appreciated negative consequences that are significant clinical problems for patients undergoing anti-MDR-TB treatment due to longer duration of treatment and more need for concurrent use of multiple second-line drugs. The aim of this study was to determine the incidence of adverse events and their impact on MDR-TB therapy and treatment outcome, and to identify possible drug-event pairs in China. MATERIAL AND METHODS An ambispective cohort study was conducted based on hospital medical records, which included a retrospective study that enrolled 751 MDR-TB patients receiving standardized regimen between May 2009 and July 2013, and a follow-up investigation of treatment outcome conducted in December 2016 in China. Adverse events were determined according to laboratory results or clinical criteria. Cox's proportional hazards regression models were used for evaluating associations. RESULTS There were 681(90.7%) patients experienced at least 1 type of adverse event and 55.2% of them required a changed MDR-TB treatment; 51(6.8%) patients required permanent discontinuation of the offending drug due to adverse events. The occurrence of adverse events was associated with poor treatment outcome (adjusted hazard ratio, 1.54; 95% CI 1.21, 1.87). A total of 10 different drug-event pairs were identified. CONCLUSIONS Adverse events occurred commonly during MDR-TB treatment in China, and often resulted in MDR-TB treatment change. The occurrence of adverse events affected MDR-TB poor outcome after treatment.
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ItemCaspase-2-mediated cell death is required for deleting aneuploid cells.(Springer Science and Business Media LLC, 2017-05-11)Caspase-2, one of the most evolutionarily conserved of the caspase family, has been implicated in maintenance of chromosomal stability and tumour suppression. Caspase-2 deficient (Casp2-/-) mice develop normally but show premature ageing-related traits and when challenged by certain stressors, succumb to enhanced tumour development and aneuploidy. To test how caspase-2 protects against chromosomal instability, we utilized an ex vivo system for aneuploidy where primary splenocytes from Casp2-/- mice were exposed to anti-mitotic drugs and followed up by live cell imaging. Our data show that caspase-2 is required for deleting mitotically aberrant cells. Acute silencing of caspase-2 in cultured human cells recapitulated these results. We further generated Casp2C320S mutant mice to demonstrate that caspase-2 catalytic activity is essential for its function in limiting aneuploidy. Our results provide direct evidence that the apoptotic activity of caspase-2 is necessary for deleting cells with mitotic aberrations to limit aneuploidy.
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ItemProtocadherin 19 (PCDH19) interacts with paraspeckle protein NONO to co-regulate gene expression with estrogen receptor alpha (ERα).(Oxford University Press (OUP), 2017-06-01)De novo and inherited mutations of X-chromosome cell adhesion molecule protocadherin 19 (PCDH19) cause frequent, highly variable epilepsy, autism, cognitive decline and behavioural problems syndrome. Intriguingly, hemizygous null males are not affected while heterozygous females are, contradicting established X-chromosome inheritance. The disease mechanism is not known. Cellular mosaicism is the likely driver. We have identified p54nrb/NONO, a multifunctional nuclear paraspeckle protein with known roles in nuclear hormone receptor gene regulation, as a PCDH19 protein interacting partner. Using breast cancer cells we show that PCDH19-NONO complex is a positive co-regulator of ERα-mediated gene expression. Expression of mutant PCDH19 affects at least a subset of known ERα-regulated genes. These data are consistent with our findings that genes regulated by nuclear hormone receptors and those involved in the metabolism of neurosteroids in particular are dysregulated in PCDH19-epilepsy girls and affected mosaic males. Overall we define and characterize a novel mechanism of gene regulation driven by PCDH19, which is mediated by paraspeckle constituent NONO and is ERα-dependent. This PCDH19-NONO-ERα axis is of relevance not only to PCDH19-epilepsy and its comorbidities but likely also to ERα and generally nuclear hormone receptor-associated cancers.
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ItemKnockout of the epilepsy gene Depdc5 in mice causes severe embryonic dysmorphology with hyperactivity of mTORC1 signalling.(Springer Science and Business Media LLC, 2017-10-03)DEPDC5 mutations have recently been shown to cause epilepsy in humans. Evidence from in vitro studies has implicated DEPDC5 as a negative regulator of mTORC1 during amino acid insufficiency as part of the GATOR1 complex. To investigate the role of DEPDC5 in vivo we generated a null mouse model using targeted CRISPR mutagenesis. Depdc5 homozygotes display severe phenotypic defects between 12.5-15.5 dpc, including hypotrophy, anaemia, oedema, and cranial dysmorphology as well as blood and lymphatic vascular defects. mTORC1 hyperactivity was observed in the brain of knockout embryos and in fibroblasts and neurospheres isolated from knockout embryos and cultured in nutrient deprived conditions. Heterozygous mice appeared to be normal and we found no evidence of increased susceptibility to seizures or tumorigenesis. Together, these data support mTORC1 hyperactivation as the likely pathogenic mechanism that underpins DEPDC5 loss of function in humans and highlights the potential utility of mTORC1 inhibitors in the treatment of DEPDC5-associated epilepsy.
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ItemVersatile single-step-assembly CRISPR/Cas9 vectors for dual gRNA expression.(Public Library of Science (PLoS), 2017)CRISPR/Cas9 technology enables efficient, rapid and cost-effective targeted genomic modification in a wide variety of cellular contexts including cultured cells. Some applications such as generation of double knock-outs, large deletions and paired-nickase cleavage require simultaneous expression of two gRNAs. Although single plasmids that enable multiplex expression of gRNAs have been developed, these require multiple rounds of cloning and/or PCR for generation of the desired construct. Here, we describe a series of vectors that enable generation of customized dual-gRNA expression constructs via an easy one-step golden gate cloning reaction using two annealed oligonucleotide inserts with different overhangs. Through nucleofection of mouse embryonic stem cells, we demonstrate highly efficient cleavage of the target loci using the dual-guide plasmids, which are available as Cas9-nuclease or Cas9-nickase expression constructs, with or without selection markers. These vectors are a valuable addition to the CRISPR/Cas9 toolbox and will be made available to all interested researchers via the Addgene plasmid repository.