Reduction of false positives in Newborn Screening

Posted by & filed under Part 03: GENERAL THEMES.

False positive newborn screening can represent a significant burden both emotionally and financially. Morris et al (Genetics in Medicine, 6(16), 2014) describes a protocol designed to reduce false positive screens in infants in a neonatal intensive care unit (NICU) setting. As part of this protocol,  total parenteral nutrition was replaced with dextrose/electrolyte fluid containing IV fluids for 3 h before newborn screening collection. This resulted in a 74% reduction in false-positive results in a NICU.

This simple protocol should be considered in NICUs in which a significant portion of patients are receiving total parenteral nutrition.


Hilary Vernon, MD PhD

MECP2 expression influences aggressive behavior


The X-chromosomal MECP2 gene encodes methyl-CpG-binding protein 2, a transcriptional activator and repressor regulating many other genes. MECP2 is mutated in Rett syndrome. This paper found that that mice with ~50% transgenic overexpression of Mecp2 have enhanced aggressive behavior. When the same transgene was expressed in C57BL/6N mice, transgenics showed reduced aggression and social interaction. This suggests that Mecp2 modulates aggressive social behavior. To test this hypothesis in humans, the authors performed a phenotype-based genetic association study in schizophrenic individuals and found MECP2 SNPs rs2239464 (G/A) and rs2734647 (C/T; 3’UTR) associated with aggression, with the G and C carriers, respectively, being more aggressive. This finding was replicated in an independent schizophrenia cohort. MECP2 mRNA expression in peripheral blood mononuclear cells was also affected by ~50% (rs2734647: C > T).  Finally, the authors showed that the brain-expressed, species-conserved miR-511 binds to MECP2 3′ UTR only in T carriers, thereby suppressing gene expression. In conclusion, this study provides mouse data supporting an interaction between genetic background and mild Mecp2 overexpression and human data that link a genetic variation to MECP2 expression and behavior.

Posted by Nicola Brunetti-Pierri, MD, FACMG

Disease modeling with iPS cell and heart-on-chip technologies

Posted by & filed under Tools.

The study by Wang et al combined patient-derived and genetically engineered induced pluripotent stem cells (iPSCs) and tissue engineering to elucidate the pathophysiology underlying the cardiomyopathy of Barth syndrome, a mitochondrial disorder caused by mutation of the gene encoding tafazzin (TAZ). Using Barth syndrome iPSC-derived cardiomyocytes (iPSC-CMs), they defined metabolic, structural and functional abnormalities associated with TAZ mutation. Barth syndrome iPSC-CMs assembled sparse and irregular sarcomeres, and engineered Barth syndrome ‘heart-on-chip’ tissues contracted weakly. Gene replacement and genome editing demonstrated that TAZ mutation was necessary and sufficient for these phenotypes. Sarcomere assembly and myocardial contraction abnormalities occurred in the context of normal whole-cell ATP levels. Excess levels of reactive oxygen species mechanistically linked TAZ mutation to impaired cardiomyocyte function. This study may serve as a model for elucidating pathophysiology and to investigate novel therapies for other diseases.

Posted by Nicola Brunetti-Pierri, MD, FACMG

Neu-Laxova syndrome is an inborn error of serine biosynthesis

Posted by & filed under Part 08: AMINO ACIDS.

Neu-Laxova syndrome (NLS) is a rare autosomal-recessive disorder characterized by severe fetal growth restriction, microcephaly, a distinct facial appearance, ichthyosis, skeletal anomalies, and perinatal lethality. Through a positional-mapping study combining autozygosity mapping and whole-exome sequencing, Shaheen and colleagues surprisingly found that NLS is caused by mutations in PHGDH encoding for 3-phosphoglycerate dehydrogenase is the first enzyme in the phosphorylated pathway of de novo serine synthesis.  PHGDH mutations were previously known to be responsible for microcephaly and developmental delay. Complete deficiency of PHGDH mouse ortholog recapitulates the key features of NLS. In conclusion, NLS is the extreme end of an inborn error of serine metabolism.

Posted by Nicola Brunetti-Pierri, MD, FACMG

Oxidative stress and X-linked ALD

Posted by & filed under Part 15: PEROXISOMES.

X-linked adrenoleukodystrophy (X-ALD), caused by a mutation in ABCD1, leads to an accumulation of long-chain fatty acids. However, the mechanism by which this accumulation causes disease is not yet understood.  One of the potential factors thought to contribute to disease is oxidative stress and subsequent free-radical damage.

Petrillo et al (Molecular Genetics and Metabolism, 109 (4): 366–370) offers further evidence for oxidative stress in X-ALD. They analyzed glutathione homeostasis in lymphocytes of  a cohort of patients with X-ALD, and found decreased total and reduced glutathione and elevated oxidized glutathione forms. Additionally, they identified decreased of plasma thiols and the high level of carbonyls, another marker of oxidative stress.

For me, results such as these bring up the question of whether or not patients with X-ALD would benefit from therapies aimed at reducing oxidative stress.

Hilary Vernon, MD PhD

A semi-synthetic organism with an expanded genetic alphabet.

Posted by & filed under Part 03: GENERAL THEMES.

Genomic information is generally encoded based on a two-base-pair genetic alphabet (A–T and G–C). In vitro, the alphabet has been expanded to include several unnatural base pairs (UBPs).

One such UBP is the pair formed between d5SICS and dNaM (d5SICS–dNaM) nucleotides bearing hydrophobic nucleobases. d5SICS–dNaM is efficiently PCR-amplified and transcribed in vitro, and its unique mechanism of replication has been previously characterized.

Malyshev et al. (2014) show that an exogenously expressed algal nucleotide triphosphate transporter efficiently imports the triphosphates of both d5SICS and dNaM (d5SICSTP and dNaMTP) into E. coli, and that the endogenous replication machinery uses them to accurately replicate a plasmid containing d5SICS–dNaM. No notable growth burden was reported and the findings suggest that UBP is not efficiently excised by DNA repair pathways.

The authors conclude that the resulting bacterium is the first organism to propagate stably an expanded genetic alphabet.

A semi-synthetic organism with an expanded genetic alphabet. Malyshev DA, Dhami K, Lavergne T, Chen T, Dai N, Foster JM, Corrêa IR Jr, Romesberg FE. Nature. 2014 May 15;509(7500):385-8. PMID: 24805238

Posted by Yannis Trakadis, MD

Genomically recoded organisms

Posted by & filed under Part 03: GENERAL THEMES, Treatment.

The fact that the canonical genetic code is nearly universal is thought to allow natural organisms to share beneficial traits via horizontal gene transfer. However, given that genetically modified organisms also share the same genetic code, they are susceptible to viruses and capable of releasing recombinant genetic material, such as resistance genes, into the environment.

By redefining the genetic code, Lajoie et al hope to produce genomically recoded organisms that are safe and useful. They reassigned the translation function of the UAG stop codon in Escherichia coli without impairing fitness. More specifically, they replaced all known UAG stop codons with synonymous UAA stop codons thereby eliminating natural UAG translation function. UAG was then reassigned as a dedicated codon to genetically encode nonstandard amino acids while avoiding deleterious incorporation at native UAG positions. The engineered E. coli incorporated nonstandard amino acids into its proteins and showed enhanced resistance to bacteriophage T7.

In a second paper, Lajoie et al. demonstrated the recoding of 13 codons in 42 highly expressed essential genes in E. coli. Reassigning sense codons is more challenging because they are more prevalent, and their usage regulates gene expression in ways that are difficult to predict. The results by Lajoie et al. suggest that the genome-wide removal of 13 codons was feasible; however, synonymous codons occasionally were nonequivalent in unpredictable ways.


Genomically recoded organisms expand biological functions. Lajoie MJ, Rovner AJ, Goodman DB, Aerni HR, Haimovich AD, Kuznetsov G, Mercer JA, Wang HH, Carr PA, Mosberg JA, Rohland N, Schultz PG, Jacobson JM, Rinehart J, Church GM, Isaacs FJ. Science. 2013 Oct 18;342(6156):357-60. PMID: 24136966

Probing the limits of genetic recoding in essential genes. Lajoie MJ, Kosuri S, Mosberg JA, Gregg CJ, Zhang D, Church GM. Science. 2013 Oct 18;342(6156):361-3. doi: 10.1126/science.1241460. PMID: 24136967

Posted by Yannis Trakadis, MD

TDP2 protects transcription from abortive topoisomerase activity and is required for normal neural function.

Posted by & filed under Part 28: NEUROGENETICS.

Gomez-Herreros et al. TDP2 protects transcription from abortive topoisomerase activity and is required for normal neural function. Nat Genet. 2014 May;46(5):516-21. doi: 10.1038/ng.2929. Epub 2014 Mar 23.

Gomez-Herreros et al identify homozygous mutations in TDP2 as a new cause of intellectual disability, by performing homozygosity mapping and exome sequencing in a consanguineous Irish family with three brothers suffering from developmental delay accompanied by refractory epilepsy and progressive ataxia.

The molecular pathophysiological mechanism proposed by the authors is particularly interesting. In the course of its function of unwinding DNA strands in order to facilitate transcription, topoisomerase II normally creates double-strand breaks. Most of these breaks are transient, being quickly repaired by the same enzyme. The ones that remain are repaired by tyrosyl DNA phosphodiesterase-2, encoded by TDP2. Cells from affected individuals therefore appear to be hypersensitive to topoisomerase II-induced double-strand breaks, and show inhibition of topoisomerase II-dependent gene transcription; among the transcripts affected, many are required for normal neuronal development and maintenance.

Posted by Alina Levtova, MD


Chitotriosidase and Gaucher disease severity and progression

Posted by & filed under Part 16: LYSOSOMAL DISORDERS.

van Dussen et al. Value of plasma chitotriosidase to assess non-neuronopathic Gaucher disease severity and progression in the era of enzyme replacement therapy. J Inherit Metab Dis. 2014 May 16. [Epub ahead of print]

In a retrospective analysis of 80 patients with non-neuronopathic (type I) Gaucher disease, representing the majority of the cohort of such patients at the Academic Medical Centre in Amsterdam, van Dussen et al evaluate the usefulness of plasma chitotriosidase activity as a marker of response to enzyme replacement therapy and as a predictor of long-term complications/associated diseases (such as Parkinson disease, amyloidosis, pulmonary hypertension, persistent bone disease, hepatocellular carcinoma, and multiple myeloma). They confirm that, prior to the initiation of treatment, plasma chitotriosidase levels are significantly correlated with spleen and liver volumes, as well as hematological parametres and the extent of bone marrow infiltration. Likewise, the reduction of chitotriosidase levels with ERT correlate with the decrease of all of these parametres except for changes of the bone marrow fat fraction. Finally, the occurrence of late complications/associated conditions correlate with pre-treatment and residual chitotriosidase levels.

The authors caution, however, that despite the observed overall correlations, plasma chitotriosidase does not necessarily reflect or predict the clinical status of individual patients (i.e. levels sometimes rose without a clinical deterioration; clinical deterioration could sometimes be observed without an increase of chitotriosidase levels). Likewise, the small number of patients did not allow them to assess the correlation between chitotriosidase levels and each of the long-term complications/associated conditions individually. Nevertheless, the study confirms that plasma chitotriosidase levels are a useful tool in the follow-up of patients with non-neuronopathic Gaucher disease.

Posted by Alina Levtova, MD


A new model of the cardiomyopathy seen in Barth syndrome

Posted by & filed under Part 12: LIPIDS.

Wang et al (Nat Med. 2014 May 11. doi: 10.1038/nm.3545.) just published a fascinating set of experiences in which they created a model of a Barth Syndrome cardiac dysfunction by using “heart on chip” technology. This technology involves using Barth patient derived induced pluripotent stem cell derived cardiomyocytes seeded on micropatterned fibronectin rectangles in order to recapitulate cardiac tissue structure. From these experiments, the authors were able to derive information about sarcomere organization, contractile dysfunction among other relevant parameters. This model, and the success of these experiments, holds great promise in the development of therapies for cardiomyopathy, as well as creating a system in which  phenotype/genotype correlations of specific gene mutations can be better understood.

Hilary Vernon, MD PhD