Pluripotent Stem Cell Platforms for Drug Discovery.

Trends Mol Med. 2018 Jul 10;:

Authors: Chen KG, Mallon BS, Park K, Robey PG, McKay RDG, Gottesman MM, Zheng W

Abstract
Use of human pluripotent stem cells (hPSCs) and their differentiated derivatives have led to recent proof-of-principle drug discoveries, defining a pathway to the implementation of hPSC-based drug discovery (hPDD). Current hPDD strategies, however, have inevitable conceptual biases and technological limitations, including the dimensionality of cell-culture methods, cell maturity and functionality, experimental variability, and data reproducibility. In this review, we dissect representative hPDD systems via analysis of hPSC-based 2D-monolayers, 3D culture, and organoids. We discuss mechanisms of drug discovery and drug repurposing, and roles of membrane drug transporters in tissue maturation and hPDD using the example of drugs that target various mutations of CFTR, the cystic fibrosis transmembrane conductance regulator gene, in patients with cystic fibrosis.

PMID: 30006147 [PubMed - as supplied by publisher]

Age-related heterogeneity in dental caries and associated risk factors in individuals with cystic fibrosis ages 6-20 years: A pilot study.

J Cyst Fibros. 2018 Jul 10;:

Authors: Chi DL, Rosenfeld M, Mancl L, Chung WO, Presland RB, Sarvas E, Rothen M, Alkhateeb A, McNamara S, Genatossio A, Virella-Lowell I, Milla C, Scott J

Abstract
BACKGROUND: The literature conflicts regarding dental caries risk in cystic fibrosis (CF) relative to controls.
METHODS: Prospective, observational study of age-related heterogeneity in caries rates and potential risk factors in individuals with CF ages 6-20 at a single clinic in Washington state (N=85). Caries rates for enrolled CF participants and historical controls from NHANES were compared using cubic spline regression models. Generalized linear regression models identified correlates of age and caries in CF.
RESULTS: Children ages 6-9 with CF had significantly lower caries than controls (Holm's P<0.05). There was no difference for ages 10-20 by CF status (Holm's P>0.05). Various biological/intraoral, medical, and behavioral factors were associated with caries and age in CF.
CONCLUSIONS: Younger children with CF may be protected from caries, but there is apparent loss of protection in early adolescence associated with multiple risk factors. Additional studies are needed to confirm these findings.

PMID: 30005828 [PubMed - as supplied by publisher]

Cell distribution and cytokine levels in induced sputum from healthy subjects and patients with asthma after using different nebulizer techniques.

BMC Pulm Med. 2018 Jul 13;18(1):115

Authors: Koc-Günel S, Schubert R, Zielen S, Rosewich M

Abstract
BACKGROUND: Sputum induction is an important noninvasive method for analyzing bronchial inflammation in patients with asthma and other respiratory diseases. Most frequently, ultrasonic nebulizers are used for sputum induction, but breath-controlled nebulizers may target the small airways more efficiently. This treatment may produce a cell distribution similar to bronchoalveolar lavage (less neutrophils and more macrophages) and provide deeper insights into the underlying lung pathology. The goal of the study was to compare both types of nebulizer devices and their efficacy in inducing sputum to measure bronchial inflammation, i.e., cell composition and cytokines, in patients with mild allergic asthma and healthy controls.
METHODS: The population of this study consisted of 20 healthy control subjects with a median age of 17 years, range: 8-25 years, and 20 patients with a median age of 12 years, range: 8-24 years, presenting with mild, controlled allergic asthma who were not administered an inhaled steroid treatment. We induced sputum in every individual using both devices on two separate days. The sputum weight, the cell composition and cytokine levels were analyzed using a cytometric bead assay (CBA) and by real-time quantitative PCR (qRT-PCR).
RESULTS: We did not observe significant differences in the weight, cell distribution or cytokine levels in the sputum samples induced by both devices. In addition, the Bland-Altman correlation revealed good concordance of the cell distribution. As expected, eosinophils and IL-5 levels were significantly elevated in patients with asthma.
CONCLUSIONS: The hypothesis that sputum induction with a breath-controlled "smart" nebulizer is more efficient and different from an ultrasonic nebulizer was not confirmed. The Bland-Altman correlations showed good concordance when comparing the two devices.
TRIAL REGISTRATION: NCT01543516 Retrospective registration date: March 5, 2012.

PMID: 30005648 [PubMed - in process]

Further delineation of the phenotype caused by loss of function mutations in PRMT7.

Eur J Med Genet. 2018 Jul 10;:

Authors: Valenzuela I, Segura-Puimedon M, Rodríguez-Santiago B, Fernández-Alvarez P, Vendrell T, Armengol L, Tizzano E

Abstract
PRMT7 encodes for an arginine methyltransferase that methylates arginine residues on various protein substrates and has been shown to play a role in various developmental processes. Mutations in PRMT7 have been recently shown to be implicated in a phenotype with intellectual disability, short stature and brachydactyly, and considered to be a phenocopy of pseudohypoparathyroidism. We report a patient with short stature, psychomotor delay, hearing loss and brachydactyly, for whom whole exome sequencing detected two mutations in PRMT7 and parental segregation studies detected biallelic mutation inheritance. Few patients with biallelic PRMT7 mutations have been reported so far in the literature. We report a new patient and review all reported cases to date to delineate the clinical manifestations that may help in diagnosis this disorder, known as Short Stature, Brachydactyly, Intellectual Developmental Disability, and Seizures syndrome, allowing appropriate management and genetic counselling.

PMID: 30006058 [PubMed - as supplied by publisher]

Diaphanospondylodysostosis: Refining the prenatal diagnosis of a rare skeletal disorder.

Eur J Med Genet. 2018 Jul 10;:

Authors: Greenbaum L, Gilboa Y, Raas-Rothschild A, Barel O, Kol N, Wolf HR, Pode-Shakked B, Finezilber Y, Messing B, Berkenstadt M

Abstract
Diaphanospondylodysostosis (DSD) is a rare autosomal recessive skeletal disorder, characterized mainly by ossification defects in vertebrae, thorax malformations, renal cystic dysplasia and usually death in the perinatal period. DSD is caused by mutations in the BMPER (Bone Morphogenetic Protein-Binding Endothelial Cell Precursor-Derived Regulator) gene. We describe the prenatal findings of a non-consanguineous Jewish couple (shared Balkan origin), with three affected fetuses that presented with malformations in the spine and chest, reduced ossification of the skull and spine, horseshoe kidney and increased nuchal translucency. The unique combination of these ultrasound (US) features raised the possibility of DSD, which was confirmed by whole exome sequencing performed on a single fetal DNA and familial segregation. In the three fetuses, a novel homozygous mutation in BMPER (c.410T > A; p. Val137Asp) was found. This mutation, which segregated in the family, was not found in 65 controls of Jewish Balkan origin, and in several large databases. Taken together, the combination of a detailed prenatal US examination and whole exome sequencing may be highly effective in confirming the diagnosis of a rare genetic disease, in this case DSD.

PMID: 30006055 [PubMed - as supplied by publisher]

Familial hypomagnesaemia, Hypercalciuria and Nephrocalcinosis associated with a novel mutation of the highly conserved leucine residue 116 of Claudin 16 in a Chinese patient with a delayed diagnosis: a case report.

BMC Nephrol. 2018 Jul 13;19(1):181

Authors: Lu J, Zhao X, Paiardini A, Lang Y, Bottillo I, Shao L

Abstract
BACKGROUND: Sixty mutations of claudin 16 coding gene have been reported in familial hypomagnesemia with hypercalciuria and nephrocalcinosis (FHHNC) patients. Recent investigations revealed that a highly conserved glycine-leucine-tryptophan (115G-L-W117) motif in the first extracellular segment (ESC1) of claudin 16 might be essential for stabilization of the appropriately folded ECS1 structure and conservation of normal claudin 16 function. However, neither missense nor nonsense mutation has ever been described in this motif. Our study aimed at identifying mutations in a Chinese patient with FHHNC and exploring the association between genotype and phenotype.
CASE PRESENTATION: A 33-year-old female presented with 4 years history of recurrent acute pyelonephritis without other notable past medical history. Her healthy parents, who aged 56 and 53 respectively, were second cousins, and her only sibling died from renal failure without definite cause at age 25. Renal ultrasound imaging demonstrated atrophic kidneys and bilateral nephrocalcinosis. The laboratory workup revealed impaired renal function (Stage CKD IV), hypocalcemia and mild hypomagnesemia, accompanied with marked renal loss of magnesium and hypercalciuria. During the follow-up, treatment with calcitriol and calcium but not with magnesium was difficult to achieve normal serum calcium levels, whereas her serum magnesium concentration fluctuated within normal ranges. In the end, the patient unavoidably reached ESRD at 36 years old. The clinical features and family history suggested the diagnosis of FHHNC. To make a definite diagnosis, we use whole-exome sequencing to identify the disease-causing mutations and Sanger sequencing to confirm the mutation co-segregation in the family. As a result, a novel homozygous mutation (c.346C > G, p.Leu116Val) in 115G-L-W117 motif of claudin 16 was identified. Her parents, grandmother and one of her cousins carried heterozygous p.Leu116Val, whereas 200 unrelated controls did not carry this mutation.
CONCLUSIONS: We described a delayed diagnosis patient with FHHNC in the Chinese population and identified a novel missense mutation in the highly conserved 115G-L-W117 motif of claudin 16 for the first time. According to the reported data and the information deduced from 3D modeling, we speculate that this mutation probably reserve partial residual function which might be related to the slight phenotype of the patient.

PMID: 30005619 [PubMed - in process]

Alternative strategies for lignocellulose fermentation through lactic acid bacteria: the state of the art and perspectives.

FEMS Microbiol Lett. 2018 Aug 01;365(15):

Authors: Tarraran L, Mazzoli R

Abstract
Lactic acid bacteria (LAB) have a long history in industrial processes as food starters and biocontrol agents, and also as producers of high-value compounds. Lactic acid, their main product, is among the most requested chemicals because of its multiple applications, including the synthesis of biodegradable plastic polymers. Moreover, LAB are attractive candidates for the production of ethanol, polyhydroalkanoates, sweeteners and exopolysaccharides. LAB generally have complex nutritional requirements. Furthermore, they cannot directly ferment inexpensive feedstocks such as lignocellulose. This significantly increases the cost of LAB fermentation and hinders its application in the production of high volumes of low-cost chemicals. Different strategies have been explored to extend LAB fermentation to lignocellulosic biomass. Fermentation of lignocellulose hydrolysates by LAB has been frequently reported and is the most mature technology. However, current economic constraints of this strategy have driven research for alternative approaches. Co-cultivation of LAB with native cellulolytic microorganisms may reduce the high cost of exogenous cellulase supplementation. Special attention is given in this review to the construction of recombinant cellulolytic LAB by metabolic engineering, which may generate strains able to directly ferment plant biomass. The state of the art of these strategies is illustrated along with perspectives of their applications to industrial second generation biorefinery processes.

PMID: 30007320 [PubMed - in process]

Chemosensitivity profiling of osteosarcoma tumour cell lines identifies a model of BRCAness.

Sci Rep. 2018 Jul 13;8(1):10614

Authors: Holme H, Gulati A, Brough R, Fleuren EDG, Bajrami I, Campbell J, Chong IY, Costa-Cabral S, Elliott R, Fenton T, Frankum J, Jones SE, Menon M, Miller R, Pemberton HN, Postel-Vinay S, Rafiq R, Selfe JL, von Kriegsheim A, Munoz AG, Rodriguez J, Shipley J, van der Graaf WTA, Williamson CT, Ryan CJ, Pettitt S, Ashworth A, Strauss SJ, Lord CJ

Abstract
Osteosarcoma (OS) is an aggressive sarcoma, where novel treatment approaches are required. Genomic studies suggest that a subset of OS, including OS tumour cell lines (TCLs), exhibit genomic loss of heterozygosity (LOH) patterns reminiscent of BRCA1 or BRCA2 mutant tumours. This raises the possibility that PARP inhibitors (PARPi), used to treat BRCA1/2 mutant cancers, could be used to target OS. Using high-throughput drug sensitivity screening we generated chemosensitivity profiles for 79 small molecule inhibitors, including three clinical PARPi. Drug screening was performed in 88 tumour cell lines, including 18 OS TCLs. This identified known sensitivity effects in OS TCLs, such as sensitivity to FGFR inhibitors. When compared to BRCA1/2 mutant TCLs, OS TCLs, with the exception of LM7, were PARPi resistant, including those with previously determined BRCAness LoH profiles. Post-screen validation experiments confirmed PARPi sensitivity in LM7 cells as well as a defect in the ability to form nuclear RAD51 foci in response to DNA damage. LM7 provides one OS model for the study of PARPi sensitivity through a potential defect in RAD51-mediated DNA repair. The drug sensitivity dataset we generated in 88 TCLs could also serve as a resource for the study of drug sensitivity effects in OS.

PMID: 30006631 [PubMed - in process]

UTX is an escape from X-inactivation tumor-suppressor in B cell lymphoma.

Nat Commun. 2018 Jul 13;9(1):2720

Authors: Li X, Zhang Y, Zheng L, Liu M, Chen CD, Jiang H

Abstract
To explain the excess cancer rate in males, several candidates for "escape from X-inactivation tumor-suppressor" (EXITS) were recently identified. In this report we provide direct experimental evidence supporting UTX's role as an EXITS gene. Using a mouse lymphoma model, we show clear dosage effect of UTX copy number during tumorigenesis, which strongly supports the EXITS theory. Importantly, UTX deletion not only accelerates lymphomagenesis, it also strongly promotes tumor progression. UTX-knockout tumors are more aggressive, showing enhanced brain dissemination and formation of blood vessels. Efnb1 is overexpressed in UTX KO tumors and can lead to such phenotypes. In human patients, lymphomas with low UTX expression also express high levels of Efnb1, and cause significantly poor survival. Lastly, we show that UTX deficiency renders lymphoma sensitive to cytarabine treatment. Taken together, these data highlight UTX loss's profound impacts on tumor initiation and drug response.

PMID: 30006524 [PubMed - in process]

Managing Competing Interests: Partitioning S between Glutathione and Protein Synthesis.

Plant Physiol. 2018 Jul;177(3):867-868

Authors: Mhamdi A

PMID: 30006454 [PubMed - in process]

Structural Basis of Highly Specific Interaction between Nephrin and MAGI1 in Slit Diaphragm Assembly and Signaling.

J Am Soc Nephrol. 2018 Jul 13;:

Authors: Weng Z, Shang Y, Ji Z, Ye F, Lin L, Zhang R, Zhu J

Abstract
BACKGROUND: The slit diaphragm is a specialized adhesion junction between opposing podocytes, establishing the final filtration barrier that prevents passage of proteins from the capillary lumen into the urinary space. Nephrin, the key structural and signaling adhesion molecule expressed in the slit diaphragm, contains an evolutionally conserved, atypical PDZ-binding motif (PBM) reported to bind to a variety of proteins in the slit diaphragm. Several mutations in NPHS1 (the gene encoding nephrin) that result in nephrin lacking an intact PBM are associated with glomerular diseases. However, the molecular basis of nephrin-PBM-mediated protein complexes is still unclear.
METHODS: Using a combination of biochemic, biophysic, and cell biologic approaches, we systematically investigated the interactions between nephrin-PBM and PDZ domain-containing proteins in the slit diaphragm.
RESULTS: We found that nephrin-PBM specifically binds to one member of the membrane-associated guanylate kinase family of scaffolding proteins, MAGI1, but not to another, MAGI2. The complex structure of MAGI1-PDZ3/nephrin-PBM reveals that the Gly at the -3 position of nephrin-PBM is the determining feature for MAGI1-PDZ3 recognition, which sharply contrasts with the typical PDZ/PBM binding mode. A single gain-of-function mutation within MAGI2 enabled nephrin-PBM binding. In addition, using our structural analysis, we developed a highly efficient inhibitory peptide capable of specifically blocking the nephrin/MAGI1 interaction.
CONCLUSIONS: MAGI1 interacts with nephrin-PBM with exquisite specificity. A newly developed, potent inhibitory peptide that blocks this interaction may be useful for future functional investigations in vivo. Our findings also provide possible explanations for the diseases caused by NPHS1 mutations.

PMID: 30006415 [PubMed - as supplied by publisher]

Holistic Approaches in Lipid Production by Yarrowia lipolytica.

Trends Biotechnol. 2018 Jul 10;:

Authors: Lazar Z, Liu N, Stephanopoulos G

Abstract
Concerns about climate change have driven research on the production of lipid-derived biofuels as an alternative and renewable liquid fuel source. Using oleaginous yeasts for lipid synthesis creates the potential for cost-effective industrial-scale operations due to their ability to reach high lipid titer, yield, and productivity resulting from their unique metabolism. Yarrowia lipolytica is the model oleaginous yeast, with the best-studied lipid metabolism, the greatest number of genetic tools, and a fully sequenced genome. In this review we highlight multiomics studies that elucidate the mechanisms allowing this yeast to achieve lipid overaccumulation and then present several major metabolic engineering efforts that enhanced the production metrics in Y. lipolytica. Recent achievements that applied novel engineering strategies are emphasized.

PMID: 30006239 [PubMed - as supplied by publisher]

Corrigendum to "Phylogenomic re-assessment of the thermophilic genus Geobacillus" [Syst. Appl. Microbiol. 39 (2016) 527-533].

Syst Appl Microbiol. 2018 Jul 10;:

Authors: Aliyu H, Lebre P, Blom J, Cowan D, De Maayer P

PMID: 30006234 [PubMed - as supplied by publisher]

Recurrent tumor-specific regulation of alternative polyadenylation of cancer-related genes.

BMC Genomics. 2018 Jul 13;19(1):536

Authors: Xue Z, Warren RL, Gibb EA, MacMillan D, Wong J, Chiu R, Hammond SA, Yang C, Nip KM, Ennis CA, Hahn A, Reynolds S, Birol I

Abstract
BACKGROUND: Alternative polyadenylation (APA) results in messenger RNA molecules with different 3' untranslated regions (3' UTRs), affecting the molecules' stability, localization, and translation. APA is pervasive and implicated in cancer. Earlier reports on APA focused on 3' UTR length modifications and commonly characterized APA events as 3' UTR shortening or lengthening. However, such characterization oversimplifies the processing of 3' ends of transcripts and fails to adequately describe the various scenarios we observe.
RESULTS: We built a cloud-based targeted de novo transcript assembly and analysis pipeline that incorporates our previously developed cleavage site prediction tool, KLEAT. We applied this pipeline to elucidate the APA profiles of 114 genes in 9939 tumor and 729 tissue normal samples from The Cancer Genome Atlas (TCGA). The full set of 10,668 RNA-Seq samples from 33 cancer types has not been utilized by previous APA studies. By comparing the frequencies of predicted cleavage sites between normal and tumor sample groups, we identified 77 events (i.e. gene-cancer type pairs) of tumor-specific APA regulation in 13 cancer types; for 15 genes, such regulation is recurrent across multiple cancers. Our results also support a previous report showing the 3' UTR shortening of FGF2 in multiple cancers. However, over half of the events we identified display complex changes to 3' UTR length that resist simple classification like shortening or lengthening.
CONCLUSIONS: Recurrent tumor-specific regulation of APA is widespread in cancer. However, the regulation pattern that we observed in TCGA RNA-seq data cannot be described as straightforward 3' UTR shortening or lengthening. Continued investigation into this complex, nuanced regulatory landscape will provide further insight into its role in tumor formation and development.

PMID: 30005633 [PubMed - in process]

The affinity of antipsychotic drugs to dopamine and serotonin 5-HT2 receptors determines their effects on prefrontal-striatal functional connectivity.

Eur Neuropsychopharmacol. 2018 Jul 10;:

Authors: Tollens F, Gass N, Becker R, Schwarz AJ, Risterucci C, Künnecke B, Lebhardt P, Reinwald J, Sack M, Weber-Fahr W, Meyer-Lindenberg A, Sartorius A

Abstract
One of the major challenges of cross-species translation in psychiatry is the identification of quantifiable brain phenotypes linked to drug efficacy and/or side effects. A measure that has received increasing interest is the effect of antipsychotic drugs on resting-state functional connectivity (FC) in magnetic resonance imaging. However, quantitative comparisons of antipsychotic drug-induced alterations of FC patterns are missing. Consideration of receptor binding affinities provides a means for the effects of antipsychotic drugs on extended brain networks to be related directly to their molecular mechanism of action. Therefore, we examined the relationship between the affinities of three second-generation antipsychotics (amisulpride, risperidone and olanzapine) to dopamine and serotonin receptors and FC patterns related to the prefrontal cortex (PFC) and striatum in Sprague-Dawley rats. FC of the relevant regions was quantified by correlation coefficients and local network properties. Each drug group (32 animals per group) was subdivided into three dose groups and a vehicle control group. A linear relationship was discovered for the mid-dose of antipsychotic compounds, with stronger affinity to serotonin 5-HT2A, 5-HT2C and 5-HT1A receptors and decreased affinity to D3 receptors associated with increased prefrontal-striatal FC (p = 0.0004, r² = 0.46; p = 0.004, r² = 0.33; p = 0.002, r² = 0.37; p = 0.02, r² = 0.22, respectively). Interestingly, no correlation was observed for the low and high dose groups, and for D2 receptors. Our results indicate that drug-induced FC patterns may be linked to antipsychotic mechanism of action on the molecular level and suggest the technique's value for drug development, especially if our results are extended to a larger number of antipsychotics.

PMID: 30006253 [PubMed - as supplied by publisher]

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How low can we go? The implications of low bacterial load in respiratory microbiota studies.

Pneumonia (Nathan). 2018;10:7

Authors: Marsh RL, Nelson MT, Pope CE, Leach AJ, Hoffman LR, Chang AB, Smith-Vaughan HC

Abstract
Background: Culture-independent sequencing methods are increasingly used to investigate the microbiota associated with human mucosal surfaces, including sites that have low bacterial load in healthy individuals (e.g. the lungs). Standard microbiota methods developed for analysis of high bacterial load specimens (e.g. stool) may require modification when bacterial load is low, as background contamination derived from sterile laboratory reagents and kits can dominate sequence data when few bacteria are present.
Main body: Bacterial load in respiratory specimens may vary depending on the specimen type, specimen volume, the anatomic site sampled and clinical parameters. This review discusses methodological issues inherent to analysis of low bacterial load specimens and recommends strategies for successful respiratory microbiota studies. The range of methods currently used to process DNA from low bacterial load specimens, and the strategies used to identify and exclude background contamination are also discussed.
Conclusion: Microbiota studies that include low bacterial load specimens require additional tests to ensure that background contamination does not bias the results or interpretation. Several methods are currently used to analyse the microbiota in low bacterial load respiratory specimens; however, there is scant literature comparing the effectiveness and biases of different methods. Further research is needed to define optimal methods for analysing the microbiota in low bacterial load specimens.

PMID: 30003009 [PubMed]

A Case Report of Pulmonary Exacerbation after Initiation of Lumacaftor/Ivacaftor Therapy in a CF Female with Complicated Lung Disease.

Case Rep Pulmonol. 2018;2018:8394370

Authors: Hatziagorou E, Kouroukli E, Georgopoulou V, Tsanakas J

Abstract
Novel targeted treatments for Cystic Fibrosis give rise to new hope for an ever-growing number of CF patients with various mutations. However, very little evidence and guidelines exist to steer clinical decisions regarding patients whose illness takes an unexpected course. In such cases, the benefits and risks of discontinuing these treatments must be carefully and individually weighed, since their long-term effects remain mainly uncharted territory. In this report we document the case of a homozygous F508del CF patient with severe lung disease who presented with a pulmonary exacerbation shortly after the beginning of treatment with lumacaftor/ivacaftor and the complicated initial phase of therapy, which was followed by significant improvements.

PMID: 30002941 [PubMed]