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Tan, P.

Publications and source records attributed to Tan, P..

At least 55 records · Page 3Linked to original sources

Integrative Epigenomic and High-Throughput Functional Enhancer Profiling Reveals Determinants of Enhancer Heterogeneity in Gastric Cancer

BackgroundEnhancers are distal cis-regulatory elements required for cell-specific gene expression and cell fate determination. In cancer, enhancer variation has been proposed as a major cause of inter-patient heterogeneity - however, most predicted enhancer regions remain to be functionally tested. ResultsAnalyzing 128 epigenomic histone modification profiles of primary GC samples, normal gastric tissues, and GC cell lines, we report a comprehensive catalog of 75,730 recurrent predicted enhancers, the majority of which are tumor-associated in vivo (>50,000) and associated with lower somatic mutation rates inferred by whole-genome sequencing. Applying Capture-based Self-Transcribing Active Regulatory Region sequencing (CapSTARR-seq) to the enhancer catalog, we observed significant correlations between CapSTARR-seq functional activity and H3K27ac/H3K4me1 levels. Super-enhancer regions exhibited increased CapSTARR-seq signals compared to regular enhancers even when decoupled from native chromatin contexture. We show that combining histone modification and CapSTARR-seq functional enhancer data improves the prediction of enhancer-promoter interactions and pinpointing of germline single nucleotide polymorphisms (SNPs), somatic copy number alterations (SCNAs), and trans-acting TFs involved in GC expression. Specifically, we identified cancer-relevant genes (e.g. ING1, ARL4C) whose expression between patients is influenced by enhancer differences in genomic copy number and germline SNPs, and HNF4 as a master trans-acting factor associated with GC enhancer heterogeneity. ConclusionsOur study indicates that combining histone modification and functional assay data may provide a more accurate metric to assess enhancer activity than either platform individually, and provides insights into the relative contribution of genetic (cis) and regulatory (trans) mechanisms to GC enhancer functional heterogeneity.

cancer biology↗

PLK1 inhibition selectively kills ARID1A deficient cells through uncoupling of oxygen consumption from ATP production

Inhibitors of the mitotic kinase PLK1 yield objective responses in a subset of refractory cancers. However, PLK1 overexpression in cancer does not correlate with drug sensitivity, and the clinical development of PLK1 inhibitors has been hampered by the lack of patient selection marker. Using a high-throughput chemical screen, we discovered that cells deficient for the tumor suppressor ARID1A are highly sensitive to PLK1 inhibition. Interestingly this sensitivity was unrelated to canonical functions of PLK1 in mediating G2-M cell cycle transition. Instead, a whole-genome CRISPR screen revealed PLK1 inhibitor sensitivity in ARID1A deficient cells to be dependent on the mitochondrial translation machinery. We find that ARID1A knocked-out (KO) cells have an unusual mitochondrial phenotype with aberrant biogenesis, increased oxygen consumption/ expression of oxidative phosphorylation genes, but without increased ATP production. Using expansion microscopy and biochemical fractionation, we see that a subset of PLK1 localizes to the mitochondria in interphase cells. Inhibition of PLK1 in ARID1A KO cells further uncouples oxygen consumption from ATP production, with subsequent membrane depolarization and apoptosis. Knockdown of a key subunit of the mitochondrial ribosome reverses PLK1-inhibitor induced apoptosis in ARID1A deficient cells, confirming specificity of the phenotype. Together, these findings highlight a novel interphase role for PLK1 in maintaining mitochondrial fitness under metabolic stress, and a strategy for therapeutic use of PLK1 inhibitors. To translate these findings, we describe a quantitative microscopy assay for assessment of ARID1A protein loss, which could offer a novel patient selection strategy for the clinical development of PLK1 inhibitors in cancer. Statement of significanceCurrently, no predictive biomarkers have been identified for PLK1 inhibitors in cancer treatment. We show that ARID1A loss sensitizes cells to PLK1 inhibitors through a previously unrecognized vulnerability in mitochondrial oxygen metabolism.

cancer biology↗

Variation in predicted COVID-19 risk among lemurs and lorises

The novel coronavirus SARS-CoV-2, which in humans leads to the disease COVID-19, has caused global disruption and more than 1.5 million fatalities since it first emerged in late 2019. As we write, infection rates are currently at their highest point globally and are rising extremely rapidly in some areas due to more infectious variants. The primary viral target is the cellular receptor angiotensin-converting enzyme-2 (ACE2). Recent sequence analyses of the ACE2 gene predicts that many nonhuman primates are also likely to be highly susceptible to infection. However, the anticipated risk is not equal across the Order. Furthermore, some taxonomic groups show high ACE2 amino acid conservation, while others exhibit high variability at this locus. As an example of the latter, analyses of strepsirrhine primate ACE2 sequences to date indicate large variation among lemurs and lorises compared to other primate clades despite low sampling effort. Here, we report ACE2 gene and protein sequences for 71 individual strepsirrhines, spanning 51 species and 19 genera. Our study reinforces previous results and finds additional variability in other strepsirrhine species, and suggests several clades of lemurs have high potential susceptibility to SARS-CoV-2 infection. Troublingly, some species, including the rare and Endangered aye-aye (Daubentonia madagascariensis), as well as those in the genera Avahi and Propithecus, may be at high risk. Given that lemurs are endemic to Madagascar and among the primates at highest risk of extinction globally, further understanding of the potential threat of COVID-19 to their health should be a conservation priority. All feasible actions should be taken to limit their exposure to SARS-CoV-2.

genetics↗

Design of smart antibody mimetics with photosensitive switches

As two prominent examples of intracellular single-domain antibodies or antibody mimetics derived from synthetic protein scaffolds, monobodies and nanobodies are gaining wide applications in cell biology, structural biology, synthetic immunology, and theranostics. We introduce herein a generally-applicable method to engineer light-controllable monobodies and nanobodies, designated as moonbody and sunbody, respectively. These engineered antibody-like modular domains enable rapid and reversible antibody-antigen recognition by utilizing light. By paralleled insertion of two LOV2 modules into a single sunbody and the use of bivalent sunbodies, we substantially enhance the range of dynamic changes of photo-switchable sunbodies. Furthermore, we demonstrate the use of moonbodies or sunbodies to precisely control protein degradation, gene transcription, and base editing by harnessing the power of light.

synthetic biology↗

Single cell RNA-seq reveals immunosuppressive gastric stem-like cancer cells as a poor prognostic factor

ObjectiveGastric cancer (GC) tumors are highly heterogenous with different subpopulations of epithelial cells. We employed single cell RNA sequencing (scRNA-seq) to dissect the heterogeneity and identified subpopulations of cancer cells with stem-like properties. We further investigated their resistance to oxaliplatin chemotherapy and their contribution to gastric cancer outcome. DesignWe performed scRNA-seq on FACS sorted epithelial and immune cells from paired samples of GC tumors and normal adjacent tissues. We identified two epithelial subpopulations (STMN1+IQGAP3+ and STMN1+IQGAP3-) with stem-like properties. We characterized and compared them to known healthy gastric stem cell populations. We also cultivated GC derived organoids to study the chemoresistance of similarly marked populations. Lastly, we employed immunohistochemistry (IHC) staining to ascertain the predicted immunosuppressive interactions. ResultsThe STMN1+IQGAP3+ subpopulation showed a higher tumor mutation burden, upregulated proliferative pathways and transcriptomically resembled proliferative healthy gastric isthmus stem cells. The STMN1+IQGAP3- subpopulation were comparatively quiescent and transcriptomically resembled enteroendocrine cells. Both transcriptomic signatures were associated with worse mortality than other epithelial subpopulations with the quiescent being associated with the poorest patient survival. GC tissue derived organoids were dominated by STMN1+IQGAP3+ cells but the STMN1+IQGAP3- compartment was more resistant to chemotherapy. We also verified the likely suppression of CD8 T cell cytotoxicity by STMN1+IQGAP3+ cells through the NECTIN2/TIGIT interaction. ConclusionsCancer cells with stem-like characteristics are associated with poor survival through chemoresistance and immunosuppression. Reactivating the immune system through checkpoint blockade is an opportunity to eliminate these cells. What is already known on this topicMultiple gastric stem cell populations have been identified and linked to tumor initiation in rodent-based studies. However, none of them have been conclusively proven in human tumors. Isolating and characterizing tumor cells with stem-like properties will help shed light on their possible origin and possible mitigation strategies. What this study addsHere we identified two sets of stem-like gastric cancer cells that are associated with poorer patient prognosis. One set is highly proliferative and exhibits oxaliplatin susceptibility. It also engages in immunosuppressive interactions such as NECTIN2/TIGIT. The other set is quiescent and highly resistant to oxaliplatin. How this study might affect research, practice or policyThe transcriptome signatures of the identified stem-like cells can aid in patient prognosis and identify patients who can benefit from checkpoint blockade therapy to reactivate their immune response towards gastric cancer cells.

immunology↗

Engineering supramolecular organizing centers for optogenetic control of innate immune responses

The spatiotemporal organization of oligomeric protein complexes and translocons, such as the supramolecular organizing centers (SMOC) made of MyDDosome and MAVSome, are essential for transcriptional activation of host inflammatory responses and immune metabolisms. Light-inducible assembly of MyDDosome and MAVSome are presented herein to induce activation of nuclear factor-kB (NF-{kappa}B) and type-I interferons (IFNs). Engineering of SMOCs and the downstream transcription factor permits programmable and customized innate immune operations in a light-dependent manner. These synthetic molecular tools will likely enable optical and user-defined modulation of innate immunity at a high spatiotemporal resolution to facilitate mechanistic studies of distinct modes of innate immune activations and potential intervention of immune disorders and cancer.

synthetic biology↗

Curation of over 10,000 transcriptomic studies to enable data reuse

Vast amounts of transcriptomic data reside in public repositories, but effective reuse remains challenging. Issues include unstructured dataset metadata, inconsistent data processing and quality control, and inconsistent probe-gene mappings across microarray technologies. Thus, extensive curation and data reprocessing is necessary prior to any reuse. The Gemma bioinformatics system was created to help address these issues. Gemma consists of a database of curated transcriptomic datasets, analytical software, a web interface, and web services. Here we present an update on Gemmas holdings, data processing and analysis pipelines, our curation guidelines, and software features. As of June 2020, Gemma contains 10,811 manually curated datasets (primarily human, mouse, and rat), over 395,000 samples and hundreds of curated transcriptomic platforms (both microarray and RNA-sequencing). Dataset topics were represented with 10,215 distinct terms from 12 ontologies, for a total of 54,316 topic annotations (mean topics/dataset = 5.2). While Gemma has broad coverage of conditions and tissues, it captures a large majority of available brain-related datasets, accounting for 34% of its holdings. Users can access the curated data and differential expression analyses through the Gemma website, RESTful service, and an R package. Database URL: https://gemma.msl.ubc.ca/home.html

bioinformatics↗

"3G" Trial: An RNA Editing Signature for Guiding Gastric Cancer Chemotherapy

Background & AimsGastric cancer (GC) cases are often diagnosed at an advanced stage with poor prognosis. Platinum-based chemotherapy has been internationally accepted as first-line therapy for inoperable or metastatic GC. To achieve greater benefits, it is critical to select patients who are eligible for the treatment. Albeit gene expression profiling has been widely used as a genomic classifier to identify molecular subtypes of GC and stratify patients for different chemotherapy regimens, the prediction accuracy remains to be improved. More recently, adenosine-to-inosine (A-to-I) RNA editing has emerged as a new player contributing to GC development and progression, offering potential clinical utility for diagnosis and treatment. MethodsWe conducted a transcriptome-wide RNA editing analysis of a cohort of 104 patients with advanced GC and identified an RNA editing (GCRE) signature to guide GC chemotherapy, using a systematic computational approach followed by both in vitro validations and in silico validations in TCGA. ResultsWe found that RNA editing events alone stand as a prognostic and predictive biomarker in advanced GC. We developed a GCRE score based on the GCRE signature consisting of 50 editing sites associated with 29 genes and achieved a high accuracy (84%) of predicting patient response to chemotherapy. Of note, patients demonstrating higher editing levels of this panel of sites present a better overall response. Consistently, GC cell lines with higher editing levels showed higher chemosensitivity. Applying the GCRE score on TCGA dataset confirmed that responders had significantly higher levels of editing in advanced GC. ConclusionsOverall, the GCRE signature reliably stratifies patients with advanced GC and predicts response from chemotherapy. SignificanceDespite the increasing documentation of RNA editing and its functional regulation, the translational potential of RNA editome in cancer remains largely under-investigated. This study reports for the first time an RNA editing signature in advanced GC, to reliably stratify patients with advanced disease to predict response from chemotherapy independently of gene expression profiling and other genomic and epigenetic changes. For this purpose, a bioinformatics approach was used to develop a GCRE score based on a panel of 50 editing sites from 29 unique genes (GCRE signature), followed by an experimental evaluation of their clinical utility as predictive biomarker in GC cell lines and in silico validation in using RNA sequencing (RNA-Seq) datasets from TCGA. The applied methodology provides a robust means of an RNA editing signature to be investigated in patients with advanced GC. Overall, this study provides insights into the translation of RNA editing process into predictive clinical applications to direct chemotherapy against GC.

cancer biology↗

Heterogeneous expression of the SARS-Coronavirus-2 receptor ACE2 in the human respiratory tract

BackgroundZoonotically transmitted coronaviruses are responsible for three disease outbreaks since 2002, including the current COVID-19 pandemic, caused by SARS-CoV-2. Its efficient transmission and range of disease severity raise questions regarding the contributions of virus-receptor interactions. ACE2 is a host ectopeptidase and the receptor for SARS-CoV-2. Numerous reports describe ACE2 mRNA abundance and tissue distribution; however, mRNA abundance is not always representative of protein levels. Currently, there is limited data evaluating ACE2 protein and its correlation with other SARS-CoV-2 susceptibility factors. Materials and methodsWe systematically examined the human upper and lower respiratory tract using single-cell RNA sequencing and immunohistochemistry to determine receptor expression and evaluated its association with risk factors for severe COVID-19. FindingsOur results reveal that ACE2 protein is highest within regions of the sinonasal cavity and pulmonary alveoli, sites of presumptive viral transmission and severe disease development, respectively. In the lung parenchyma, ACE2 protein was found on the apical surface of a small subset of alveolar type II cells and colocalized with TMPRSS2, a cofactor for SARS-CoV2 entry. ACE2 protein was not increased by pulmonary risk factors for severe COVID-19. Additionally, ACE2 protein was not reduced in children, a demographic with a lower incidence of severe COVID-19. InterpretationThese results offer new insights into ACE2 protein localization in the human respiratory tract and its relationship with susceptibility factors to COVID-19. Research in contextO_ST_ABSEvidence before this studyC_ST_ABSPrevious studies of ACE2 mRNA transcript abundance in the human respiratory tract have suggested a possible association between ACE2 expression and age, sex, and the presence of comorbidities. However, these studies have provided conflicting results, as well as a lack of protein validation. Previous ACE2 protein studies have been limited by a paucity of lung tissue samples and reports that have produced contradictory results. Added value of this studyUsing a combination of single-cell RNA sequencing and immunohistochemistry, we describe ACE2 expression in the human respiratory tract. Staining protocols were optimized and validated to show consistent apical localization and avoid non-specific staining. We show ACE2 protein is found in subsets of airway cells and is highest within regions of the sinonasal cavity and pulmonary alveoli, sites of presumptive viral transmission and severe disease development for COVID-19, respectively. We show age, sex, and comorbidities do not increase ACE2 protein expression in the human respiratory tract. Implications of all the available evidenceACE2 protein abundance does not correlate with risk factors for severe clinical outcomes, but in some cases showed an inversed relationship. Features driving COVID-19 susceptibility and severity are complex, our data suggests factors other than ACE2 protein abundance as important determinants of clinical outcomes.

cell biology↗

Epigenomic Diversity of Cortical Projection Neurons in the Mouse Brain

Neuronal cell types are classically defined by their molecular properties, anatomy, and functions. While recent advances in single-cell genomics have led to high-resolution molecular characterization of cell type diversity in the brain, neuronal cell types are often studied out of the context of their anatomical properties. To better understand the relationship between molecular and anatomical features defining cortical neurons, we combined retrograde labeling with single-nucleus DNA methylation sequencing to link epigenomic properties of cell types to neuronal projections. We examined 11,827 single neocortical neurons from 63 cortico-cortical (CC) and cortico-subcortical long-distance projections. Our results revealed unique epigenetic signatures of projection neurons that correspond to their laminar and regional location and projection patterns. Based on their epigenomes, intra-telencephalic (IT) cells projecting to different cortical targets could be further distinguished, and some layer 5 neurons projecting to extra-telencephalic targets (L5-ET) formed separate subclusters that aligned with their axonal projections. Such separation varied between cortical areas, suggesting area-specific differences in L5-ET subtypes, which were further validated by anatomical studies. Interestingly, a population of CC projection neurons clustered with L5-ET rather than IT neurons, suggesting a population of L5-ET cortical neurons projecting to both targets (L5-ET+CC). We verified the existence of these neurons by labeling the axon terminals of CC projection neurons and observed clear labeling in ET targets including thalamus, superior colliculus, and pons. These findings highlight the power of single-cell epigenomic approaches to connect the molecular properties of neurons with their anatomical and projection properties.

neuroscience↗

Family History Assessment Significantly Enhances Delivery of Precision Medicine in the Genomics Era

BackgroundFamily history has traditionally been an essential part of clinical care to assess health risks. However, declining sequencing costs have precipitated a shift towards genomics-first approaches in population screening programs, with less emphasis on family history assessment. We evaluated the utility of family history for genomic sequencing selection. MethodsWe analysed whole genome sequences of 1750 healthy research participants, with and without preselection based on standardised family history collection, screening 95 cancer genes. ResultsThe frequency of likely pathogenic/ pathogenic (LP/P) variants in 884 participants with no family history available (FH not available group) (2%) versus 866 participants with family history available (FH available group) (3.1%) was not significant (p=0.158). However, within the FH available group, amongst 73 participants with an increased family history cancer risk (increased FH risk), 1 in 7 participants carried a LP/P variant inferring a six-fold increase compared with 1 in 47 participants assessed at average family history cancer risk (average FH risk) and a seven-fold increase compared to the FH not available group. The enrichment was further pronounced (up to 18-fold) when assessing the 25 cancer genes in the ACMG 59-gene panel. Furthermore, 63 participants had an increased family history cancer risk in absence of an apparent LP/P variant. ConclusionOur findings show that systematic family history collection remains critical for health risk assessment, providing important actionable data and augmenting the yield from genomic data. Family history also highlights the potential impact of additional hereditary, environmental and behavioural influences not reflected by genomic sequencing.

genomics↗

Bactericidal efficiency and mechanism of specifically targeted antimicrobial peptides optimized based on structural and functional relationships

In contrast to traditional broad-spectrum antibiotics, it is difficult for bacteria to develop resistance to most specifically targeted antimicrobial peptides (STAMPs), moreover, they can maintain a normal ecological balance and provide long-term protection for the body. However, therapeutic applications of STAMPS are hindered by their weak activity, and imperfect specificity as well as lack of knowledge to understand their structure-activity relationships. To further investigate the effects of different parameters on the biological activities of STAMPs, a peptide sequence, WKKIWKDPGIKKWIK, was truncated, extended, and provided with an increased charge and altered amphipathicity. In addition, a novel template modification method was introduced, in which a phage-displayed peptide that recognized and bound to E. coli cells was attached at the end of the sequence. Compared with the traditional template modification method, peptide 11, which contained a phage-displayed peptide at the C-terminus, exhibited superior narrow-spectrum antibacterial activity against E. coli compared to that of parental peptide 2, and the activity and specificity of 11 were increased by 5.0 and 2.4 times, respectively. Additionally, 11 showed low cell toxicity and relatively desirable salt, serum, acid and alkaline stability. In this study, 11 specifically killed E. coli by causing cytoplasmic membrane rupture and cytosol leakage. In summary, these findings are useful for improving the activity and specificity of STAMPs and show that peptide 11 is better able to combat the growing threat of E. coli infections.

biochemistry↗

Digital phenotyping by consumer wearables identifies sleep-associated markers of cardiovascular disease risk and biological aging

Despite growing adoption of consumer wearables, the potential for sleep metrics from these devices to contribute to sleep-related biomedical research remains largely uncharacterized. Here we analyze sleep tracking data, along with questionnaire responses and multi-modal phenotypic data, generated from 482 normal volunteers. First, we provide a detailed comparison of wearable-derived and self-reported sleep metrics, particularly total sleep time (TST) and sleep efficiency (SE). We then identified demographic, socioeconomic and lifestyle factors associated with wearable-derived sleep duration. We also analyzed our multi-modal phenotypic data and showed that wearable-derived TST and SE are associated with various cardiovascular disease risk markers, whereas self-reported measures were not. Using whole-genome sequencing data, we estimated leukocyte telomere length and showed that volunteers with insufficient sleep also exhibit premature telomere attrition. Our study highlights the potential for sleep metrics generated by consumer wearables to provide novel insights into data generated from population cohort studies.

epidemiology↗

Large-scale whole-genome sequencing of three diverse Asian populations in Singapore

Asian populations are currently underrepresented in human genetics research. Here we present whole-genome sequencing data of 4,810 Singaporeans from three diverse ethnic groups: 2,780 Chinese, 903 Malays, and 1,127 Indians. Despite a medium depth of 13.7x, we achieved essentially perfect (>99.8%) sensitivity and accuracy for detecting common variants and good sensitivity (>89%) for detecting extremely rare variants with <0.1% allele frequency. We found 89.2 million single-nucleotide polymorphisms (SNPs) and 9.1 million small insertions and deletions (INDELs), more than half of which have not been cataloged in dbSNP. In particular, we found 126 common deleterious mutations (MAF>0.01) that were absent in the existing public databases, highlighting the importance of local population reference for genetic diagnosis. We describe fine-scale genetic structure of Singapore populations and their relationship to worldwide populations from the 1000 Genomes Project. In addition to revealing noticeable amounts of admixture among three Singapore populations and a Malay-related novel ancestry component that has not been captured by the 1000 Genomes Project, our analysis also identified some fine-scale features of genetic structure consistent with two waves of prehistoric migration from south China to Southeast Asia. Finally, we demonstrate that our data can substantially improve genotype imputation not only for Singapore populations, but also for populations across Asia and Oceania. These results highlight the genetic diversity in Singapore and the potential impacts of our data as a resource to empower human genetics discovery in a broad geographic region.

genetics↗

Whole-Genome Genomics Correlates of Response To Anti-PD1 Therapy in Relapsed/Refractory Natural Killer/T Cell Lymphoma

AbstractThis study aims to identify recurrent genetic alterations in relapsed or refractory (RR) natural-killer/T-cell lymphoma (NKTL) patients who have achieved complete response (CR) with programmed cell death 1 (PD-1) blockade therapy. Seven of the eleven patients treated with pembrolizumab achieved CR while the remaining four had progressive disease (PD). Using whole genome sequencing (WGS), we found recurrent clonal structural rearrangements (SR) of the PD-L1 gene in four of the seven (57%) CR patients pretreated tumors. These PD-L1 SRs consist of inter-chromosomal translocations, tandem duplication and micro-inversion that disrupted the suppressive function of PD-L1 3UTR. Interestingly, recurrent JAK3-activating (p.A573V) mutations were also validated in two CR patients tumors that did not harbor the PD-L1 SR. Importantly, these mutations were absent in the four PD cases. With immunohistochemistry (IHC), PD-L1 positivity could not discriminate patients who archived CR (range: 6%-100%) from patients who had PD (range: 35%-90%). PD-1 blockade with pembrolizumab is a potent strategy for RR NKTL patients and genomic screening could potentially accompany PD-L1 IHC positivity to better select patients for anti-PD-1 therapy.

genomics↗

Using machine learning to guide targeted and locally-tailored empiric antibiotic prescribing in a children’s hospital in Cambodia

BackgroundEarly and appropriate empiric antibiotic treatment of patients suspected of having sepsis is associated with reduced mortality. The increasing prevalence of antimicrobial resistance risks eroding the benefits of such empiric therapy. This problem is particularly severe for children in developing country settings. We hypothesized that by applying machine learning approaches to readily collected patient data, it would be possible to obtain actionable and patient-specific predictions for antibiotic-susceptibility. If sufficient discriminatory power can be achieved, such predictions could lead to substantial improvements in the chances of choosing an appropriate antibiotic for empiric therapy, while minimizing the risk of increased selection for resistance due to use of antibiotics usually held in reserve.\n\nMethods and FindingsWe analyzed blood culture data collected from a 100-bed childrens hospital in North-West Cambodia between February 2013 and January 2016. Clinical, demographic and living condition information for each child was captured with 35 independent variables. Using these variables, we used a suite of machine learning algorithms to predict Gram stains and whether bacterial pathogens could be treated with standard empiric antibiotic therapies: i) ampicillin and gentamicin; ii) ceftriaxone; iii) at least one of the above.\n\n243 cases of bloodstream infection were available for analysis. We used 195 (80%) to train the algorithms, and 48 (20%) for evaluation. We found that the random forest method had the best predictive performance overall as assessed by the area under the receiver operating characteristic curve (AUC), though support vector machine with radial kernel had similar performance for predicting Gram stain and ceftriaxone susceptibility. Predictive performance of logistic regression, simple and boosted decision trees and k-nearest neighbors were poor in comparison. The random forest method gave an AUC of 0.91 (95%CI 0.81-1.00) for predicting susceptibility to ceftriaxone, 0.75 (0.60-0.90) for susceptibility to ampicillin and gentamicin, 0.76 (0.59-0.93) for susceptibility to neither, and 0.69 (0.53-0.85) for Gram stain result. The most important variables for predicting susceptibility were time from admission to blood culture, patient age, hospital versus community-acquired infection, and age-adjusted weight score.\n\nConclusionsApplying machine learning algorithms to patient data that are readily available even in resource-limited hospital settings can provide highly informative predictions on susceptibilities of pathogens to guide appropriate empiric antibiotic therapy. Used as a decision support tool, such approaches have the potential to lead to better targeting of empiric therapy, improve patient outcomes and reduce the burden of antimicrobial resistance.\n\nAuthor summaryO_LSTWhy was this study done?C_LSTO_LIEarly and appropriate antibiotic treatment of patients with life-threatening bacterial infections is thought to reduce the risk of mortality.\nC_LIO_LIIn hospitals that have a microbiology laboratory, it takes 3-4 days to get results which indicate which antibiotics are likely to be effective; before this information is available antibiotics have to be prescribed empirically i.e. without knowledge of the causative organism.\nC_LIO_LIIncreasing resistance to antibiotics amongst bacteria makes finding an appropriate antibiotic to use empirically difficult; this problem is particularly severe for children in developing country settings.\nC_LIO_LIIf we could predict which antibiotics were likely to be effective at the time of starting antibiotic therapy, we might be able to improve patient outcomes and reduce resistance.\nC_LI\n\nO_LSTWhat Did the Researchers Do and Find?C_LSTO_LIWe evaluated the ability of a number of different algorithms (i.e. sets of step-by-step instructions) to predict susceptibility to commonly-used antibiotics using routinely available patient data from a childrens hospital in Cambodia.\nC_LIO_LIWe found that an algorithm called random forests enabled surprisingly accurate predictions, particularly for predicting whether the infection was likely to be treatable with ceftriaxone, the most commonly used empiric antibiotic at the study hospital.\nC_LIO_LIUsing this approach it would be possible to correctly predict when a different antibiotic would be needed for empiric treatment over 80% of the time, while recommending a different antibiotic when ceftriaxone would suffice less than 20% of the time.\nC_LI\n\nO_LSTWhat Do These Findings Mean?C_LSTO_LIUsing readily available patient information, sophisticated algorithms can enable good predictions of whether antibiotics are likely to be effective several days before laboratory tests are available.\nC_LIO_LIAlgorithms would need to be trained with local hospital data, but our study shows that even with relatively limited data from a small hospital, good predictions can be obtained.\nC_LIO_LIUsed as part of a decision support system such algorithms could help choose appropriate antibiotics for empiric therapy; this would be expected to translate into better patient outcomes and may help to reduce resistance.\nC_LIO_LISuch as a decision support system would have very low costs and be easy to implement in low- and middle-income countries.\nC_LI

epidemiology↗

Naa10p promotes metastasis by stabilizing matrix metalloproteinase-2 protein in human osteosarcomas

N--Acetyltransferase 10 protein (Naa10p) mediates N-terminal acetylation of nascent proteins. Oncogenic or tumor suppressive roles of Naa10p were reported in cancers. Here, we report an oncogenic role of Naa10p in promoting metastasis of osteosarcomas. Higher NAA10 transcripts were observed in metastatic osteosarcoma tissues compared to non-metastatic tissues and were also correlated with a worse prognosis of patients. Knockdown and overexpression of Naa10p in osteosarcoma cells respectively led to decreased and increased cell migratory/invasive abilities. Re-expression of Naa10p, but not an enzymatically inactive mutant, relieved suppression of the invasive ability in vitro and metastasis in vivo imposed by Naa10p-knockdown. According to protease array screening, we identified that matrix metalloproteinase (MMP)-2 was responsible for the Naa10p-induced invasive phenotype. Naa10p was directly associated with MMP-2 protein through its acetyltransferase domain and maintained MMP-2 protein stability via NatA complex activity. MMP-2 expression levels were also significantly correlated with Naa10p levels in osteosarcoma tissues. These results reveal a novel function of Naa10p in the regulation of cell invasiveness by preventing MMP-2 protein degradation that is crucial during osteosarcoma metastasis.

cancer biology↗

Transmission patterns of hyper-endemic multi-drug resistant Klebsiella pneumoniae in a Cambodian neonatal unit: a longitudinal study with whole genome sequencing

BackgroundKlebsiella pneumoniae is an important and increasing cause of life-threatening disease in hospitalised neonates. Third generation cephalosporin resistance (3GC-R) is frequently a marker of multi-drug resistance, and can complicate management of infections. 3GC-R K. pneumoniae is hyper-endemic in many developing country settings, but its epidemiology is poorly understood and prospective studies of endemic transmission are lacking. We aimed to determine the transmission dynamics of 3GC-R K. pneumoniae in a newly opened neonatal unit (NU) in Cambodia.\n\nMethodsWe performed a prospective longitudinal study between September and November 2013. Rectal swabs from 37 consented patients were collected upon NU admission and every three days thereafter. Morphologically different colonies from swabs growing cefpodoxime-resistant K. pneumoniae were selected for whole-genome sequencing (WGS).\n\nResults32/37 (86%) patients screened positive for 3GC-R K. pneumoniae and 93 colonies from 119 swabs were sequenced. Isolates were resistant to a median of six (range 3-9) antimicrobials. WGS revealed high diversity; pairwise distances between isolates from the same patient were either 0-1 SNV or >1,000 SNVs; 19/32 colonized patients harboured K. pneumoniae colonies differing by >1000 SNVs. Diverse lineages accounted for 18 probable importations to the NU and nine probable transmission clusters involving 19/37 (51%) of screened patients. Median cluster size was 5 patients (range 3-9).\n\nConclusionsThe epidemiology of 3GC-R K. pneumoniae was characterised by multiple introductions and a dense network of cross-infection, with half of screened neonates part of a transmission cluster. Efforts to reduce the 3GC-R K. pneumoniae disease burden should consider targeting both processes.

epidemiology↗