Assistant Professor, Institute for Immunity, Transplantation and Infection (2014 - Present)
Longitudinal Gene Expression Profiling in Adults After Traumatic Injury
The purpose of this study is to examine the immune response to traumatic injury and subsequent infections in critically ill adults. Traumatic injuries lead to severe dysregulation of the immune system, and predispose to severe infections. Diagnosing these infections in a timely manner is paramount in reducing morbidity and mortality, but diagnosis is made difficult by the inflammatory response to trauma. The main purpose of the study is to prospectively test the diagnostic power of the expression of an 11-gene set which the investigators recently published (Sweeney et al., Sci Transl Med, 2015). Since the timing of an acquired infection cannot be determined a priori, this study is designed to be a longitudinal examination of a cohort of traumatically injured adults. The investigators will draw blood at regular intervals, as well as at day of diagnosis of infection for any patient that are diagnosed with an infection. The investigators will then assay the blood for gene expression levels post hoc, and correlate the molecular profiles with clinical information to establish a prospective estimate of diagnostic power.
Stanford is currently not accepting patients for this trial. For more information, please contact Timothy E Sweeney, MD, PhD, 720-201-6689.
- Essential Methods in Computational and Systems Immunology
IMMUNOL 207 (Spr)
- Seminars in Computational and Systems Immunology
IMMUNOL 310 (Sum)
Independent Studies (15)
- Biomedical Informatics Teaching Methods
BIOMEDIN 290 (Aut, Win, Spr)
- Directed Reading and Research
BIOMEDIN 299 (Aut, Win, Spr, Sum)
- Directed Reading in Immunology
IMMUNOL 299 (Win, Spr)
- Directed Reading in Medicine
MED 299 (Win, Spr)
- Early Clinical Experience in Immunology
IMMUNOL 280 (Win, Spr)
- Early Clinical Experience in Medicine
MED 280 (Win, Spr)
- Graduate Research
CBIO 399 (Aut, Win, Spr, Sum)
- Graduate Research
IMMUNOL 399 (Aut, Win, Spr, Sum)
- Graduate Research
MED 399 (Win, Spr, Sum)
- Medical Scholars Research
BIOMEDIN 370 (Aut, Win, Spr, Sum)
- Medical Scholars Research
MED 370 (Win, Spr)
- Ph.D. Research
CME 400 (Aut, Win, Spr)
- Teaching in Immunology
IMMUNOL 290 (Win, Spr, Sum)
- Undergraduate Research
IMMUNOL 199 (Win, Spr)
- Undergraduate Research
MED 199 (Win, Spr)
- Biomedical Informatics Teaching Methods
Prior Year Courses
- Essential Methods in Computational and Systems Immunology
IMMUNOL 207 (Spr)
- Seminars in Computational and Systems Immunology
IMMUNOL 310 (Sum)
- Essential Methods in Computational and Systems Immunology
IMMUNOL 207 (Spr)
- Seminars in Computational and Systems Immunology
IMMUNOL 310 (Sum)
- Essential Methods in Computational and Systems Immunology
IMMUNOL 207 (Spr)
- Seminars in Computational and Systems Immunology
IMMUNOL 310 (Sum)
- Essential Methods in Computational and Systems Immunology
Doctoral Dissertation Reader (AC)
Postdoctoral Faculty Sponsor
Sanjana Gupta, Lara Jones
Doctoral Dissertation Advisor (AC)
Ananthakrishnan Ganesan, Hacer Umay Geyikci, Larry Kalesinskas, Yiran Liu, Madeleine Scott
Doctoral Dissertation Co-Advisor (AC)
Kalani Ratnasiri, Jiaying Toh
Postdoctoral Research Mentor
Sanjana Gupta, Lara Jones, Krishna Martinez-Singh
Graduate and Fellowship Programs
Biomedical Informatics (Phd Program)
Increasing reproducibility, robustness, and generalizability of biomarker selection from meta-analysis using Bayesian methodology.
PLoS computational biology
2022; 18 (6): e1010260
A major limitation of gene expression biomarker studies is that they are not reproducible as they simply do not generalize to larger, real-world, heterogeneous populations. Frequentist multi-cohort gene expression meta-analysis has been frequently used as a solution to this problem to identify biomarkers that are truly differentially expressed. However, the frequentist meta-analysis framework has its limitations-it needs at least 4-5 datasets with hundreds of samples, is prone to confounding from outliers and relies on multiple-hypothesis corrected p-values. To address these shortcomings, we have created a Bayesian meta-analysis framework for the analysis of gene expression data. Using real-world data from three different diseases, we show that the Bayesian method is more robust to outliers, creates more informative estimates of between-study heterogeneity, reduces the number of false positive and false negative biomarkers and selects more generalizable biomarkers with less data. We have compared the Bayesian framework to a previously published frequentist framework and have developed a publicly available R package for use.
View details for DOI 10.1371/journal.pcbi.1010260
View details for PubMedID 35759523
Host protease activity classifies pneumonia etiology.
Proceedings of the National Academy of Sciences of the United States of America
2022; 119 (25): e2121778119
Community-acquired pneumonia (CAP) has been brought to the forefront of global health priorities due to the COVID-19 pandemic. However, classification of viral versus bacterial pneumonia etiology remains a significant clinical challenge. To this end, we have engineered a panel of activity-based nanosensors that detect the dysregulated activity of pulmonary host proteases implicated in the response to pneumonia-causing pathogens and produce a urinary readout of disease. The nanosensor targets were selected based on a human protease transcriptomic signature for pneumonia etiology generated from 33 unique publicly available study cohorts. Five mouse models of bacterial or viral CAP were developed to assess the ability of the nanosensors to produce etiology-specific urinary signatures. Machine learning algorithms were used to train diagnostic classifiers that could distinguish infected mice from healthy controls and differentiate those with bacterial versus viral pneumonia with high accuracy. This proof-of-concept diagnostic approach demonstrates a way to distinguish pneumonia etiology based solely on the host proteolytic response to infection.
View details for DOI 10.1073/pnas.2121778119
View details for PubMedID 35696579
An 8-gene machine learning model improves clinical prediction of severe dengue progression.
2022; 14 (1): 33
BACKGROUND: Each year 3-6 million people develop life-threatening severe dengue (SD). Clinical warning signs for SD manifest late in the disease course and are nonspecific, leading to missed cases and excess hospital burden. Better SD prognostics are urgently needed.METHODS: We integrated 11 public datasets profiling the blood transcriptome of 365 dengue patients of all ages and from seven countries, encompassing biological, clinical, and technical heterogeneity. We performed an iterative multi-cohort analysis to identify differentially expressed genes (DEGs) between non-severe patients and SD progressors. Using only these DEGs, we trained an XGBoost machine learning model on public data to predict progression to SD. All model parameters were "locked" prior to validation in an independent, prospectively enrolled cohort of 377 dengue patients in Colombia. We measured expression of the DEGs in whole blood samples collected upon presentation, prior to SD progression. We then compared the accuracy of the locked XGBoost model and clinical warning signs in predicting SD.RESULTS: We identified eight SD-associated DEGs in the public datasets and built an 8-gene XGBoost model that accurately predicted SD progression in the independent validation cohort with 86.4% (95% CI 68.2-100) sensitivity and 79.7% (95% CI 75.5-83.9) specificity. Given the 5.8% proportion of SD cases in this cohort, the 8-gene model had a positive and negative predictive value (PPV and NPV) of 20.9% (95% CI 16.7-25.6) and 99.0% (95% CI 97.7-100.0), respectively. Compared to clinical warning signs at presentation, which had 77.3% (95% CI 58.3-94.1) sensitivity and 39.7% (95% CI 34.7-44.9) specificity, the 8-gene model led to an 80% reduction in the number needed to predict (NNP) from 25.4 to 5.0. Importantly, the 8-gene model accurately predicted subsequent SD in the first three days post-fever onset and up to three days prior to SD progression.CONCLUSIONS: The 8-gene XGBoost model, trained on heterogeneous public datasets, accurately predicted progression to SD in a large, independent, prospective cohort, including during the early febrile stage when SD prediction remains clinically difficult. The model has potential to be translated to a point-of-care prognostic assay to reduce dengue morbidity and mortality without overwhelming limited healthcare resources.
View details for DOI 10.1186/s13073-022-01034-w
View details for PubMedID 35346346
Increases in ambient air pollutants during pregnancy are linked to increases in methylation of IL4, IL10, and IFNgamma.
2022; 14 (1): 40
BACKGROUND: Ambient air pollutant (AAP) exposure is associated with adverse pregnancy outcomes, such as preeclampsia, preterm labor, and low birth weight. Previous studies have shown methylation of immune genes associate with exposure to air pollutants in pregnant women, but the cell-mediated response in the context of typical pregnancy cell alterations has not been investigated. Pregnancy causes attenuation in cell-mediated immunity with alterations in the Th1/Th2/Th17/Treg environment, contributing to maternal susceptibility. We recruited women (n=186) who were 20weeks pregnant from Fresno, CA, an area with chronically elevated AAP levels. Associations of average pollution concentration estimates for 1week, 1month, 3months, and 6months prior to blood draw were associated with Th cell subset (Th1, Th2, Th17, and Treg) percentages and methylation of CpG sites (IL4, IL10, IFNgamma, and FoxP3). Linear regression models were adjusted for weight, age, season, race, and asthma, using a Q value as the false-discovery-rate-adjusted p-value across all genes.RESULTS: Short-term and mid-term AAP exposures to fine particulate matter (PM2.5), nitrogen dioxide (NO2) carbon monoxide (CO), and polycyclic aromatic hydrocarbons (PAH456) were associated with percentages of immune cells. A decrease in Th1 cell percentage was negatively associated with PM2.5 (1 mo/3 mo: Q<0.05), NO2 (1 mo/3 mo/6 mo: Q<0.05), and PAH456 (1week/1 mo/3 mo: Q<0.05). Th2 cell percentages were negatively associated with PM2.5 (1week/1 mo/3 mo/6 mo: Q<0.06), and NO2 (1week/1 mo/3 mo/6 mo: Q<0.06). Th17 cell percentage was negatively associated with NO2 (3 mo/6 mo: Q<0.01), CO (1week/1 mo: Q<0.1), PM2.5 (3 mo/6 mo: Q<0.05), and PAH456 (1 mo/3 mo/6 mo: Q<0.08). Methylation of the IL10 gene was positively associated with CO (1week/1 mo/3 mo: Q<0.01), NO2 (1 mo/3 mo/6 mo: Q<0.08), PAH456 (1week/1 mo/3 mo: Q<0.01), and PM2.5 (3 mo: Q=0.06) while IL4 gene methylation was positively associated with concentrations of CO (1week/1 mo/3 mo/6 mo: Q<0.09). Also, IFNgamma gene methylation was positively associated with CO (1week/1 mo/3 mo: Q<0.05) and PAH456 (1week/1 mo/3 mo: Q<0.06).CONCLUSION: Exposure to several AAPs was negatively associated with T-helper subsets involved in pro-inflammatory and anti-inflammatory responses during pregnancy. Methylation of IL4, IL10, and IFNgamma genes with pollution exposure confirms previous research. These results offer insights into the detrimental effects of air pollution during pregnancy, the demand for more epigenetic studies, and mitigation strategies to decrease pollution exposure during pregnancy.
View details for DOI 10.1186/s13148-022-01254-2
View details for PubMedID 35287715
Mechanisms of innate and adaptive immunity to the Pfizer-BioNTech BNT162b2 vaccine.
Despite the success of the BNT162b2 mRNA vaccine, the immunological mechanisms that underlie its efficacy are poorly understood. Here we analyzed the innate and adaptive responses to BNT162b2 in mice, and show that immunization stimulated potent antibody and antigen-specific T cell responses, as well as strikingly enhanced innate responses after secondary immunization, which was concurrent with enhanced serum interferon (IFN)-gamma levels 1d following secondary immunization. Notably, we found that natural killer cells and CD8+ T cells in the draining lymph nodes are the major producers of this circulating IFN-gamma. Analysis of knockout mice revealed that induction of antibody and T cell responses to BNT162b2 was not dependent on signaling via Toll-like receptors 2, 3, 4, 5 and 7 nor inflammasome activation, nor the necroptosis or pyroptosis cell death pathways. Rather, the CD8+ T cell response induced by BNT162b2 was dependent on type I interferon-dependent MDA5 signaling. These results provide insights into the molecular mechanisms by which the BNT162b2 vaccine stimulates immune responses.
View details for DOI 10.1038/s41590-022-01163-9
View details for PubMedID 35288714
- Author Correction: The immunoregulatory landscape of human tuberculosis granulomas. Nature immunology 2022
Serum proteome analysis of systemic JIA and related lung disease identifies distinct inflammatory programs and biomarkers.
Arthritis & rheumatology (Hoboken, N.J.)
OBJECTIVES: Recent observations in systemic Juvenile Idiopathic Arthritis (sJIA) suggest an increasing incidence of high-mortality interstitial lung disease (sJIA-LD) often characterized by a variant of pulmonary alveolar proteinosis (PAP). Co-occurrence of macrophage activation syndrome (MAS) and PAP in sJIA suggested a shared pathology, but sJIA-LD patients also commonly experience features of drug reaction such as atypical rashes and eosinophilia. We sought to investigate immunopathology and identify biomarkers in sJIA, MAS, and sJIA-LD.METHODS: We used SOMAscan to measure >1300 analytes in sera from healthy controls and patients with sJIA, MAS, sJIA-LD and other related diseases. We verified selected findings by ELISA and lung immunostaining. Because the proteome of a sample may reflect multiple states (sJIA, MAS, sJIA-LD), we used regression modeling to identify subsets of altered proteins associated with each state. We tested key findings in a validation cohort.RESULTS: Proteome alterations in active sJIA and MAS overlapped substantially, including known sJIA biomarkers like SAA and S100A9, and novel elevations of heat shock proteins and glycolytic enzymes. IL-18 was elevated in all sJIA groups, particularly MAS and sJIA-LD. We also identified an MAS-independent sJIA-LD signature notable for elevated ICAM5, MMP7, and allergic/eosinophilic chemokines, which have been previously associated with lung damage. Immunohistochemistry localized ICAM5 and MMP7 in sJIA-LD lung. ICAM5's ability to distinguish sJIA-LD from sJIA/MAS was independently validated.CONCLUSION: Serum proteins support an sJIA-to-MAS continuum, help distinguish sJIA, sJIA/MAS, and sJIA-LD and suggest etiologic hypotheses. Select biomarkers, such as ICAM5, could aid in early detection and management of sJIA-LD.
View details for DOI 10.1002/art.42099
View details for PubMedID 35189047
A GMR-based assay for quantification of the human response to influenza.
Biosensors & bioelectronics
2022; 205: 114086
Detecting and quantifying the host transcriptional response to influenza virus infection can serve as a real-time diagnostic tool for clinical management. We have employed the multiplexing capabilities of GMR sensors to develop a novel assay based on the influenza metasignature (IMS), which can classify influenza infection based on transcript levels. We show that the assay can reliably detect ten IMS transcripts and distinguish subjects with naturally acquired influenza infection from those with other symptomatic viral infections (AUC 0.93, 95% CI: 0.82-1.00). Separately, we validated that the gene IFI27, not included in the IMS panel, has very high single-biomarker accuracy (AUC 0.95, 95% CI: 0.90-0.99) in stratifying patients with influenza. We demonstrate that a portable GMR biosensor can be used as a tool to diagnose influenza infection by measuring the host response, simultaneously highlighting the power of immune system metrics and advancing the field of gene expression-based diagnostics.
View details for DOI 10.1016/j.bios.2022.114086
View details for PubMedID 35192997
A robust gene expression signature for NASH in liver expression data.
2022; 12 (1): 2571
Non-Alcoholic Fatty Liver Disease (NAFLD) is a progressive liver disease that affects up to 30% of worldwide population, of which up to 25% progress to Non-Alcoholic SteatoHepatitis (NASH), a severe form of the disease that involves inflammation and predisposes the patient to liver cirrhosis. Despite its epidemic proportions, there is no reliable diagnostics that generalizes to global patient population for distinguishing NASH from NAFLD. We performed a comprehensive multicohort analysis of publicly available transcriptome data of liver biopsies from Healthy Controls (HC), NAFLD and NASH patients. Altogether we analyzed 812 samples from 12 different datasets across 7 countries, encompassing real world patient heterogeneity. We used 7 datasets for discovery and 5 datasets were held-out for independent validation. Altogether we identified 130 genes significantly differentially expressed in NASH versus a mixed group of NAFLD and HC. We show that our signature is not driven by one particular group (NAFLD or HC) and reflects true biological signal. Using a forward search we were able to downselect to a parsimonious set of 19 mRNA signature with mean AUROC of 0.98 in discovery and 0.79 in independent validation. Methods for consistent diagnosis of NASH relative to NAFLD are urgently needed. We showed that gene expression data combined with advanced statistical methodology holds the potential to serve basis for development of such diagnostic tests for the unmet clinical need.
View details for DOI 10.1038/s41598-022-06512-0
View details for PubMedID 35173224
Cytokine signatures differentiate systemic sclerosis patients at high versus low risk for pulmonary arterial hypertension.
Arthritis research & therapy
2022; 24 (1): 39
BACKGROUND: Pulmonary arterial hypertension (PAH) affects approximately 10% of patients with systemic sclerosis (SSc) and is a leading cause of death. We sought to identify serum cytokine signatures that risk stratify SSc patients for this potentially fatal complication.METHODS: Subjects at high risk for PAH and with incident PAH based on right heart catheterization (RHC) were enrolled in the multi-center prospective registry, Pulmonary Hypertension Assessment and Recognition of Outcomes in Scleroderma (PHAROS). Low-risk SSc patients were enrolled at Stanford and had normal pulmonary function test and echocardiogram parameters. Serum was available from 71 high-risk patients, 81 incident PAH patients, 10 low-risk patients, and 20 healthy controls (HC). Custom 14- and 65-plex arrays were used for cytokine analysis. Cytokine expression was compared between patient groups by principal component analysis and Tukey's test result. A multiple hypotheses corrected p value <0.05 was considered significant.RESULTS: Exploratory analysis using principal components showed unique clustering for each patient group. There was a significant difference in cytokine expression in at least one group comparison for every cytokine. Overall, there was very little difference in cytokine expression comparing high-risk and PAH patient groups; however, these groups had substantially different cytokine profiles compared to low-risk patients and HC.CONCLUSION: These data suggest that cytokine profiles can distinguish SSc patients who are at high-risk for or have PAH from SSc patients who may be at lower risk for PAH and HC. However, high-risk and PAH patients had very similar cytokine profiles, suggesting that these patients are on a disease continuum.
View details for DOI 10.1186/s13075-022-02734-9
View details for PubMedID 35139913
The Single Cell Transcriptomic and Epigenomic Map of the Innate Immune Response to Vaccination in Lymph Nodes
MOSBY-ELSEVIER. 2022: AB316
View details for Web of Science ID 000778999300708
A molecular atlas of innate immunity to adjuvanted and live attenuated vaccines, in mice.
1800; 13 (1): 549
Adjuvants hold great potential in enhancing vaccine efficacy, making the understanding and improving of adjuvants critical goals in vaccinology. The TLR7/8 agonist, 3M-052, induces long-lived humoral immunity in non-human primates and is currently being evaluated in human clinical trials. However, the innate mechanisms of 3M-052 have not been fully characterized. Here, we perform flow cytometry, single cell RNA-seq and ATAC-seq to profile the kinetics, transcriptomics and epigenomics of innate immune cells in murine draining lymph nodes following 3M-052-Alum/Ovalbumin immunization. We find that 3M-052-Alum/OVA induces a robust antiviral and interferon gene program, similar to the yellow fever vaccine, which is known to confer long-lasting protection. Activation of myeloid cells in dLNs persists through day 28 and single cell analysis reveals putative TF-gene regulatory programs in distinct myeloid cells and heterogeneity of monocytes. This study provides a comprehensive characterization of the transcriptomics and epigenomics of innate populations in the dLNs after vaccination.
View details for DOI 10.1038/s41467-022-28197-9
View details for PubMedID 35087093
A 6-mRNA host response classifier in whole blood predicts outcomes in COVID-19 and other acute viral infections.
1800; 12 (1): 889
Predicting the severity of COVID-19 remains an unmet medical need. Our objective was to develop a blood-based host-gene-expression classifier for the severity of viral infections and validate it in independent data, including COVID-19. We developed a logistic regression-based classifier for the severity of viral infections and validated it in multiple viral infection settings including COVID-19. We used training data (N=705) from 21 retrospective transcriptomic clinical studies of influenza and other viral illnesses looking at a preselected panel of host immune response messenger RNAs. We selected 6 host RNAs and trained logistic regression classifier with a cross-validation area under curve of 0.90 for predicting 30-day mortality in viral illnesses. Next, in 1417 samples across 21 independent retrospective cohorts the locked 6-RNA classifier had an area under curve of 0.94 for discriminating patients with severe vs. non-severe infection. Next, in independent cohorts of prospectively (N=97) and retrospectively (N=100) enrolled patients with confirmed COVID-19, the classifier had an area under curve of 0.89 and 0.87, respectively, for identifying patients with severe respiratory failure or 30-day mortality. Finally, we developed a loop-mediated isothermal gene expression assay for the 6-messenger-RNA panel to facilitate implementation as a rapid assay. With further study, the classifier could assist in the risk assessment of COVID-19 and other acute viral infections patients to determine severity and level of care, thereby improving patient management and reducing healthcare burden.
View details for DOI 10.1038/s41598-021-04509-9
View details for PubMedID 35042868
Disease characteristics and serological responses in patients with differing severity of COVID-19 infection: A longitudinal cohort study in Dhaka, Bangladesh.
PLoS neglected tropical diseases
2022; 16 (1): e0010102
COVID-19 caused by SARS-CoV-2 ranges from asymptomatic to severe disease and can cause fatal and devastating outcome in many cases. In this study, we have compared the clinical, biochemical and immunological parameters across the different disease spectrum of COVID-19 in Bangladeshi patients.This longitudinal study was conducted in two COVID-19 hospitals and also around the community in Dhaka city in Bangladesh between November 2020 to March 2021. A total of 100 patients with COVID-19 infection were enrolled and classified into asymptomatic, mild, moderate and severe cases (n = 25/group). In addition, thirty age and sex matched healthy participants were enrolled and 21 were analyzed as controls based on exclusion criteria. After enrollment (study day1), follow-up visits were conducted on day 7, 14 and 28 for the cases. Older age, male gender and co-morbid conditions were the risk factors for severe COVID-19 disease. Those with moderate and severe cases of infection had low lymphocyte counts, high neutrophil counts along with a higher neutrophil-lymphocyte ratio (NLR) at enrollment; this decreased to normal range within 42 days after the onset of symptom. At enrollment, D-dimer, CRP and ferritin levels were elevated among moderate and severe cases. The mild, moderate, and severe cases were seropositive for IgG antibody by day 14 after enrollment. Moderate and severe cases showed significantly higher IgM and IgG levels of antibodies to SARS-CoV-2 compared to mild and asymptomatic cases.We report on the clinical, biochemical, and hematological parameters associated with the different severity of COVID-19 infection. We also show different profile of antibody response against SARS-CoV-2 in relation to disease severity, especially in those with moderate and severe disease manifestations compared to the mild and asymptomatic infection.
View details for DOI 10.1371/journal.pntd.0010102
View details for PubMedID 34982773
The immunoregulatory landscape of human tuberculosis granulomas.
Tuberculosis (TB) in humans is characterized by formation of immune-rich granulomas in infected tissues, the architecture and composition of which are thought to affect disease outcome. However, our understanding of the spatial relationships that control human granulomas is limited. Here, we used multiplexed ion beam imaging by time of flight (MIBI-TOF) to image 37 proteins in tissues from patients with active TB. We constructed a comprehensive atlas that maps 19 cell subsets across 8 spatial microenvironments. This atlas shows an IFN-γ-depleted microenvironment enriched for TGF-β, regulatory T cells and IDO1+ PD-L1+ myeloid cells. In a further transcriptomic meta-analysis of peripheral blood from patients with TB, immunoregulatory trends mirror those identified by granuloma imaging. Notably, PD-L1 expression is associated with progression to active TB and treatment response. These data indicate that in TB granulomas, there are local spatially coordinated immunoregulatory programs with systemic manifestations that define active TB.
View details for DOI 10.1038/s41590-021-01121-x
View details for PubMedID 35058616
Comparison of the Transcriptomic Signatures in Pediatric and Adult CML.
1800; 13 (24)
Children with chronic myeloid leukemia (CML) tend to present with higher white blood counts and larger spleens than adults with CML, suggesting that the biology of pediatric and adult CML may differ. To investigate whether pediatric and adult CML have unique molecular characteristics, we studied the transcriptomic signature of pediatric and adult CML CD34+ cells and healthy pediatric and adult CD34+ control cells. Using high-throughput RNA sequencing, we found 567 genes (207 up- and 360 downregulated) differentially expressed in pediatric CML CD34+ cells compared to pediatric healthy CD34+ cells. Directly comparing pediatric and adult CML CD34+ cells, 398 genes (258 up- and 140 downregulated), including many in the Rho pathway, were differentially expressed in pediatric CML CD34+ cells. Using RT-qPCR to verify differentially expressed genes, VAV2 and ARHGAP27 were significantly upregulated in adult CML CD34+ cells compared to pediatric CML CD34+ cells. NCF1, CYBB, and S100A8 were upregulated in adult CML CD34+ cells but not in pediatric CML CD34+ cells, compared to healthy controls. In contrast, DLC1 was significantly upregulated in pediatric CML CD34+ cells but not in adult CML CD34+ cells, compared to healthy controls. These results demonstrate unique molecular characteristics of pediatric CML, such as dysregulation of the Rho pathway, which may contribute to clinical differences between pediatric and adult patients.
View details for DOI 10.3390/cancers13246263
View details for PubMedID 34944883
Identification of a LAMC2-regulated network featuring targetable effectors for dual therapies in pancreatic cancer.
AMER ASSOC CANCER RESEARCH. 2021: 59-60
View details for Web of Science ID 000720117400101
A Multi-mRNA Prognostic Signature for Anti-TNFalpha Therapy Response in Patients with Inflammatory Bowel Disease.
Diagnostics (Basel, Switzerland)
2021; 11 (10)
BACKGROUND: Anti-TNF-alpha (anti-TNFalpha) therapies have transformed the care and management of inflammatory bowel disease (IBD). However, they are expensive and ineffective in greater than 50% of patients, and they increase the risk of infections, liver issues, arthritis, and lymphoma. With 1.6 million Americans suffering from IBD and global prevalence on the rise, there is a critical unmet need in the use of anti-TNFalpha therapies: a test for the likelihood of therapy response. Here, as a proof-of-concept, we present a multi-mRNA signature for predicting response to anti-TNFalpha treatment to improve the efficacy and cost-to-benefit ratio of these biologics.METHODS: We surveyed public data repositories and curated four transcriptomic datasets (n = 136) from colonic and ileal mucosal biopsies of IBD patients (pretreatment) who were subjected to anti-TNFalpha therapy and subsequently adjudicated for response. We applied a multicohort analysis with a leave-one-study-out (LOSO) approach, MetaIntegrator, to identify significant differentially expressed (DE) genes between responders and non-responders and then used a greedy forward search to identify a parsimonious gene signature. We then calculated an anti-TNFalpha response (ATR) score based on this parsimonious gene signature to predict responder status and assessed discriminatory performance via an area-under-receiver operating-characteristic curve (AUROC).RESULTS: We identified 324 significant DE genes between responders and non-responders. The greedy forward search yielded seven genes that robustly distinguish anti-TNFalpha responders from non-responders, with an AUROC of 0.88 (95% CI: 0.70-1). The Youden index yielded a mean sensitivity of 91%, mean specificity of 76%, and mean accuracy of 86%.CONCLUSIONS: Our findings suggest that there is a robust transcriptomic signature for predicting anti-TNFalpha response in mucosal biopsies from IBD patients prior to treatment initiation. This seven-gene signature should be further investigated for its potential to be translated into a predictive test for clinical use.
View details for DOI 10.3390/diagnostics11101902
View details for PubMedID 34679598
Functional Consequences of Memory Inflation after Solid Organ Transplantation.
Journal of immunology (Baltimore, Md. : 1950)
CMV is a major infectious complication following solid organ transplantation. Reactivation of CMV leads to memory inflation, a process in which CD8 T cells expand over time. Memory inflation is associated with specific changes in T cell function, including increased oligoclonality, decreased cytokine production, and terminal differentiation. To address whether memory inflation during the first year after transplantation in human subjects alters T cell differentiation and function, we employed single-cell-matched TCRalphabeta and targeted gene expression sequencing. Expanded T cell clones exhibited a terminally differentiated, immunosenescent, and polyfunctional phenotype whereas rare clones were less differentiated. Clonal expansion occurring between pre- and 3 mo posttransplant was accompanied by enhancement of polyfunctionality. In contrast, polyfunctionality and differentiation state were largely maintained between 3 and 12 mo posttransplant. Highly expanded clones had a higher degree of polyfunctionality than rare clones. Thus, CMV-responsive CD8 T cells differentiated during the pre- to posttransplant period then maintained their differentiation state and functional capacity despite posttransplant clonal expansion.
View details for DOI 10.4049/jimmunol.2100405
View details for PubMedID 34551963
Evolution of Cytomegalovirus-Responsive T Cell Clonality following Solid Organ Transplantation.
Journal of immunology (Baltimore, Md. : 1950)
CMV infection is a significant complication after solid organ transplantation. We used single cell TCR alphabeta sequencing to determine how memory inflation impacts clonality and diversity of the CMV-responsive CD8 and CD4 T cell repertoire in the first year after transplantation in human subjects. We observed CD8 T cell inflation but no changes in clonal diversity, indicating homeostatic stability in clones. In contrast, the CD4 repertoire was diverse and stable over time, with no evidence of CMV-responsive CD4 T cell expansion. We identified shared CDR3 TCR motifs among patients but no public CMV-specific TCRs. Temporal changes in clonality in response to transplantation and in the absence of detectable viral reactivation suggest changes in the repertoire immediately after transplantation followed by an expansion with stable clonal competition that may mediate protection.
View details for DOI 10.4049/jimmunol.2100404
View details for PubMedID 34551964
Computational drug repositioning of atorvastatin for ulcerative colitis.
Journal of the American Medical Informatics Association : JAMIA
OBJECTIVE: Ulcerative colitis (UC) is a chronic inflammatory disorder with limited effective therapeutic options for long-term treatment and disease maintenance. We hypothesized that a multi-cohort analysis of independent cohorts representing real-world heterogeneity of UC would identify a robust transcriptomic signature to improve identification of FDA-approved drugs that can be repurposed to treat patients with UC.MATERIALS AND METHODS: We performed a multi-cohort analysis of 272 colon biopsy transcriptome samples across 11 publicly available datasets to identify a robust UC disease gene signature. We compared the gene signature to in vitro transcriptomic profiles induced by 781 FDA-approved drugs to identify potential drug targets. We used a retrospective cohort study design modeled after a target trial to evaluate the protective effect of predicted drugs on colectomy risk in patients with UC from the Stanford Research Repository (STARR) database and Optum Clinformatics DataMart.RESULTS: Atorvastatin treatment had the highest inverse-correlation with the UC gene signature among non-oncolytic FDA-approved therapies. In both STARR (n = 827) and Optum (n = 7821), atorvastatin intake was significantly associated with a decreased risk of colectomy, a marker of treatment-refractory disease, compared to patients prescribed a comparator drug (STARR: HR = 0.47, P = .03; Optum: HR = 0.66, P = .03), irrespective of age and length of atorvastatin treatment.DISCUSSION & CONCLUSION: These findings suggest that atorvastatin may serve as a novel therapeutic option for ameliorating disease in patients with UC. Importantly, we provide a systematic framework for integrating publicly available heterogeneous molecular data with clinical data at a large scale to repurpose existing FDA-approved drugs for a wide range of human diseases.
View details for DOI 10.1093/jamia/ocab165
View details for PubMedID 34529084
Macrophage-derived IL-6 trans-signaling as a novel target in the pathogenesis of bronchopulmonary dysplasia.
The European respiratory journal
RATIONALE: Premature infants exposed to oxygen are at risk for bronchopulmonary dysplasia (BPD), which is characterised by lung growth arrest. Inflammation is important, but the mechanisms remain elusive. Here, we investigated inflammatory pathways and therapeutic targets in severe clinical and experimental BPD.METHODS AND RESULTS: First, transcriptomic analysis with in-silico cellular deconvolution identified a lung-intrinsic M1-like-driven cytokine pattern in newborn mice after hyperoxia. These findings were confirmed by gene expression of macrophage-regulating chemokines (Ccl2, Ccl7, Cxcl5) and markers (Il6, Il17A, Mmp12). Second, hyperoxia-activated IL-6/STAT3 signaling was measured in vivo and related to loss of alveolar epithelial type II cells (ATII) as well as increased mesenchymal marker. Il6 null mice exhibited preserved ATII survival, reduced myofibroblasts and improved elastic fiber assembly, thus enabling lung growth and protecting lung function. Pharmacological inhibition of global IL-6 signaling and IL-6 trans-signaling promoted alveolarisation and ATII survival after hyperoxia. Third, hyperoxia triggered M1-like polarisation, possibly via Klf4; hyperoxia-conditioned medium of macrophages and IL-6 impaired ATII proliferation. Finally, clinical data demonstrate elevated macrophage-related plasma cytokines as potential biomarkers that identify infants receiving oxygen at increased risk of developing BPD. Moreover, macrophage-derived IL6 and active STAT3 were related to loss of epithelial cells in BPD lungs.CONCLUSION: We present a novel IL-6-mediated mechanism by which hyperoxia activates macrophages in immature lungs, impairs ATII homeostasis, and disrupts elastic fiber formation, thereby inhibiting lung growth. The data provide evidence that IL-6 trans-signaling could offer an innovative pharmacological target to enable lung growth in severe neonatal chronic lung disease.
View details for DOI 10.1183/13993003.02248-2020
View details for PubMedID 34446466
The single-cell epigenomic and transcriptional landscape of immunity to influenza vaccination in humans
WILEY. 2021: 31
View details for Web of Science ID 000753366400071
A novel blood-based assay for treatment monitoring of tuberculosis.
BMC research notes
2021; 14 (1): 247
OBJECTIVES: A novel 3-gene host transcriptional signature (GBP5, DUSP3 and KLF2) has been validated for tuberculosis (TB) treatment monitoring using laboratory-based RNA sequencing platforms. The signature was recently translated by Cepheid into a prototype cartridge-based test that can be run on the GeneXpert instrument. In this study, we prospectively evaluated the change in the expression of the cartridge-based 3-gene signature following treatment initiation among pulmonary TB patients who were microbiologically cured at the end of treatment.RESULTS: The 3-gene signature expression level (TB score) changed significantly over time with respect to baseline among 31 pulmonary TB patients. The greatest increase in TB score occurred within the first month of treatment (median fold-increase in TB score: 1.08 [IQR 0.54-1.52]) and plateaued after 4months of treatment (median TB score: 1.97 [IQR: 1.03-2.33]). The rapid and substantial increase of the TB score in the first month of treatment holds promise for the early identification of patients that respond to TB treatment. The plateau in TB score at 4months may indicate early clearance of disease and could direct treatment to be shortened. These hypotheses need to be further explored with larger prospective treatment monitoring studies.
View details for DOI 10.1186/s13104-021-05663-z
View details for PubMedID 34193258
Prospective validation of an 11-gene mRNA host response score for mortality risk stratification in the intensive care unit.
2021; 11 (1): 13062
Several clinical calculators predict intensive care unit (ICU) mortality, however these are cumbersome and often require 24h of data to calculate. Retrospective studies have demonstrated the utility of whole blood transcriptomic analysis in predicting mortality. In this study, we tested prospective validation of an 11-gene messenger RNA (mRNA) score in an ICU population. Whole blood mRNA from 70 subjects in the Stanford ICU Biobank with samples collected within 24h of Emergency Department presentation were used to calculate an 11-gene mRNA score. We found that the 11-gene score was highly associated with 60-day mortality, with an area under the receiver operating characteristic curve of 0.68 in all patients, 0.77 in shock patients, and 0.98 in patients whose primary determinant of prognosis was acute illness. Subjects with the highest quartile of mRNA scores were more likely to die in hospital (40% vs 7%, p<0.01) and within 60days (40% vs 15%, p=0.06). The 11-gene score improved prognostication with a categorical Net Reclassification Improvement index of 0.37 (p=0.03) and an Integrated Discrimination Improvement index of 0.07 (p=0.02) when combined with Simplified Acute Physiology Score 3 or Acute Physiology and Chronic Health Evaluation II score. The test performed poorly in the 95 independent samples collected>24h after emergency department presentation. Tests will target a 30-min turnaround time, allowing for rapid results early in admission. Moving forward, this test may provide valuable real-time prognostic information to improve triage decisions and allow for enrichment of clinical trials.
View details for DOI 10.1038/s41598-021-91201-7
View details for PubMedID 34158514
The single-cell epigenomic and transcriptional landscape of immunity to influenza vaccination.
Emerging evidence indicates a fundamental role for the epigenome in immunity. Here, we mapped the epigenomic and transcriptional landscape of immunity to influenza vaccination in humans at the single-cell level. Vaccination against seasonal influenza induced persistently diminished H3K27ac in monocytes and myeloid dendritic cells (mDCs), which was associated with impaired cytokine responses to Toll-like receptor stimulation. Single-cell ATAC-seq analysis revealed an epigenomically distinct subcluster of monocytes with reduced chromatin accessibility at AP-1-targeted loci after vaccination. Similar effects were observed in response to vaccination with the AS03-adjuvanted H5N1 pandemic influenza vaccine. However, this vaccine also stimulated persistently increased chromatin accessibility at interferon response factor (IRF) loci in monocytes and mDCs. This was associated with elevated expression of antiviral genes and heightened resistance to the unrelated Zika and Dengue viruses. These results demonstrate that vaccination stimulates persistent epigenomic remodeling of the innate immune system and reveal AS03's potential as an epigenetic adjuvant.
View details for DOI 10.1016/j.cell.2021.05.039
View details for PubMedID 34174187
Repression of CTSG, ELANE and PRTN3-mediated histone H3 proteolytic cleavage promotes monocyte-to-macrophage differentiation.
Chromatin undergoes extensive reprogramming during immune cell differentiation. Here we report the repression of controlled histone H3 amino terminus proteolytic cleavage (H3DeltaN) during monocyte-to-macrophage development. This abundant histone mark in human peripheral blood monocytes is catalyzed by neutrophil serine proteases (NSPs) cathepsin G, neutrophil elastase and proteinase 3. NSPs are repressed as monocytes mature into macrophages. Integrative epigenomic analysis reveals widespread H3DeltaN distribution across the genome in a monocytic cell line and primary monocytes, which becomes largely undetectable in fully differentiated macrophages. H3DeltaN is enriched at permissive chromatin and actively transcribed genes. Simultaneous NSP depletion in monocytic cells results in H3DeltaN loss and further increase in chromatin accessibility, which likely primes the chromatin for gene expression reprogramming. Importantly, H3DeltaN is reduced in monocytes from patients with systemic juvenile idiopathic arthritis, an autoinflammatory disease with prominent macrophage involvement. Overall, we uncover an epigenetic mechanism that primes the chromatin to facilitate macrophage development.
View details for DOI 10.1038/s41590-021-00928-y
View details for PubMedID 34017121
- Diversity in immunogenomics: the value and the challenge. Nature methods 2021
iPSC-endothelial cell phenotypic drug screening and in silico analyses identify tyrphostin-AG1296 for pulmonary arterial hypertension.
Science translational medicine
2021; 13 (592)
Pulmonary arterial hypertension (PAH) is a progressive disorder leading to occlusive vascular remodeling. Current PAH therapies improve quality of life but do not reverse structural abnormalities in the pulmonary vasculature. Here, we used high-throughput drug screening combined with in silico analyses of existing transcriptomic datasets to identify a promising lead compound to reverse PAH. Induced pluripotent stem cell-derived endothelial cells generated from six patients with PAH were exposed to 4500 compounds and assayed for improved cell survival after serum withdrawal using a chemiluminescent caspase assay. Subsequent validation of caspase activity and improved angiogenesis combined with data analyses using the Gene Expression Omnibus and Library of Integrated Network-Based Cellular Signatures databases revealed that the lead compound AG1296 was positively associated with an anti-PAH gene signature. AG1296 increased abundance of bone morphogenetic protein receptors, downstream signaling, and gene expression and suppressed PAH smooth muscle cell proliferation. AG1296 induced regression of PA neointimal lesions in lung organ culture and PA occlusive changes in the Sugen/hypoxia rat model and reduced right ventricular systolic pressure. Moreover, AG1296 improved vascular function and BMPR2 signaling and showed better correlation with the anti-PAH gene signature than other tyrosine kinase inhibitors. Specifically, AG1296 up-regulated small mothers against decapentaplegic (SMAD) 1/5 coactivators, cAMP response element-binding protein 3 (CREB3), and CREB5: CREB3 induced inhibitor of DNA binding 1 and downstream genes that improved vascular function. Thus, drug discovery for PAH can be accelerated by combining phenotypic screening with in silico analyses of publicly available datasets.
View details for DOI 10.1126/scitranslmed.aba6480
View details for PubMedID 33952674
Gene Expression-Based Diagnosis of Infections in Critically Ill Patients-Prospective Validation of the SepsisMetaScore in a Longitudinal Severe Trauma Cohort.
Critical care medicine
OBJECTIVES: Early diagnosis of infections is pivotal in critically ill patients. Innovative gene expression-based approaches promise to deliver precise, fast, and clinically practicable diagnostic tools to bedside. This study aimed to validate the SepsisMetaScore, an 11-gene signature previously reported by our study group, in a representative longitudinal cohort of trauma patients.DESIGN: Prospective observational cohort study.SETTING: Surgical ICUs of the University Medical Center Goettingen, Germany.PATIENTS: Critically ill patients with severe traumatic injuries.INTERVENTIONS: None.MEASUREMENTS AND MAIN RESULTS: Paired box gene (PAXgene) RNA blood tubes were drawn at predefined time points over the course of disease. The performance of the SepsisMetaScore was tested using targeted polymerase chain reaction and compared with Procalcitonin using area under the receiver operating characteristics analyses. The SepsisMetaScore showed significant differences between infected and noninfected patients (n = 52). It was able to accurately discriminate infectious from noninfectious acute inflammation with an area under the receiver operating characteristics of 0.92 (95% CI, 0.85-0.99) and significantly outperformed Procalcitonin (area under the receiver operating characteristics curve = 0.53; 95% CI, 0.42-0.64) early in the course of infection (p = 0.014).CONCLUSIONS: We demonstrated the clinical utility for diagnosis of infections with higher accuracy using the SepsisMetaScore compared with Procalcitonin in a prospective cohort of severe trauma patients. Future studies should assess whether the SepsisMetaScore may substantially improve clinical practice by accurate differentiation of infections from sterile inflammation and identification of patients at risk for sepsis. Our results support further investigation of the SepsisMetaScore for the development of tailored precision treatment of critically ill patients.
View details for DOI 10.1097/CCM.0000000000005027
View details for PubMedID 33883455
Multisystem inflammatory syndrome in children: a microcosm of challenges and opportunities for translational bioinformatics in pediatric research.
Current opinion in pediatrics
PURPOSE OF REVIEW: Despite significant progress in our understanding and clinical management of multisystem inflammatory syndrome in children (MIS-C), significant challenges remain. Here, we review recently published studies on the clinical diagnosis, risk stratification, and treatment of MIS-C, highlighting key gaps in research progress that are a microcosm for challenges in translational pediatric research. We then discuss potential solutions in the realm of translational bioinformatics.RECENT FINDINGS: Current case definitions are inconsistent and do not capture the underlying pathophysiology of MIS-C, which remains poorly understood. Although overall mortality is low, some patients rapidly decompensate, and a test to identify those at risk for severe outcomes remains an unmet need. Treatment consists of various combinations of immunoglobulins, corticosteroids, and biologics, based on extrapolated data and expert opinion, while the benefits remain unclear as we await the completion of clinical trials.SUMMARY: The small size and heterogeneity of the pediatric population contribute to unmet needs because of financial and logistical constraints of the current research infrastructure focused on eliminating most sources of heterogeneity, leading to ungeneralizable results. Data sharing and meta-analysis of gene expression shows promise to accelerate progress in the field of MIS-C as well as other childhood diseases beyond the current pandemic.
View details for DOI 10.1097/MOP.0000000000001012
View details for PubMedID 33871421
Multi-cohort analysis of host immune response identifies conserved protective and detrimental modules associated with severity across viruses.
Viral infections induce a conserved host response distinct from bacterial infections. We hypothesized that the conserved response is associated with disease severity and is distinct between patients with different outcomes. To test this, we integrated 4,780 blood transcriptome profiles from patients aged 0 to 90 years infected with one of 16 viruses, including SARS-CoV-2, Ebola, chikungunya, and influenza, across 34 cohorts from 18 countries, and single-cell RNA sequencing profiles of 702,970 immune cells from 289 samples across three cohorts. Severe viral infection was associated with increased hematopoiesis, myelopoiesis, and myeloid-derived suppressor cells. We identified protective and detrimental gene modules that defined distinct trajectories associated with mild versus severe outcomes. The interferon response was decoupled from the protective host response in patients with severe outcomes. These findings were consistent, irrespective of age and virus, and provide insights to accelerate the development of diagnostics and host-directed therapies to improve global pandemic preparedness.
View details for DOI 10.1016/j.immuni.2021.03.002
View details for PubMedID 33765435
Transcriptomic similarities and differences in host response between SARS-CoV-2 and other viral infections.
2021; 24 (1): 101947
The pandemic 2019 novel coronavirus disease (COVID-19) shares certain clinical characteristics with other acute viral infections. We studied the whole-blood transcriptomic host response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) using RNAseq from 24 healthy controls and 62 prospectively enrolled patients with COVID-19. We then compared these data to non-COVID-19 viral infections, curated from 23 independent studies profiling 1,855 blood samples covering six viruses (influenza, respiratory syncytial virus (RSV), human rhinovirus (HRV), severe acute respiratory syndrome coronavirus 1 (SARS-CoV-1), Ebola, dengue). We show gene expression changes in COVID-19 versus non-COVID-19 viral infections are highly correlated (r= 0.74, p< 0.001). However, we also found 416 genes specific to COVID-19. Inspection of top genes revealed dynamic immune evasion and counter host responses specific to COVID-19. Statistical deconvolution of cell proportions maps many cell type proportions concordantly shifting. Discordantly increased in COVID-19 were CD56bright natural killer cells and M2 macrophages. The concordant and discordant responses mapped out here provide a window to explore the pathophysiology of the host response to SARS-CoV-2.
View details for DOI 10.1016/j.isci.2020.101947
View details for PubMedID 33437935
SIMON: Open-Source Knowledge Discovery Platform.
Patterns (New York, N.Y.)
2021; 2 (1): 100178
Data analysis and knowledge discovery has become more and more important in biology and medicine with the increasing complexity of biological datasets, but the necessarily sophisticated programming skills and in-depth understanding of algorithms needed pose barriers to most biologists and clinicians to perform such research. We have developed a modular open-source software, SIMON, to facilitate the application of 180+ state-of-the-art machine-learning algorithms to high-dimensional biomedical data. With an easy-to-use graphical user interface, standardized pipelines, and automated approach for machine learning and other statistical analysis methods, SIMON helps to identify optimal algorithms and provides a resource that empowers non-technical and technical researchers to identify crucial patterns in biomedical data.
View details for DOI 10.1016/j.patter.2020.100178
View details for PubMedID 33511368
A multi-scale integrated analysis identifies KRT8 as a pan-cancer early biomarker.
Pacific Symposium on Biocomputing. Pacific Symposium on Biocomputing
2021; 26: 297–308
An early biomarker would transform our ability to screen and treat patients with cancer. The large amount of multi-scale molecular data in public repositories from various cancers provide unprecedented opportunities to find such a biomarker. However, despite identification of numerous molecular biomarkers using these public data, fewer than 1% have proven robust enough to translate into clinical practice. One of the most important factors affecting the successful translation to clinical practice is lack of real-world patient population heterogeneity in the discovery process. Almost all biomarker studies analyze only a single cohort of patients with the same cancer using a single modality. Recent studies in other diseases have demonstrated the advantage of leveraging biological and technical heterogeneity across multiple independent cohorts to identify robust disease biomarkers. Here we analyzed 17149 samples from patients with one of 23 cancers that were profiled using either DNA methylation, bulk and single-cell gene expression, or protein expression in tumor and serum. First, we analyzed DNA methylation profiles of 9855 samples across 23 cancers from The Cancer Genome Atlas (TCGA). We then examined the gene expression profile of the most significantly hypomethylated gene, KRT8, in 6781 samples from 57 independent microarray datasets from NCBI GEO. KRT8 was significantly over-expressed across cancers except colon cancer (summary effect size=1.05; p < 0.0001). Further, single-cell RNAseq analysis of 7447 single cells from lung tumors showed that genes that significantly correlated with KRT8 (p < 0.05) were involved in p53-related pathways. Immunohistochemistry in tumor biopsies from 294 patients with lung cancer showed that high protein expression of KRT8 is a prognostic marker of poor survival (HR = 1.73, p = 0.01). Finally, detectable KRT8 in serum as measured by ELISA distinguished patients with pancreatic cancer from healthy controls with an AUROC=0.94. In summary, our analysis demonstrates that KRT8 is (1) differentially expressed in several cancers across all molecular modalities and (2) may be useful as a biomarker to identify patients that should be further tested for cancer.
View details for PubMedID 33691026
Multicohort Analysis Identifies Monocyte Gene Signatures to Accurately Monitor Subset-Specific Changes in Human Diseases.
Frontiers in immunology
2021; 12: 659255
Monocytes are crucial regulators of inflammation, and are characterized by three distinct subsets in humans, of which classical and non-classical are the most abundant. Different subsets carry out different functions and have been previously associated with multiple inflammatory conditions. Dissecting the contribution of different monocyte subsets to disease is currently limited by samples and cohorts, often resulting in underpowered studies and poor reproducibility. Publicly available transcriptome profiles provide an alternative source of data characterized by high statistical power and real-world heterogeneity. However, most transcriptome datasets profile bulk blood or tissue samples, requiring the use of in silico approaches to quantify changes in cell levels. Here, we integrated 853 publicly available microarray expression profiles of sorted human monocyte subsets from 45 independent studies to identify robust and parsimonious gene expression signatures, consisting of 10 genes specific to each subset. These signatures maintain their accuracy regardless of disease state in an independent cohort profiled by RNA-sequencing and are specific to their respective subset when compared to other immune cells from both myeloid and lymphoid lineages profiled across 6160 transcriptome profiles. Consequently, we show that these signatures can be used to quantify changes in monocyte subsets levels in expression profiles from patients in clinical trials. Finally, we show that proteins encoded by our signature genes can be used in cytometry-based assays to specifically sort monocyte subsets. Our results demonstrate the robustness, versatility, and utility of our computational approach and provide a framework for the discovery of new cellular markers.
View details for DOI 10.3389/fimmu.2021.659255
View details for PubMedID 34054824
A multi-scale integrated analysis identifies KRT8 as a pan-cancer early biomarker
WORLD SCIENTIFIC PUBL CO PTE LTD. 2021: 297-308
View details for Web of Science ID 000759784400028
Systems vaccinology of the BNT162b2 mRNA vaccine in humans.
The emergency use authorization of two mRNA vaccines in less than a year since the emergence of SARS-CoV-2 represents a landmark in vaccinology1,2. Yet, how mRNA vaccines stimulate the immune system to elicit protective immune responses is unknown. Here we used a systems vaccinology approach to comprehensively profile the innate and adaptive immune responses of 56 healthy volunteers vaccinated with the Pfizer-BioNTech mRNA vaccine. Vaccination resulted in robust production of neutralizing antibodies (nAbs) against the parent Wuhan strain and, to a lesser extent, the B.1.351 strain, and significant increases in antigen-specific polyfunctional CD4 and CD8 T cells after the second dose. Booster vaccination stimulated a strikingly enhanced innate immune response compared to primary vaccination, evidenced by a greater: (i) frequency of CD14+CD16+ inflammatory monocytes; (ii) concentration of plasma IFN-g; (iii) transcriptional signature of innate antiviral immunity. Consistent with these observations, single-cell transcriptomics analysis demonstrated a ~100-fold increase in the frequency of a myeloid cell cluster, enriched in interferon-response transcription factors (TFs) and reduced in AP-1 TFs, following secondary immunization. Finally, we identified distinct innate pathways associated with CD8 T cell and nAb responses, and show that a monocyte-related signature correlates with the nAb response against the B.1.351 variant strain. Collectively, these data provide insights into immune responses induced by mRNA vaccination and demonstrate its capacity to prime the innate immune system to mount a more potent response following booster immunization.
View details for DOI 10.1038/s41586-021-03791-x
View details for PubMedID 34252919
- Formulating a Gene Signature for Diagnosis of Autoimmune and Infectious Diseases ASSOC COMPUTING MACHINERY. 2021
Signatures of immune dysfunction in HIV and HCV infection share features with chronic inflammation in aging and persist after viral reduction or elimination.
Proceedings of the National Academy of Sciences of the United States of America
2021; 118 (14)
Chronic inflammation is thought to be a major cause of morbidity and mortality in aging, but whether similar mechanisms underlie dysfunction in infection-associated chronic inflammation is unclear. Here, we profiled the immune proteome, and cellular composition and signaling states in a cohort of aging individuals versus a set of HIV patients on long-term antiretroviral therapy therapy or hepatitis C virus (HCV) patients before and after sofosbuvir treatment. We found shared alterations in aging-associated and infection-associated chronic inflammation including T cell memory inflation, up-regulation of intracellular signaling pathways of inflammation, and diminished sensitivity to cytokines in lymphocytes and myeloid cells. In the HIV cohort, these dysregulations were evident despite viral suppression for over 10 y. Viral clearance in the HCV cohort partially restored cellular sensitivity to interferon-α, but many immune system alterations persisted for at least 1 y posttreatment. Our findings indicate that in the HIV and HCV cohorts, a broad remodeling and degradation of the immune system can persist for a year or more, even after the removal or drastic reduction of the pathogen load and that this shares some features of chronic inflammation in aging.
View details for DOI 10.1073/pnas.2022928118
View details for PubMedID 33811141
Blood-based host biomarker diagnostics in active case finding for pulmonary tuberculosis:
EClinicalMedicine, published by The Lancet
View details for DOI 10.1016/j.eclinm.2021.100776
Diagnostic accuracy study of a novel blood-based assay for identification of TB in people living with HIV.
Journal of clinical microbiology
A non-sputum triage test to rule out TB disease is a WHO high-priority diagnostic and a combinatory score based on a 3-gene host-signature has shown promise in discriminating TB from other illnesses. We evaluated the accuracy of an early-prototype cartridge-assay ("Xpert MTB Host Response", or Xpert-MTB-HR-Prototype) of this 3-gene signature on bio-banked blood-samples from PLHIV against a comprehensive microbiological reference standard (CMRS) and against Xpert MTB/RIF on first sputum alone. We depict results based on performance targets set by WHO in comparison with a laboratory-based CRP assay. Of 201 patients included, 67 were culture-positive for Mycobacterium tuberculosis AUC for the Xpert-MTB-HR-Prototype was 0·89 (CI 0·83-0·94) against the CMRS and 0·94 (CI 0·89-0·98) against Xpert MTB/RIF. Considering Xpert-MTB-HR-Prototype as a triage test (at nearest upper value of sensitivity to 90%), specificity was 55·8% (CI 47·2-64·1) compared to the CMRS and 85·9% (CI 79·3-90·7) compared to Xpert MTB/RIF as confirmatory tests. Considering Xpert-MTB-HR-Prototype as a stand-alone diagnostic test, at a specificity near 95%, the test achieved a sensitivity of 65·7% (CI 53·7-75·9) while CRP achieved a sensitivity of only 13·6% (CI 7·3-23·4). In this first accuracy study of a prototype blood-based host-marker assay, we show the possible value of the assay for triage and diagnosis in PLHIV.
View details for DOI 10.1128/JCM.01643-20
View details for PubMedID 33298607
Data Heterogeneity: The Enzyme to Catalyze Translational Bioinformatics?
Journal of medical Internet research
2020; 22 (8): e18044
Up to 95% of novel interventions demonstrating significant effects at the bench fail to translate to the bedside. In recent years, the windfalls of "big data" have afforded investigators more substrate for research than ever before. However, issues with translation have persisted: although countless biomarkers for diagnostic and therapeutic targeting have been proposed, few of these generalize effectively. We assert that inadequate heterogeneity in datasets used for discovery and validation causes their nonrepresentativeness of the diversity observed in real-world patient populations. This nonrepresentativeness is contrasted with advantages rendered by the solicitation and utilization of data heterogeneity for multisystemic disease modeling. Accordingly, we propose the potential benefits of models premised on heterogeneity to promote the Institute for Healthcare Improvement's Triple Aim. In an era of personalized medicine, these models can confer higher quality clinical care for individuals, increased access to effective care across all populations, and lower costs for the health care system.
View details for DOI 10.2196/18044
View details for PubMedID 32784182
Systems biological assessment of immunity to mild versus severe COVID-19 infection in humans.
Science (New York, N.Y.)
COVID-19 represents a global crisis, yet major knowledge gaps remain about human immunity to SARS-CoV-2. We analyzed immune responses in 76 COVID-19 patients and 69 healthy individuals from Hong Kong and Atlanta. In PBMCs of COVID-19 patients, there was reduced expression of HLA-DR and pro-inflammatory cytokines by myeloid cells, and impaired mTOR-signaling and IFN-alpha production by plasmacytoid DCs. In contrast, there were enhanced plasma levels of inflammatory mediators, including EN-RAGE, TNFSF14, and oncostatin-M, which correlated with disease severity and increased bacterial products in human plasma. Single-cell transcriptomics revealed no type-I IFN, reduced HLA-DR in myeloid cells of severe patients, and transient expression of IFN-stimulated genes. This was consistent with bulk PBMC transcriptomics, and transient, low plasma IFN-alpha levels during infection. These results reveal mechanisms and potential therapeutic targets for COVID-19.
View details for DOI 10.1126/science.abc6261
View details for PubMedID 32788292
Cellular senescence impairs the reversibility of pulmonary arterial hypertension.
Science translational medicine
2020; 12 (554)
Pulmonary arterial hypertension (PAH) in congenital cardiac shunts can be reversed by hemodynamic unloading (HU) through shunt closure. However, this reversibility potential is lost beyond a certain point in time. The reason why PAH becomes irreversible is unknown. In this study, we used MCT+shunt-induced PAH in rats to identify a dichotomous reversibility response to HU, similar to the human situation. We compared vascular profiles of reversible and irreversible PAH using RNA sequencing. Cumulatively, we report that loss of reversibility is associated with a switch from a proliferative to a senescent vascular phenotype and confirmed markers of senescence in human PAH-CHD tissue. In vitro, we showed that human pulmonary endothelial cells of patients with PAH are more vulnerable to senescence than controls in response to shear stress and confirmed that the senolytic ABT263 induces apoptosis in senescent, but not in normal, endothelial cells. To support the concept that vascular cell senescence is causal to the irreversible nature of end-stage PAH, we targeted senescence using ABT263 and induced reversal of the hemodynamic and structural changes associated with severe PAH refractory to HU. The factors that drive the transition from a reversible to irreversible pulmonary vascular phenotype could also explain the irreversible nature of other PAH etiologies and provide new leads for pharmacological reversal of end-stage PAH.
View details for DOI 10.1126/scitranslmed.aaw4974
View details for PubMedID 32727916
- Response to: 'Successful treatment of plasma exchange for refractory systemic juvenile idiopathic arthritis complicated with macrophage activation syndrome and severe lung disease' by Sato et al. Annals of the rheumatic diseases 2020
A generalizable 29-mRNA neural-network classifier for acute bacterial and viral infections.
2020; 11 (1): 1177
Improved identification of bacterial and viral infections would reduce morbidity from sepsis, reduce antibiotic overuse, and lower healthcare costs. Here, we develop a generalizable host-gene-expression-based classifier for acute bacterial and viral infections. We use training data (N=1069) from 18 retrospective transcriptomic studies. Using only 29 preselected host mRNAs, we train a neural-network classifier with a bacterial-vs-other area under the receiver-operating characteristic curve (AUROC) 0.92 (95% CI 0.90-0.93) and a viral-vs-other AUROC 0.92 (95% CI 0.90-0.93). We then apply this classifier, inflammatix-bacterial-viral-noninfected-version 1(IMX-BVN-1), without retraining, to an independent cohort (N=163). In this cohort, IMX-BVN-1 AUROCs are: bacterial-vs.-other 0.86 (95% CI 0.77-0.93), and viral-vs.-other 0.85 (95% CI 0.76-0.93). In patients enrolled within 36h of hospital admission (N=70), IMX-BVN-1 AUROCs are: bacterial-vs.-other 0.92 (95% CI 0.83-0.99), and viral-vs.-other 0.91 (95% CI 0.82-0.98). With further study, IMX-BVN-1 could provide a tool for assessing patients with suspected infection and sepsis at hospital admission.
View details for DOI 10.1038/s41467-020-14975-w
View details for PubMedID 32132525
- Response to: 'Effectiveness and safety of ruxolitinib for the treatment of refractory systemic idiopathic juvenile arthritis like associated with interstitial lung disease: case report' by Bader-Meunier et al. Annals of the rheumatic diseases 2020
High-throughput quantitative histology in systemic sclerosis skin disease using computer vision.
Arthritis research & therapy
2020; 22 (1): 48
Skin fibrosis is the clinical hallmark of systemic sclerosis (SSc), where collagen deposition and remodeling of the dermis occur over time. The most widely used outcome measure in SSc clinical trials is the modified Rodnan skin score (mRSS), which is a semi-quantitative assessment of skin stiffness at seventeen body sites. However, the mRSS is confounded by obesity, edema, and high inter-rater variability. In order to develop a new histopathological outcome measure for SSc, we applied a computer vision technology called a deep neural network (DNN) to stained sections of SSc skin. We tested the hypotheses that DNN analysis could reliably assess mRSS and discriminate SSc from normal skin.We analyzed biopsies from two independent (primary and secondary) cohorts. One investigator performed mRSS assessments and forearm biopsies, and trichrome-stained biopsy sections were photomicrographed. We used the AlexNet DNN to generate a numerical signature of 4096 quantitative image features (QIFs) for 100 randomly selected dermal image patches/biopsy. In the primary cohort, we used principal components analysis (PCA) to summarize the QIFs into a Biopsy Score for comparison with mRSS. In the secondary cohort, using QIF signatures as the input, we fit a logistic regression model to discriminate between SSc vs. control biopsy, and a linear regression model to estimate mRSS, yielding Diagnostic Scores and Fibrosis Scores, respectively. We determined the correlation between Fibrosis Scores and the published Scleroderma Skin Severity Score (4S) and between Fibrosis Scores and longitudinal changes in mRSS on a per patient basis.In the primary cohort (n = 6, 26 SSc biopsies), Biopsy Scores significantly correlated with mRSS (R = 0.55, p = 0.01). In the secondary cohort (n = 60 SSc and 16 controls, 164 biopsies; divided into 70% training and 30% test sets), the Diagnostic Score was significantly associated with SSc-status (misclassification rate = 1.9% [training], 6.6% [test]), and the Fibrosis Score significantly correlated with mRSS (R = 0.70 [training], 0.55 [test]). The DNN-derived Fibrosis Score significantly correlated with 4S (R = 0.69, p = 3 × 10- 17).DNN analysis of SSc biopsies is an unbiased, quantitative, and reproducible outcome that is associated with validated SSc outcomes.
View details for DOI 10.1186/s13075-020-2127-0
View details for PubMedID 32171325
T cell-inducing vaccine durably prevents mucosal SHIV infection even with lower neutralizing antibody titers.
Recent efforts toward an HIV vaccine focus on inducing broadly neutralizing antibodies, but eliciting both neutralizing antibodies (nAbs) and cellular responses may be superior. Here, we immunized macaques with an HIV envelope trimer, either alone to induce nAbs, or together with a heterologous viral vector regimen to elicit nAbs and cellular immunity, including CD8+ tissue-resident memory T cells. After ten vaginal challenges with autologous virus, protection was observed in both vaccine groups at 53.3% and 66.7%, respectively. A nAb titer >300 was generally associated with protection but in the heterologous viral vector + nAb group, titers <300 were sufficient. In this group, protection was durable as the animals resisted six more challenges 5 months later. Antigen stimulation of T cells in ex vivo vaginal tissue cultures triggered antiviral responses in myeloid and CD4+ T cells. We propose that cellular immune responses reduce the threshold of nAbs required to confer superior and durable protection.
View details for DOI 10.1038/s41591-020-0858-8
View details for PubMedID 32393800
Integrated, multicohort analysis reveals unified signature of systemic lupus erythematosus.
2020; 5 (4)
Systemic lupus erythematosus (SLE) is a complex autoimmune disease that follows an unpredictable disease course and affects multiple organs and tissues. We performed an integrated, multicohort analysis of 7,471 transcriptomic profiles from 40 independent studies to identify robust gene expression changes associated with SLE. We identified a 93-gene signature (SLE MetaSignature) that is differentially expressed in the blood of patients with SLE compared with healthy volunteers; distinguishes SLE from other autoimmune, inflammatory, and infectious diseases; and persists across diverse tissues and cell types. The SLE MetaSignature correlated significantly with disease activity and other clinical measures of inflammation. We prospectively validated the SLE MetaSignature in an independent cohort of pediatric patients with SLE using a microfluidic quantitative PCR (qPCR) array. We found that 14 of the 93 genes in the SLE MetaSignature were independent of IFN-induced and neutrophil-related transcriptional profiles that have previously been associated with SLE. Pathway analysis revealed dysregulation associated with nucleic acid biosynthesis and immunometabolism in SLE. We further refined a neutropoiesis signature and identified underappreciated transcripts related to immune cells and oxidative stress. In our multicohort, transcriptomic analysis has uncovered underappreciated genes and pathways associated with SLE pathogenesis, with the potential to advance clinical diagnosis, biomarker development, and targeted therapeutics for SLE.
View details for DOI 10.1172/jci.insight.122312
View details for PubMedID 31971918
Pilot study of a novel serum mRNA gene panel for diagnosis of acute septic arthritis.
World journal of orthopedics
2019; 10 (12): 424–33
BACKGROUND: Septic arthritis is an orthopedic emergency requiring immediate surgical intervention. Current diagnostic standard of care is an invasive joint aspiration. Aspirations provide information about the inflammatory cells in the sample within a few hours, but there is often ambiguity about whether the source is infectious (e.g. bacterial) or non-infectious (e.g. gout). Cultures can take days to result, so decisions about surgery are often made with incomplete data. Novel diagnostics are thus needed. The "Sepsis MetaScore" (SMS) is an 11-mRNA host immune blood signature that can distinguish between infectious and non-infectious acute inflammation. It has been validated in multiple cohorts across heterogeneous clinical settings.AIM: To study whether the SMS holds diagnostic validity in determining the etiology of acute arthritis.METHODS: We conducted a blinded, prospective, non-interventional clinical study of the SMS. All patients undergoing work-up for a septic primary joint were enrolled. Patients proceeded through the normal standard-of-care pathway, including joint aspiration and inflammatory labs [white blood cell (WBC), erythrocyte sedimentation rate (ESR), C-reactive protein (CRP)]. Venous blood was also drawn into PAX gene RNA-stabilizing tubes and mRNAs were measured using Nano String nCounter. SMS was calculated blinded to clinical results.RESULTS: A total of 20 samples were included, of which 11 were infected based on aspiration or intra-operative cultures. The SMS had an area under the ROC curve (AUROC) of 0.87 for separating infectious from non-infectious conditions. For comparison, the AUROCs for ESR = 0.58, CRP = 0.6, and WBC = 0.59. At 100% sensitivity for infection, the specificity of the SMS was 40%, meaning nearly half of non-septic patients could have been ruled out for further intervention.CONCLUSION: In this pilot study, SMS showed a high level of diagnostic accuracy in predicting septic joints compared to other diagnostic biomarkers. This quick blood test could be an important tool for early, accurate identification of acute septic joints and need for emergent surgery, improving clinical care and healthcare spending.
View details for DOI 10.5312/wjo.v10.i12.424
View details for PubMedID 31908991
Sex Differences in the Blood Transcriptome Identify Robust Changes in Immune Cell Proportions with Aging and Influenza Infection.
2019; 29 (7): 1961
Sex differences in autoimmunity and infection suggest that a better understanding of molecular sex differences will improve the diagnosis and treatment of immune-related disease. We identified 144 differentially expressed genes, referred to as immune sex expression signature (iSEXS), between human males and females using an integrated multi-cohort analysis of blood transcriptome profiles from six discovery cohorts from five continents with 458 healthy individuals. We validated iSEXS in 11 additional cohorts of 524 peripheral blood samples. When we separated iSEXS into genes located on sex chromosomes (XY-iSEXS) or autosomes (autosomal-iSEXS), both modules distinguished males and females. iSEXS reflects sex differences in immune cell proportions, with female-associated genes showing higher expression by CD4+ Tcells and male-associated genes showing higher expression by myeloid cells. Autosomal-iSEXS detected an increase in monocytes with age in females, reflected sex-differential immune cell dynamics during influenza infection, and predicted antibody response in males, but not females.
View details for DOI 10.1016/j.celrep.2019.10.019
View details for PubMedID 31722210
Multiplex Serum Analysis Identifies Potential Biomarkers of Systemic Juvenile Idiopathic Arthritis, Macrophage Activation Syndrome, and Associated Pulmonary Alveolar Proteinosis: Evidence for Independently-regulated Hyperinflammatory and Eosinophilic Inflammation
View details for Web of Science ID 000507466901341
Cytokine Signatures Differentiate Systemic Sclerosis Patients at High versus Low Risk for Pulmonary Arterial Hypertension
View details for Web of Science ID 000507466901493
Transplantation Alters Function and Clonality of Cytomegalovirus-Responsive T Cells
LIPPINCOTT WILLIAMS & WILKINS. 2019
View details for Web of Science ID 000511467800256
Computational and Systems Immunology: A Student's Perspective.
Trends in immunology
The big data revolution has transformed the landscape of immunology research. As inaugural students of Stanford's new Computational and Systems Immunology PhD track, we share our experiences and advice with other institutions considering a similar program.
View details for DOI 10.1016/j.it.2019.06.006
View details for PubMedID 31288986
- Increased monocyte count as a cellular biomarker for poor outcomes in fibrotic diseases: a retrospective, multicentre cohort study LANCET RESPIRATORY MEDICINE 2019; 7 (6): 497–508
- Single-cell technologies - studying rheumatic diseases one cell at a time NATURE REVIEWS RHEUMATOLOGY 2019; 15 (6): 340–54
- Single cell immune profiling in transplantation research AMERICAN JOURNAL OF TRANSPLANTATION 2019; 19 (5): 1278–87
- CD22 blockade restores homeostatic microglial phagocytosis in ageing brains NATURE 2019; 568 (7751): 187-+
CD22 blockade restores homeostatic microglial phagocytosis in ageing brains.
Microglia maintain homeostasis in the central nervous system through phagocytic clearance of protein aggregates and cellular debris. This function deteriorates during ageing and neurodegenerative disease, concomitant with cognitive decline. However, the mechanisms of impaired microglial homeostatic function and the cognitive effects of restoring this function remain unknown. We combined CRISPR-Cas9 knockout screens with RNAsequencing analysis to discover age-related genetic modifiers of microglial phagocytosis. These screens identified CD22, a canonical Bcell receptor, as a negative regulator of phagocytosis that is upregulated on aged microglia. CD22 mediates the anti-phagocytic effect of alpha2,6-linked sialic acid, and inhibition of CD22 promotes the clearance of myelin debris, amyloid-beta oligomers and alpha-synuclein fibrils in vivo. Long-term central nervous system delivery of an antibody that blocks CD22 function reprograms microglia towards a homeostatic transcriptional state and improves cognitive function in aged mice. These findings elucidate a mechanism of age-related microglial impairment and a strategy to restore homeostasis in the ageing brain.
View details for PubMedID 30944478
- Host-response-based gene signatures for tuberculosis diagnosis: A systematic comparison of 16 signatures PLOS MEDICINE 2019; 16 (4)
Increased monocyte count as a cellular biomarker for poor outcomes in fibrotic diseases: a retrospective, multicentre cohort study.
The Lancet. Respiratory medicine
BACKGROUND: There is an urgent need for biomarkers to better stratify patients with idiopathic pulmonary fibrosis by risk for lung transplantation allocation who have the same clinical presentation. We aimed to investigate whether a specific immune cell type from patients with idiopathic pulmonary fibrosis could identify those at higher risk of poor outcomes. We then sought to validate our findings using cytometry and electronic health records.METHODS: We first did a discovery analysis with transcriptome data from the Gene Expression Omnibus at the National Center for Biotechnology Information for 120 peripheral blood mononuclear cell (PBMC) samples of patients with idiopathic pulmonary fibrosis. We estimated percentages of 13 immune cell types using statistical deconvolution, and investigated the association of these cell types with transplant-free survival. We validated these results using PBMC samples from patients with idiopathic pulmonary fibrosis in two independent cohorts (COMET and Yale). COMET profiled monocyte counts in 45 patients with idiopathic pulmonary fibrosis from March 12, 2010, to March 10, 2011, using flow cytometry; we tested if increased monocyte count was associated with the primary outcome of disease progression. In the Yale cohort, 15 patients with idiopathic pulmonary fibrosis (with five healthy controls) were classed as high risk or low risk from April 28, 2014, to Aug 20, 2015, using a 52-gene signature, and we assessed whether monocyte percentage (measured by cytometry by time of flight) was higher in high-risk patients. We then examined complete blood count values in the electronic health records (EHR) of 45 068 patients with idiopathic pulmonary fibrosis, systemic sclerosis, hypertrophic cardiomyopathy, or myelofibrosis from Stanford (Jan 01, 2008, to Dec 31, 2015), Northwestern (Feb 15, 2001 to July 31, 2017), Vanderbilt (Jan 01, 2008, to Dec 31, 2016), and Optum Clinformatics DataMart (Jan 01, 2004, to Dec 31, 2016) cohorts, and examined whether absolute monocyte counts of 0·95 K/muL or greater were associated with all-cause mortality in these patients.FINDINGS: In the discovery analysis, estimated CD14+ classical monocyte percentages above the mean were associated with shorter transplant-free survival times (hazard ratio [HR] 1·82, 95% CI 1·05-3·14), whereas higher percentages of T cells and B cells were not (0·97, 0·59-1·66; and 0·78, 0·45-1·34 respectively). In two validation cohorts (COMET trial and the Yale cohort), patients with higher monocyte counts were at higher risk for poor outcomes (COMET Wilcoxon p=0·025; Yale Wilcoxon p=0·049). Monocyte counts of 0·95 K/muL or greater were associated with mortality after adjusting for forced vital capacity (HR 2·47, 95% CI 1·48-4·15; p=0·0063), and the gender, age, and physiology index (HR 2·06, 95% CI 1·22-3·47; p=0·0068) across the COMET, Stanford, and Northwestern datasets). Analysis of medical records of 7459 patients with idiopathic pulmonary fibrosis showed that patients with monocyte counts of 0·95 K/muL or greater were at increased risk of mortality with lung transplantation as a censoring event, after adjusting for age at diagnosis and sex (Stanford HR=2·30, 95% CI 0·94-5·63; Vanderbilt 1·52, 1·21-1·89; Optum 1·74, 1·33-2·27). Likewise, higher absolute monocyte count was associated with shortened survival in patients with hypertrophic cardiomyopathy across all three cohorts, and in patients with systemic sclerosis or myelofibrosis in two of the three cohorts.INTERPRETATION: Monocyte count could be incorporated into the clinical assessment of patients with idiopathic pulmonary fibrosis and other fibrotic disorders. Further investigation into the mechanistic role of monocytes in fibrosis might lead to insights that assist the development of new therapies.FUNDING: Bill & Melinda Gates Foundation, US National Institute of Allergy and Infectious Diseases, and US National Library of Medicine.
View details for PubMedID 30935881
- Discovery of Distinct Immune Phenotypes Using Machine Learning in Pulmonary Arterial Hypertension CIRCULATION RESEARCH 2019; 124 (6): 904–19
A clinically meaningful metric of immune age derived from high-dimensional longitudinal monitoring.
Immune responses generally decline with age. However, the dynamics of this process at the individual level have not been characterized, hindering quantification of an individual's immune age. Here, we use multiple 'omics' technologies to capture population- and individual-level changes in the human immune system of 135 healthy adult individuals of different ages sampled longitudinally over a nine-year period. We observed high inter-individual variability in the rates of change of cellular frequencies that was dictated by their baseline values, allowing identification of steady-state levels toward which a cell subset converged and the ordered convergence of multiple cell subsets toward an older adult homeostasis. These data form a high-dimensional trajectory of immune aging (IMM-AGE) that describes a person's immune status better than chronological age. We show that the IMM-AGE score predicted all-cause mortality beyond well-established risk factors in the Framingham Heart Study, establishing its potential use in clinics for identification of patients at risk.
View details for PubMedID 30842675
- A clinically meaningful metric of immune age derived from high-dimensional longitudinal monitoring NATURE MEDICINE 2019; 25 (3): 487-+
Single cell immune profiling in transplantation research.
American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons
Recently developed single-cell profiling technologies hold promise to provide new insights including analysis of population heterogeneity and linkage of antigen receptors with gene expression. These technologies produce complex data sets that require knowledge of bioinformatics for appropriate analysis. In this minireview, we discuss several single-cell immune profiling technologies for gene and protein expression, including cytometry by time-of-flight, RNA sequencing, and antigen receptor sequencing, as well as key considerations for analysis that apply to each. Because of the critical importance of data analysis for high parameter single cell analysis, we discuss essential factors in analysis of these data, including quality control, quantification, examples of methods for high dimensional analysis, immune repertoire analysis, and preparation of analysis pipelines. We provide examples of, and suggestions for, application of these innovative methods to transplantation research. This article is protected by copyright. All rights reserved.
View details for PubMedID 30768832
A 20-Gene Set Predictive of Progression to Severe Dengue.
2019; 26 (5): 1104
There is a need to identify biomarkers predictive of severe dengue. Single-cohort transcriptomics has not yielded generalizable results or parsimonious, predictive gene sets. We analyzed blood samples of dengue patients from seven gene expression datasets (446 samples, five countries) using an integrated multi-cohort analysis framework and identified a 20-gene set that predicts progression to severe dengue. We validated the predictive power of this 20-gene set in three retrospective dengue datasets (84 samples, three countries) and a prospective Colombia cohort (34 patients), with an area under the receiver operating characteristic curve of 0.89, 100% sensitivity, and 76% specificity. The 20-gene dengue severity scores declined during the diseasecourse, suggesting an infection-triggered host response. This 20-gene set is strongly associated with the progression to severe dengue and represents a predictive signature, generalizable across ages, host genetic factors, and virus strains, with potential implications for the development of a host response-based dengue prognostic assay.
View details for PubMedID 30699342
- A 20-Gene Set Predictive of Progression to Severe Dengue CELL REPORTS 2019; 26 (5): 1104-+
Single-cell technologies - studying rheumatic diseases one cell at a time.
Nature reviews. Rheumatology
Cells, the basic units of life, have striking differences at transcriptomic, proteomic and epigenomic levels across tissues, organs, organ systems and organisms. The coordination of individual immune cells is essential for the generation of effective immune responses to pathogens while immune tolerance is maintained to protect the host. In rheumatic diseases, when immune responses are dysregulated, pathologically important cells might represent only a small fraction of the immune system. Interrogation of the contributions of individual immune cells to pathogenesis and disease progression should therefore reveal important insights into the complicated aetiology of rheumatic diseases. Technological advances are enabling the high-dimensional dissection of single cells at multiple omics levels, which could facilitate the identification of dysregulated molecular mechanisms in patients with rheumatic diseases and the discovery of new therapeutic targets and biomarkers. The single-cell technologies that have been developed over the past decade and the experimental platforms that enable multi-omics integrative analyses have already made inroads into immunology-related fields of study and have potential for use in rheumatology. Layers of omics data derived from single cells are likely to fundamentally change our understanding of the molecular pathways that underpin the pathogenesis of rheumatic diseases.
View details for PubMedID 31065108
- FHIT, a Novel Modifier Gene in Pulmonary Arterial Hypertension AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE 2019; 199 (1): 83-98
- Data analytics for precision medicine GENOMIC AND PRECISION MEDICINE: INFECTIOUS AND INFLAMMATORY DISEASE, 3RD EDITION 2019: 25–33
Pregnancy-Induced Alterations in NK Cell Phenotype and Function.
Frontiers in immunology
2019; 10: 2469
Pregnant women are particularly susceptible to complications of influenza A virus infection, which may result from pregnancy-induced changes in the function of immune cells, including natural killer (NK) cells. To better understand NK cell function during pregnancy, we assessed the ability of the two main subsets of NK cells, CD56dim, and CD56bright NK cells, to respond to influenza-virus infected cells and tumor cells. During pregnancy, CD56dim and CD56bright NK cells displayed enhanced functional responses to both infected and tumor cells, with increased expression of degranulation markers and elevated frequency of NK cells producing IFN-gamma. To better understand the mechanisms driving this enhanced function, we profiled CD56dim and CD56bright NK cells from pregnant and non-pregnant women using mass cytometry. NK cells from pregnant women displayed significantly increased expression of several functional and activation markers such as CD38 on both subsets and NKp46 on CD56dim NK cells. NK cells also displayed diminished expression of the chemokine receptor CXCR3 during pregnancy. Overall, these data demonstrate that functional and phenotypic shifts occur in NK cells during pregnancy that can influence the magnitude of the immune response to both infections and tumors.
View details for DOI 10.3389/fimmu.2019.02469
View details for PubMedID 31708922
Emergent high fatality lung disease in systemic juvenile arthritis.
Annals of the rheumatic diseases
To investigate the characteristics and risk factors of a novel parenchymal lung disease (LD), increasingly detected in systemic juvenile idiopathic arthritis (sJIA).In a multicentre retrospective study, 61 cases were investigated using physician-reported clinical information and centralised analyses of radiological, pathological and genetic data.LD was associated with distinctive features, including acute erythematous clubbing and a high frequency of anaphylactic reactions to the interleukin (IL)-6 inhibitor, tocilizumab. Serum ferritin elevation and/or significant lymphopaenia preceded LD detection. The most prevalent chest CT pattern was septal thickening, involving the periphery of multiple lobes ± ground-glass opacities. The predominant pathology (23 of 36) was pulmonary alveolar proteinosis and/or endogenous lipoid pneumonia (PAP/ELP), with atypical features including regional involvement and concomitant vascular changes. Apparent severe delayed drug hypersensitivity occurred in some cases. The 5-year survival was 42%. Whole exome sequencing (20 of 61) did not identify a novel monogenic defect or likely causal PAP-related or macrophage activation syndrome (MAS)-related mutations. Trisomy 21 and young sJIA onset increased LD risk. Exposure to IL-1 and IL-6 inhibitors (46 of 61) was associated with multiple LD features. By several indicators, severity of sJIA was comparable in drug-exposed subjects and published sJIA cohorts. MAS at sJIA onset was increased in the drug-exposed, but was not associated with LD features.A rare, life-threatening lung disease in sJIA is defined by a constellation of unusual clinical characteristics. The pathology, a PAP/ELP variant, suggests macrophage dysfunction. Inhibitor exposure may promote LD, independent of sJIA severity, in a small subset of treated patients. Treatment/prevention strategies are needed.
View details for DOI 10.1136/annrheumdis-2019-216040
View details for PubMedID 31562126
Discovery of Distinct Immune Phenotypes Using Machine Learning in Pulmonary Arterial Hypertension.
Accumulating evidence implicates inflammation in pulmonary arterial hypertension (PAH) and therapies targeting immunity are under investigation, though it remains unknown if distinct immune phenotypes exist.Identify PAH immune phenotypes based on unsupervised analysis of blood proteomic profiles.In a prospective observational study of Group 1 PAH patients evaluated at Stanford University (discovery cohort, n=281) and University of Sheffield (validation cohort, n=104) between 2008-2014, we measured a circulating proteomic panel of 48 cytokines, chemokines, and factors using multiplex immunoassay. Unsupervised machine learning (consensus clustering) was applied in both cohorts independently to classify patients into proteomic immune clusters, without guidance from clinical features. To identify central proteins in each cluster, we performed partial correlation network analysis. Clinical characteristics and outcomes were subsequently compared across clusters. Four PAH clusters with distinct proteomic immune profiles were identified in the discovery cohort. Cluster 2 (n=109) had low cytokine levels similar to controls. Other clusters had unique sets of upregulated proteins central to immune networks- cluster 1 (n=58)(TRAIL, CCL5, CCL7, CCL4, MIF), cluster 3 (n=77)(IL-12, IL-17, IL-10, IL-7, VEGF), and cluster 4 (n=37)(IL-8, IL-4, PDGF-β, IL-6, CCL11). Demographics, PAH etiologies, comorbidities, and medications were similar across clusters. Non-invasive and hemodynamic surrogates of clinical risk identified cluster 1 as high-risk and cluster 3 as low-risk groups. Five-year transplant-free survival rates were unfavorable for cluster 1 (47.6%, CI 35.4-64.1%) and favorable for cluster 3 (82.4%, CI 72.0-94.3%)(across-cluster p<0.001). Findings were replicated in the validation cohort, where machine learning classified four immune clusters with comparable proteomic, clinical, and prognostic features.Blood cytokine profiles distinguish PAH immune phenotypes with differing clinical risk that are independent of World Health Organization Group 1 subtypes. These phenotypes could inform mechanistic studies of disease pathobiology and provide a framework to examine patient responses to emerging therapies targeting immunity.
View details for PubMedID 30661465
Cell-centred meta-analysis reveals baseline predictors of anti-TNFα non-response in biopsy and blood of patients with IBD.
2019; 68 (4): 604–14
Although anti-tumour necrosis factor alpha (anti-TNFα) therapies represent a major breakthrough in IBD therapy, their cost-benefit ratio is hampered by an overall 30% non-response rate, adverse side effects and high costs. Thus, finding predictive biomarkers of non-response prior to commencing anti-TNFα therapy is of high value.We analysed publicly available whole-genome expression profiles of colon biopsies obtained from multiple cohorts of patients with IBD using a combined computational deconvolution-meta-analysis paradigm which allows to estimate immune cell contribution to the measured expression and capture differential regulatory programmes otherwise masked due to variation in cellular composition. Insights from this in silico approach were experimentally validated in biopsies and blood samples of three independent test cohorts.We found the proportion of plasma cells as a robust pretreatment biomarker of non-response to therapy, which we validated in two independent cohorts of immune-stained colon biopsies, where a plasma cellular score from inflamed biopsies was predictive of non-response with an area under the curve (AUC) of 82%. Meta-analysis of the cell proportion-adjusted gene expression data suggested that an increase in inflammatory macrophages in anti-TNFα non-responding individuals is associated with the upregulation of the triggering receptor expressed on myeloid cells 1 (TREM-1) and chemokine receptor type 2 (CCR2)-chemokine ligand 7 (CCL7) -axes. Blood gene expression analysis of an independent cohort, identified TREM-1 downregulation in non-responders at baseline, which was predictive of response with an AUC of 94%.Our study proposes two clinically feasible assays, one in biopsy and one in blood, for predicting non-response to anti-TNFα therapy prior to initiation of treatment. Moreover, it suggests that mechanism-driven novel drugs for non-responders should be developed.
View details for PubMedID 29618496
Host-response-based gene signatures for tuberculosis diagnosis: A systematic comparison of 16 signatures.
2019; 16 (4): e1002786
The World Health Organization (WHO) and Foundation for Innovative New Diagnostics (FIND) have published target product profiles (TPPs) calling for non-sputum-based diagnostic tests for the diagnosis of active tuberculosis (ATB) disease and for predicting the progression from latent tuberculosis infection (LTBI) to ATB. A large number of host-derived blood-based gene-expression biomarkers for diagnosis of patients with ATB have been proposed to date, but none have been implemented in clinical settings. The focus of this study is to directly compare published gene signatures for diagnosis of patients with ATB across a large, diverse list of publicly available gene expression datasets, and evaluate their performance against the WHO/FIND TPPs.We searched PubMed, Gene Expression Omnibus (GEO), and ArrayExpress in June 2018. We included all studies irrespective of study design and enrollment criteria. We found 16 gene signatures for the diagnosis of ATB compared to other clinical conditions in PubMed. For each signature, we implemented a classification model as described in the corresponding original publication of the signature. We identified 24 datasets containing 3,083 transcriptome profiles from whole blood or peripheral blood mononuclear cell samples of healthy controls or patients with ATB, LTBI, or other diseases from 14 countries in GEO. Using these datasets, we calculated weighted mean area under the receiver operating characteristic curve (AUROC), specificity at 90% sensitivity, and negative predictive value (NPV) for each gene signature across all datasets. We also compared the diagnostic odds ratio (DOR), heterogeneity in DOR, and false positive rate (FPR) for each signature using bivariate meta-analysis. Across 9 datasets of patients with culture-confirmed diagnosis of ATB, 11 signatures had weighted mean AUROC > 0.8, and 2 signatures had weighted mean AUROC ≤ 0.6. All but 2 signatures had high NPV (>98% at 2% prevalence). Two gene signatures achieved the minimal WHO TPP for a non-sputum-based triage test. When including datasets with clinical diagnosis of ATB, there was minimal reduction in the weighted mean AUROC and specificity of all but 3 signatures compared to when using only culture-confirmed ATB data. Only 4 signatures had homogeneous DOR and lower FPR when datasets with clinical diagnosis of ATB were included; other signatures either had heterogeneous DOR or higher FPR or both. Finally, 7 of 16 gene signatures predicted progression from LTBI to ATB 6 months prior to sputum conversion with positive predictive value > 6% at 2% prevalence. Our analyses may have under- or overestimated the performance of certain ATB diagnostic signatures because our implementation may be different from the published models for those signatures. We re-implemented published models because the exact models were not publicly available.We found that host-response-based diagnostics could accurately identify patients with ATB and predict individuals with high risk of progression from LTBI to ATB prior to sputum conversion. We found that a higher number of genes in a signature did not increase the accuracy of the signature. Overall, the Sweeney3 signature performed robustly across all comparisons. Our results provide strong evidence for the potential of host-response-based diagnostics in achieving the WHO goal of ending tuberculosis by 2035, and host-response-based diagnostics should be pursued for clinical implementation.
View details for PubMedID 31013272
MECHANISMS DRIVING ALTERED V Delta 2+Gamma Delta T CELL FUNCTION DURING RECURRENT MALARIA INFECTION
AMER SOC TROP MED & HYGIENE. 2019: 111
View details for Web of Science ID 000507364502362
High-throughput Drug Screen to Reverse Phenotype of Pulmonary Artery Hypertension Ipsc Derived Vascular Cells Combined with Bioinformatics Uncovers Promising Therapies
LIPPINCOTT WILLIAMS & WILKINS. 2018: E70
View details for Web of Science ID 000454198400020
- Comparison of the Transcriptomic Signature of Pediatric Vs. Adult CML and Normal Bone Marrow Stem Cells AMER SOC HEMATOLOGY. 2018
Early life immunity in the era of systems biology: understanding development and disease.
2018; 10 (1): 88
Systems immunology has the potential to offer invaluable insights into the development of the immune system. Two recent studies offer an in-depth view of both the dynamics of immune system development and the heritability of the levels of key immune modulators at birth.
View details for PubMedID 30470248
- Early life immunity in the era of systems biology: understanding development and disease GENOME MEDICINE 2018; 10
Leveraging heterogeneity across multiple datasets increases cell-mixture deconvolution accuracy and reduces biological and technical biases.
2018; 9 (1): 4735
In silico quantification of cell proportions from mixed-cell transcriptomics data (deconvolution) requires a reference expression matrix, called basis matrix. We hypothesize that matrices created using only healthy samples from a single microarray platform would introduce biological and technical biases in deconvolution. We show presence of such biases in two existing matrices, IRIS and LM22, irrespective of deconvolution method. Here, we present immunoStates, a basis matrix built using 6160 samples with different disease states across 42 microarray platforms. We find that immunoStates significantly reduces biological and technical biases. Importantly, we find that different methods have virtually no or minimal effect once the basis matrix is chosen. We further show that cellular proportion estimates using immunoStates are consistently more correlated with measured proportions than IRIS and LM22, across all methods. Our results demonstrate the need and importance of incorporating biological and technical heterogeneity in a basis matrix for achieving consistently high accuracy.
View details for PubMedID 30413720
- Leveraging heterogeneity across multiple datasets increases cell-mixture deconvolution accuracy and reduces biological and technical biases NATURE COMMUNICATIONS 2018; 9
- Single-cell epigenetics - Chromatin modification atlas unveiled by mass cytometry CLINICAL IMMUNOLOGY 2018; 196: 40–48
Author Correction: A multi-cohort study of the immune factors associated with M. tuberculosis infection outcomes.
The spelling of author Qianting Yang was corrected; the affiliation of author Stephanus T. Malherbe was corrected; and graphs in Fig. 4b and c were corrected owing to reanalysis of the data into the correct timed intervals.
View details for PubMedID 30377311
Assessment of Validity of a Blood-Based 3-Gene Signature Score for Progression and Diagnosis of Tuberculosis, Disease Severity, and Treatment Response.
JAMA network open
2018; 1 (6): e183779
The World Health Organization identified the need for a non-sputum-based triage test to identify those in need of further tuberculosis (TB) testing.To determine whether the 3-gene TB score can be a diagnostic tool throughout the course of TB disease, from latency to diagnosis to treatment response, and posttreatment residual inflammation.This nested case-control study analyzed the 3-gene TB score in 3 cohorts, each focusing on a different stage of TB disease: (1) the Adolescent Cohort Study profiled whole-blood samples from adolescents with latent Mycobacterium tuberculosis infection, some of which progressed to active TB (ATB), using RNA sequencing; (2) the Brazil Active Screen Study collected whole blood from an actively screened case-control cohort of adult inmates from 2 prisons in Mato Grosso do Sul, Brazil, for ATB from January 2016 to February 2016; and (3) the Catalysis Treatment Response Cohort (CTRC) identified culture-positive adults in primary health care clinics in Cape Town, South Africa, from 2005 to 2007 and collected whole blood for RNA sequencing from patients with ATB at diagnosis and weeks 1, 4, and 24. The CTRC patients also had positron emission tomography-computed tomography scans at diagnosis, week 4, and week 24. Analyses were performed from September 2017 to June 2018.A 3-gene messenger RNA expression score, measured by quantitative polymerase chain reaction or RNA sequencing, was evaluated for distinguishing the following: individuals who progressed to ATB from those who did not, individuals with ATB from those without, and individuals with slower treatment response during TB therapy.Patients evaluated in this study included 144 adolescents from the Adolescent Cohort Study (aged 12-18 years; 96 female and 48 male), 81 adult prison inmates from the Brazil Active Screen Study (aged 20-72 years; 81 male), and 138 adult community members from the CTRC (aged 17-64 years; 81 female and 57 male). The 3-gene TB score identified progression from latent M tuberculosis infection to ATB 6 months prior to sputum conversion with 86% sensitivity and 84% specificity (area under the curve [AUC], 0.86; 95% CI, 0.77-0.96) and patients with ATB in the Brazil Active Screen Study cohort (AUC, 0.87; 95% CI, 0.78-0.95) and CTRC (AUC, 0.94; 95% CI, 0.88-0.99). It also identified CTRC patients with failed treatment at the end of treatment (AUC, 0.93; 95% CI, 0.83-1.00). Collectively, across all cohorts, the 3-gene TB score identified patients with ATB with 90% sensitivity, 70% specificity, and 99.3% negative predictive value at 4% prevalence.Across 3 independent prospective cohorts, the 3-gene TB score approaches the World Health Organization target product profile benchmarks for non-sputum-based triage test with high negative predictive value. This gene expression diagnostic approach should be considered for further validation and future implementation.
View details for DOI 10.1001/jamanetworkopen.2018.3779
View details for PubMedID 30646264
View details for PubMedCentralID PMC6324428
- A multi-cohort study of the immune factors associated with M. tuberculosis infection outcomes NATURE 2018; 560 (7720): 644-+
A multi-cohort study of the immune factors associated with M. tuberculosis infection outcomes.
Most infections with Mycobacterium tuberculosis (Mtb) manifest as a clinically asymptomatic, contained state, known as latent tuberculosis infection, that affects approximately one-quarter of the global population1. Although fewer than one in ten individuals eventually progress to active disease2, tuberculosis is a leading cause of death from infectious disease worldwide3. Despite intense efforts, immune factors that influence the infection outcomes remain poorly defined. Here we used integrated analyses of multiple cohorts to identify stage-specific host responses to Mtb infection. First, using high-dimensional mass cytometry analyses and functional assays of a cohort of South African adolescents, we show that latent tuberculosis is associated with enhanced cytotoxic responses, which are mostly mediated by CD16 (also known as FcgammaRIIIa) and natural killer cells, and continuous inflammation coupled with immune deviations in both T and B cell compartments.Next, using cell-type deconvolution of transcriptomic data from several cohorts of different ages, genetic backgrounds, geographical locations and infection stages, we show that although deviations in peripheral B and T cell compartments generally start at latency, they are heterogeneous across cohorts. However, an increase in the abundance of circulating natural killer cells in tuberculosis latency, with a corresponding decrease during active disease and a return to baseline levels upon clinical cure are features that are common to all cohorts. Furthermore, by analysing three longitudinal cohorts, we find that changes inperipheral levels of natural killer cells can inform disease progression and treatment responses, and inversely correlate with the inflammatory state of the lungs of patients with active tuberculosis. Together, our findings offer crucial insights into the underlying pathophysiology of tuberculosis latency, and identify factors that may influence infection outcomes.
View details for PubMedID 30135583
Fragile Histidine Triad (FHIT), a Novel Modifier Gene in Pulmonary Arterial Hypertension.
American journal of respiratory and critical care medicine
RATIONALE: Pulmonary arterial hypertension (PAH) is characterized by progressive narrowing of pulmonary arteries resulting in right heart failure and death. Bone Morphogenetic Protein Receptor type-2 (BMPR2) mutations account for most familial PAH (FPAH) forms while reduced BMPR2 is present in many idiopathic PAH (IPAH) forms, suggesting dysfunctional BMPR2 signaling to be a key feature of PAH. Modulating BMPR2 signaling is therapeutically promising, yet how BMPR2 is downregulated in PAH is unclear.OBJECTIVES: We intended to identify and pharmaceutically target BMPR2 modifier genes to improve PAH.METHODS: We combined siRNA High Throughput Screening (HTS) of >20,000 genes with a multi-cohort analysis of publicly available PAH RNA expression data to identify clinically relevant BMPR2-modifiers. After confirming gene dysregulation in PAH patient tissue, we determined the functional roles of BMPR2-modifiers in vitro and tested the repurposed drug Enzastaurin for its propensity to improve experimental PH.MEASUREMENTS AND MAIN RESULTS: We discovered Fragile Histidine Triad (FHIT) as a novel BMPR2-modifier. BMPR2 and FHIT expression were reduced in PAH patients. FHIT reductions were associated with endothelial and smooth muscle cell dysfunction, rescued by Enzastaurin through a dual mechanism: upregulation of FHIT as well as miR17-5 repression. Fhit-/- mice had exaggerated hypoxic PH and failed to recover in normoxia. Enzastaurin reversed PH in the Sugen5416/Hypoxia/Normoxia rat model, by improving Right Ventricular Systolic Pressure (RVSP), RV hypertrophy, cardiac fibrosis and vascular remodeling.CONCLUSIONS: This study highlights the importance of the novel BMPR2 modifier FHIT in PH and the clinical value of the repurposed drug Enzastaurin as a potential novel therapeutic strategy to improve PAH.
View details for PubMedID 30107138
Future Research Directions in Pneumonia NHLBI Working Group Report
AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE
2018; 198 (2): 256–63
Pneumonia is a complex pulmonary disease in need of new clinical approaches. Although triggered by a pathogen, pneumonia often results from dysregulations of host defense that likely precede infection. The coordinated activities of immune resistance and tissue resilience then dictate whether and how pneumonia progresses or resolves. Inadequate or inappropriate host responses lead to more severe outcomes such as acute respiratory distress syndrome and to organ dysfunction beyond the lungs and over extended time frames after pathogen clearance, some of which increase the risk for subsequent pneumonia. Improved understanding of such host responses will guide the development of novel approaches for preventing and curing pneumonia and for mitigating the subsequent pulmonary and extrapulmonary complications of pneumonia. The NHLBI assembled a working group of extramural investigators to prioritize avenues of host-directed pneumonia research that should yield novel approaches for interrupting the cycle of unhealthy decline caused by pneumonia. This report summarizes the working group's specific recommendations in the areas of pneumonia susceptibility, host response, and consequences. Overarching goals include the development of more host-focused clinical approaches for preventing and treating pneumonia, the generation of predictive tools (for pneumonia occurrence, severity, and outcome), and the elucidation of mechanisms mediating immune resistance and tissue resilience in the lung. Specific areas of research are highlighted as especially promising for making advances against pneumonia.
View details for PubMedID 29546996
Single-cell epigenetics - Chromatin modification atlas unveiled by mass cytometry.
Clinical immunology (Orlando, Fla.)
Modifications of histone proteins are fundamental to the regulation of epigenetic phenotypes. Dysregulations of histone modifications have been linked to the pathogenesis of diverse human diseases. However, identifying differential histone modifications in patients with immune-mediated diseases has been challenging, in part due to the lack of a powerful analytic platform to study histone modifications in the complex human immune system. We recently developed a highly multiplexed platform, Epigenetic landscape profiling using cytometry by Time-Of-Flight (EpiTOF), to analyze the global levels of a broad array of histone modifications in single cells using mass cytometry. In this review, we summarize the development of EpiTOF and discuss its potential applications in biomedical research. We anticipate that this platform will provide new insights into the roles of epigenetic regulation in hematopoiesis, immune cell functions and immune system aging, and reveal aberrant epigenetic patterns associated with immune-mediated diseases.
View details for PubMedID 29960011
KLRD1-expressing natural killer cells predict influenza susceptibility
2018; 10: 45
Influenza infects tens of millions of people every year in the USA. Other than notable risk groups, such as children and the elderly, it is difficult to predict what subpopulations are at higher risk of infection. Viral challenge studies, where healthy human volunteers are inoculated with live influenza virus, provide a unique opportunity to study infection susceptibility. Biomarkers predicting influenza susceptibility would be useful for identifying risk groups and designing vaccines.We applied cell mixture deconvolution to estimate immune cell proportions from whole blood transcriptome data in four independent influenza challenge studies. We compared immune cell proportions in the blood between symptomatic shedders and asymptomatic nonshedders across three discovery cohorts prior to influenza inoculation and tested results in a held-out validation challenge cohort.Natural killer (NK) cells were significantly lower in symptomatic shedders at baseline in both discovery and validation cohorts. Hematopoietic stem and progenitor cells (HSPCs) were higher in symptomatic shedders at baseline in discovery cohorts. Although the HSPCs were higher in symptomatic shedders in the validation cohort, the increase was statistically nonsignificant. We observed that a gene associated with NK cells, KLRD1, which encodes CD94, was expressed at lower levels in symptomatic shedders at baseline in discovery and validation cohorts. KLRD1 expression in the blood at baseline negatively correlated with influenza infection symptom severity. KLRD1 expression 8 h post-infection in the nasal epithelium from a rhinovirus challenge study also negatively correlated with symptom severity.We identified KLRD1-expressing NK cells as a potential biomarker for influenza susceptibility. Expression of KLRD1 was inversely correlated with symptom severity. Our results support a model where an early response by KLRD1-expressing NK cells may control influenza infection.
View details for PubMedID 29898768
- Unsupervised Analysis of Transcriptomics in Bacterial Sepsis Across Multiple Datasets Reveals Three Robust Clusters CRITICAL CARE MEDICINE 2018; 46 (6): 915-925
- Validation of the Sepsis MetaScore for Diagnosis of Neonatal Sepsis JOURNAL OF THE PEDIATRIC INFECTIOUS DISEASES SOCIETY 2018; 7 (2): 129-135
- Single-Cell Chromatin Modification Profiling Reveals Increased Epigenetic Variations with Aging CELL 2018; 173 (6): 1385-+
Single-Cell Chromatin Modification Profiling Reveals Increased Epigenetic Variations with Aging.
Post-translational modifications of histone proteins and exchanges of histone variants of chromatin are central to the regulation of nearly all DNA-templated biological processes. However, the degree and variability of chromatin modifications in specific human immune cells remain largely unknown. Here, we employ a highly multiplexed mass cytometry analysis to profile the global levels of a broad array of chromatin modifications in primary human immune cells at the single-cell level. Our data reveal markedly different cell-type- and hematopoietic-lineage-specific chromatin modification patterns. Differential analysis between younger and older adults shows that aging is associated with increased heterogeneity between individuals and elevated cell-to-cell variability in chromatin modifications. Analysis of a twin cohort unveils heritability of chromatin modifications and demonstrates that aging-related chromatin alterations are predominantly driven by non-heritable influences. Together, we present a powerful platform for chromatin and immunology research. Our discoveries highlight the profound impacts of aging on chromatin modifications.
View details for PubMedID 29706550
Interpretation of biological experiments changes with evolution of the Gene Ontology and its annotations
2018; 8: 5115
Gene Ontology (GO) enrichment analysis is ubiquitously used for interpreting high throughput molecular data and generating hypotheses about underlying biological phenomena of experiments. However, the two building blocks of this analysis - the ontology and the annotations - evolve rapidly. We used gene signatures derived from 104 disease analyses to systematically evaluate how enrichment analysis results were affected by evolution of the GO over a decade. We found low consistency between enrichment analyses results obtained with early and more recent GO versions. Furthermore, there continues to be a strong annotation bias in the GO annotations where 58% of the annotations are for 16% of the human genes. Our analysis suggests that GO evolution may have affected the interpretation and possibly reproducibility of experiments over time. Hence, researchers must exercise caution when interpreting GO enrichment analyses and should reexamine previous analyses with the most recent GO version.
View details for PubMedID 29572502
A community approach to mortality prediction in sepsis via gene expression analysis
2018; 9: 694
Improved risk stratification and prognosis prediction in sepsis is a critical unmet need. Clinical severity scores and available assays such as blood lactate reflect global illness severity with suboptimal performance, and do not specifically reveal the underlying dysregulation of sepsis. Here, we present prognostic models for 30-day mortality generated independently by three scientific groups by using 12 discovery cohorts containing transcriptomic data collected from primarily community-onset sepsis patients. Predictive performance is validated in five cohorts of community-onset sepsis patients in which the models show summary AUROCs ranging from 0.765-0.89. Similar performance is observed in four cohorts of hospital-acquired sepsis. Combining the new gene-expression-based prognostic models with prior clinical severity scores leads to significant improvement in prediction of 30-day mortality as measured via AUROC and net reclassification improvement index These models provide an opportunity to develop molecular bedside tests that may improve risk stratification and mortality prediction in patients with sepsis.
View details for PubMedID 29449546
Antigen Identification for Orphan T Cell Receptors Expressed on Tumor-Infiltrating Lymphocytes
2018; 172 (3): 549-+
The immune system can mount T cell responses against tumors; however, the antigen specificities of tumor-infiltrating lymphocytes (TILs) are not well understood. We used yeast-display libraries of peptide-human leukocyte antigen (pHLA) to screen for antigens of "orphan" T cell receptors (TCRs) expressed on TILs from human colorectal adenocarcinoma. Four TIL-derived TCRs exhibited strong selection for peptides presented in a highly diverse pHLA-A∗02:01 library. Three of the TIL TCRs were specific for non-mutated self-antigens, two of which were present in separate patient tumors, and shared specificity for a non-mutated self-antigen derived from U2AF2. These results show that the exposed recognition surface of MHC-bound peptides accessible to the TCR contains sufficient structural information to enable the reconstruction of sequences of peptide targets for pathogenic TCRs of unknown specificity. This finding underscores the surprising specificity of TCRs for their cognate antigens and enables the facile indentification of tumor antigens through unbiased screening.
View details for PubMedID 29275860
View details for PubMedCentralID PMC5786495
Gene annotation bias impedes biomedical research
2018; 8: 1362
We found tremendous inequality across gene and protein annotation resources. We observed that this bias leads biomedical researchers to focus on richly annotated genes instead of those with the strongest molecular data. We advocate that researchers reduce these biases by pursuing data-driven hypotheses.
View details for PubMedID 29358745
Inflammatory macrophage-associated 3-gene signature predicts subclinical allograft injury and graft survival.
2018; 3 (2)
Late allograft failure is characterized by cumulative subclinical insults manifesting over many years. Although immunomodulatory therapies targeting host T cells have improved short-term survival rates, rates of chronic allograft loss remain high. We hypothesized that other immune cell types may drive subclinical injury, ultimately leading to graft failure. We collected whole-genome transcriptome profiles from 15 independent cohorts composed of 1,697 biopsy samples to assess the association of an inflammatory macrophage polarization-specific gene signature with subclinical injury. We applied penalized regression to a subset of the data sets and identified a 3-gene inflammatory macrophage-derived signature. We validated discriminatory power of the 3-gene signature in 3 independent renal transplant data sets with mean AUC of 0.91. In a longitudinal cohort, the 3-gene signature strongly correlated with extent of injury and accurately predicted progression of subclinical injury 18 months before clinical manifestation. The 3-gene signature also stratified patients at high risk of graft failure as soon as 15 days after biopsy. We found that the 3-gene signature also distinguished acute rejection (AR) accurately in 3 heart transplant data sets but not in lung transplant. Overall, we identified a parsimonious signature capable of diagnosing AR, recognizing subclinical injury, and risk-stratifying renal transplant patients. Our results strongly suggest that inflammatory macrophages may be a viable therapeutic target to improve long-term outcomes for organ transplantation patients.
View details for PubMedID 29367465
Unsupervised Analysis of Transcriptomics in Bacterial Sepsis Across Multiple Datasets Reveals Three Robust Clusters.
Critical care medicine
To find and validate generalizable sepsis subtypes using data-driven clustering.We used advanced informatics techniques to pool data from 14 bacterial sepsis transcriptomic datasets from eight different countries (n = 700).Retrospective analysis.Persons admitted to the hospital with bacterial sepsis.None.A unified clustering analysis across 14 discovery datasets revealed three subtypes, which, based on functional analysis, we termed "Inflammopathic, Adaptive, and Coagulopathic." We then validated these subtypes in nine independent datasets from five different countries (n = 600). In both discovery and validation data, the Adaptive subtype is associated with a lower clinical severity and lower mortality rate, and the Coagulopathic subtype is associated with higher mortality and clinical coagulopathy. Further, these clusters are statistically associated with clusters derived by others in independent single sepsis cohorts.The three sepsis subtypes may represent a unifying framework for understanding the molecular heterogeneity of the sepsis syndrome. Further study could potentially enable a precision medicine approach of matching novel immunomodulatory therapies with septic patients most likely to benefit.
View details for PubMedID 29537985
A Human Genome-wide RNAi Screen Reveals Diverse Modulators that Mediate IRE1α-XBP1 Activation.
Molecular cancer research : MCR
Activation of the unfolded protein response (UPR) signaling pathways is linked to multiple human diseases including cancer. The inositol-requiring kinase 1 (IRE1)-X-box binding protein 1 (XBP1) pathway is the most evolutionarily conserved of the three major signaling branches of the UPR. Here, we performed a genome-wide siRNA screen to obtain a systematic assessment of genes integrated in the IRE1-XBP1 axis. We monitored the expression of an XBP1-luciferase chimeric protein in which luciferase was fused in-frame with the spliced (active) form of XBP1. Using cells expressing this reporter construct, we identified 162 genes for which siRNA inhibition resulted in alteration in XBP1 splicing. These genes express diverse types of proteins modulating a wide range of cellular processes. Pathway analysis identified a set of genes implicated in the pathogenesis of breast cancer. Several genes including BCL10, GCLM, and IGF1R correlated with worse relapse-free survival (RFS) in an analysis of patients with triple negative breast cancer (TNBC). However, in this cohort of 1908 patients, only high GCLM expression correlated with worse RFS in both TNBC and non-TNBC patients. Altogether, our study revealed unidentified roles of novel pathways regulating the UPR and these findings may serve as a paradigm for exploring novel therapeutic opportunities based on modulating the UPR.Genome-wide RNAi screen identifies novel genes/pathways that modulate IRE1-XBP1 signaling in human tumor cells and leads to the development of improved therapeutic approaches targeting the UPR.
View details for PubMedID 29440447
Multicohort Analysis of Whole-Blood Gene Expression Data Does Not Form a Robust Diagnostic for Acute Respiratory Distress Syndrome.
Critical care medicine
2018; 46 (2): 244–51
To identify a novel, generalizable diagnostic for acute respiratory distress syndrome using whole-blood gene expression arrays from multiple acute respiratory distress syndrome cohorts of varying etiologies.We performed a systematic search for human whole-blood gene expression arrays of acute respiratory distress syndrome in National Institutes of Health Gene Expression Omnibus and ArrayExpress. We also included the Glue Grant gene expression cohorts.We included investigator-defined acute respiratory distress syndrome within 48 hours of diagnosis and compared these with relevant critically ill controls.We used multicohort analysis of gene expression to identify genes significantly associated with acute respiratory distress syndrome, both with and without adjustment for clinical severity score. We performed gene ontology enrichment using Database for Annotation, Visualization and Integrated Discovery and cell type enrichment tests for both immune cells and pneumocyte gene expression. Finally, we selected a gene set optimized for diagnostic power across the datasets and used leave-one-dataset-out cross validation to assess robustness of the model.We identified datasets from three adult cohorts with sepsis, one pediatric cohort with acute respiratory failure, and two datasets of adult patients with trauma and burns, for a total of 148 acute respiratory distress syndrome cases and 268 critically ill controls. We identified 30 genes that were significantly associated with acute respiratory distress syndrome (false discovery rate < 20% and effect size >1.3), many of which had been previously associated with sepsis. When metaregression was used to adjust for clinical severity scores, none of these genes remained significant. Cell type enrichment was notable for bands and neutrophils, suggesting that the gene expression signature is one of acute inflammation rather than lung injury per se. Finally, an attempt to develop a generalizable diagnostic gene set for acute respiratory distress syndrome showed a mean area under the receiver-operating characteristic curve of only 0.63 on leave-one-dataset-out cross validation.The whole-blood gene expression signature across a wide clinical spectrum of acute respiratory distress syndrome is likely confounded by systemic inflammation, limiting the utility of whole-blood gene expression studies for uncovering a generalizable diagnostic gene signature.
View details for PubMedID 29337789
Pediatric Sepsis Endotypes Among Adults With Sepsis
CRITICAL CARE MEDICINE
2017; 45 (12): e1289–e1291
Recent transcriptomic studies describe two subgroups of adults with sepsis differentiated by a sepsis response signature. The implied biology and related clinical associations are comparable with recently reported pediatric sepsis endotypes, labeled "A" and "B." We classified adults with sepsis using the pediatric endotyping strategy and the sepsis response signature and determined how endotype assignment, sepsis response signature membership, and age interact with respect to mortality.Retrospective analysis of publically available transcriptomic data representing critically ill adults with sepsis from which the sepsis response signature groups were derived and validated.Multiple ICUs.Adults with sepsis.None.Transcriptomic data were conormalized into a single dataset yielding 549 unique cases with sepsis response signature assignments. Each subject was assigned to endotype A or B using the expression data for the 100 endotyping genes. There were 163 subjects (30%) assigned to endotype A and 386 to endotype B. There was a weak, positive correlation between endotype assignment and sepsis response signature membership. Mortality rates were similar between patients assigned endotype A and those assigned endotype B. A multivariable logistic regression model fit to endotype assignment, sepsis response signature membership, age, and the respective two-way interactions revealed that endotype A, sepsis response signature 1 membership, older age, and the interactions between them were associated with mortality. Subjects coassigned to endotype A, and sepsis response signature 1 had the highest mortality.Combining the pediatric endotyping strategy with sepsis response signature membership might provide complementary, age-dependent, biological, and prognostic information.
View details for PubMedID 28991828
View details for PubMedCentralID PMC5693699
Higher Baseline Monocyte Count Is Associated with More Extensive Skin Involvement and Higher Mortality in Systemic Sclerosis
View details for Web of Science ID 000411824103204
Unique transcriptomic response to sepsis is observed among patients of different age groups
2017; 12 (9): e0184159
Sepsis is a major cause of morbidity and mortality, especially at the extremes of age. To understand the human age-specific transcriptomic response to sepsis, a multi-cohort, pooled analysis was conducted on adults, children, infants, and neonates with and without sepsis. Nine public whole-blood gene expression datasets (636 patients) were employed. Age impacted the transcriptomic host response to sepsis. Gene expression from septic neonates and adults was more dissimilar whereas infants and children were more similar. Neonates showed reductions in inflammatory recognition and signaling pathways compared to all other age groups. Likewise, adults demonstrated decreased pathogen sensing, inflammation, and myeloid cell function, as compared to children. This may help to explain the increased incidence of sepsis-related organ failure and death in adults. The number of dysregulated genes in septic patients was proportional to age and significantly differed among septic adults, children, infants, and neonates. Overall, children manifested a greater transcriptomic intensity to sepsis as compared to the other age groups. The transcriptomic magnitude for adults and neonates was dramatically reduced as compared to children and infants. These findings suggest that the transcriptomic response to sepsis is age-dependent, and diagnostic and therapeutic efforts to identify and treat sepsis will have to consider age as an important variable.
View details for PubMedID 28886074
Multicohort analysis reveals baseline transcriptional predictors of influenza vaccination responses
2017; 2 (14)
View details for Web of Science ID 000434326500001
A B-Cell Gene Signature Correlates With the Extent of Gluten-Induced Intestinal Injury in Celiac Disease.
Cellular and molecular gastroenterology and hepatology
2017; 4 (1): 1-17
Celiac disease (CeD) provides an opportunity to study autoimmunity and the transition in immune cells as dietary gluten induces small intestinal lesions.Seventy-three celiac disease patients on a long-term, gluten-free diet ingested a known amount of gluten daily for 6 weeks. A peripheral blood sample and intestinal biopsy specimens were taken before and 6 weeks after initiating the gluten challenge. Biopsy results were reported on a continuous numeric scale that measured the villus-height-to-crypt-depth ratio to quantify gluten-induced intestinal injury. Pooled B and T cells were isolated from whole blood, and RNA was analyzed by DNA microarray looking for changes in peripheral B- and T-cell gene expression that correlated with changes in villus height to crypt depth, as patients maintained a relatively healthy intestinal mucosa or deteriorated in the face of a gluten challenge.Gluten-dependent intestinal damage from baseline to 6 weeks varied widely across all patients, ranging from no change to extensive damage. Genes differentially expressed in B cells correlated strongly with the extent of intestinal damage. A relative increase in B-cell gene expression correlated with a lack of sensitivity to gluten whereas their relative decrease correlated with gluten-induced mucosal injury. A core B-cell gene module, representing a subset of B-cell genes analyzed, accounted for the correlation with intestinal injury.Genes comprising the core B-cell module showed a net increase in expression from baseline to 6 weeks in patients with little to no intestinal damage, suggesting that these individuals may have mounted a B-cell immune response to maintain mucosal homeostasis and circumvent inflammation. DNA microarray data were deposited at the GEO repository (accession number: GSE87629; available: https://www.ncbi.nlm.nih.gov/geo/).
View details for DOI 10.1016/j.jcmgh.2017.01.011
View details for PubMedID 28508029
AGE-SPECIFIC TRANSCRIPTOMIC RESPONSE TO SEPSIS
LIPPINCOTT WILLIAMS & WILKINS. 2017: 92
View details for Web of Science ID 000401276500203
Validation of the Sepsis MetaScore for Diagnosis of Neonatal Sepsis.
Journal of the Pediatric Infectious Diseases Society
Neonates are at increased risk for developing sepsis, but this population often exhibits ambiguous clinical signs that complicate the diagnosis of infection. No biomarker has yet shown enough diagnostic accuracy to rule out sepsis at the time of clinical suspicion.We show that a gene-expression-based signature is an accurate objective measure of the risk of sepsis in a neonate or preterm infant, and it substantially improves diagnostic accuracy over that of commonly used laboratory-based testing. Implementation might decrease inappropriate antibiotic use.Neonatal sepsis can have devastating consequences, but accurate diagnosis is difficult. As a result, up to 200 neonates with suspected sepsis are treated with empiric antibiotics for every 1 case of microbiologically confirmed sepsis. These unnecessary antibiotics enhance bacterial antibiotic resistance, increase economic costs, and alter gut microbiota composition. We recently reported an 11-gene diagnostic test for sepsis (Sepsis MetaScore) based on host whole-blood gene expression in children and adults, but this test has not been evaluated in neonates.We identified existing gene expression microarray-based cohorts of neonates with sepsis. We then tested the accuracy of the Sepsis MetaScore both alone and in combination with standard diagnostic laboratory tests in diagnosing sepsis.We found 3 cohorts with a total of 213 samples from control neonates and neonates with sepsis. The Sepsis MetaScore had an area under the receiver operating characteristic curve of 0.92-0.93 in all 3 cohorts. We also found that, as a diagnostic test for sepsis, it outperformed standard laboratory measurements alone and, when used in combination with another test(s), resulted in a significant net reclassification index (0.3-0.69) in 5 of 6 comparisons. The mean point estimates for sensitivity and specificity were 95% and 60%, respectively, which, if confirmed prospectively and applied in a high-risk cohort, could reduce inappropriate antibiotic usage substantially.The Sepsis MetaScore had excellent diagnostic accuracy across 3 separate cohorts of neonates from 3 different countries. Further prospective targeted study will be needed before clinical application.
View details for DOI 10.1093/jpids/pix021
View details for PubMedID 28419265
Host-response biomarkers: A disease-defining diagnostic for sepsis
AMER CHEMICAL SOC. 2017
View details for Web of Science ID 000430568501270
Chemical Space Mimicry for Drug Discovery
JOURNAL OF CHEMICAL INFORMATION AND MODELING
2017; 57 (4): 875-882
We describe a new library generation method, Machine-based Identification of Molecules Inside Characterized Space (MIMICS), that generates sets of molecules inspired by a text-based input. MIMICS-generated libraries were found to preserve distributions of properties while simultaneously increasing structural diversity. Newly identified MIMICS-generated compounds were found to be bioactive as inhibitors of specific components of the unfolded protein response (UPR) and the VEGFR2 pathway in cell-based assays, thus confirming the applicability of this methodology toward drug design applications. Wider application of MIMICS could facilitate the efficient utilization of chemical space.
View details for DOI 10.1021/acs.jcim.6b00754
View details for Web of Science ID 000400204900023
View details for PubMedID 28257191
- The authors reply. Critical care medicine 2017; 45 (4): e457-e458
Gene Expression Analysis to Assess the Relevance of Rodent Models to Human Lung Injury.
American journal of respiratory cell and molecular biology
Rationale The relevance of animal models to human diseases is an area of intense scientific debate. The degree to which mouse models of lung injury recapitulate human lung injury has never been assessed. Integrating data from both human and animal expression studies allows for increased statistical power and identification of conserved differential gene expression across organisms and conditions. Objectives Comprehensive integration of gene expression data in experimental ALI in rodents compared to humans. Methods We performed two separate gene expression multi-cohort analyses to determine differential gene expression in experimental animal and human lung injury. We used correlational and pathway analyses combined with external in vitro gene expression data to identify both potential drivers of underlying inflammation and therapeutic drug candidates. Main Results We identified 21 animal lung tissue datasets and 3 human lung injury BAL datasets. We show that the meta-signatures of animal and human experimental ALI are significantly correlated despite these widely varying experimental conditions. The gene expression changes among mice and rats across diverse injury models (ozone, VILI, LPS) are significantly correlated with human models of lung injury (Pearson r 0.33-0.45, P<1e-16). Neutrophil signatures are enriched in both animal and human lung injury. Predicted therapeutic targets, peptide ligand signatures, and pathway analyses are also all highly overlapping. Conclusions Gene expression changes are similar in animal and human experimental ALI, and provide several physiologic and therapeutic insights to the disease.
View details for DOI 10.1165/rcmb.2016-0395OC
View details for PubMedID 28324666
- The authors reply. Critical care medicine 2017; 45 (3)
- Septic Cardiomyopathy: Getting to the Heart of the Matter. Critical care medicine 2017; 45 (3): 556-557
An integrative approach unveils FOSL1 as an oncogene vulnerability in KRAS-driven lung and pancreatic cancer.
2017; 8: 14294-?
KRAS mutated tumours represent a large fraction of human cancers, but the vast majority remains refractory to current clinical therapies. Thus, a deeper understanding of the molecular mechanisms triggered by KRAS oncogene may yield alternative therapeutic strategies. Here we report the identification of a common transcriptional signature across mutant KRAS cancers of distinct tissue origin that includes the transcription factor FOSL1. High FOSL1 expression identifies mutant KRAS lung and pancreatic cancer patients with the worst survival outcome. Furthermore, FOSL1 genetic inhibition is detrimental to both KRAS-driven tumour types. Mechanistically, FOSL1 links the KRAS oncogene to components of the mitotic machinery, a pathway previously postulated to function orthogonally to oncogenic KRAS. FOSL1 targets include AURKA, whose inhibition impairs viability of mutant KRAS cells. Lastly, combination of AURKA and MEK inhibitors induces a deleterious effect on mutant KRAS cells. Our findings unveil KRAS downstream effectors that provide opportunities to treat KRAS-driven cancers.
View details for DOI 10.1038/ncomms14294
View details for PubMedID 28220783
Methods to increase reproducibility in differential gene expression via meta-analysis.
Nucleic acids research
2017; 45 (1)
Findings from clinical and biological studies are often not reproducible when tested in independent cohorts. Due to the testing of a large number of hypotheses and relatively small sample sizes, results from whole-genome expression studies in particular are often not reproducible. Compared to single-study analysis, gene expression meta-analysis can improve reproducibility by integrating data from multiple studies. However, there are multiple choices in designing and carrying out a meta-analysis. Yet, clear guidelines on best practices are scarce. Here, we hypothesized that studying subsets of very large meta-analyses would allow for systematic identification of best practices to improve reproducibility. We therefore constructed three very large gene expression meta-analyses from clinical samples, and then examined meta-analyses of subsets of the datasets (all combinations of datasets with up to N/2 samples and K/2 datasets) compared to a 'silver standard' of differentially expressed genes found in the entire cohort. We tested three random-effects meta-analysis models using this procedure. We showed relatively greater reproducibility with more-stringent effect size thresholds with relaxed significance thresholds; relatively lower reproducibility when imposing extraneous constraints on residual heterogeneity; and an underestimation of actual false positive rate by Benjamini-Hochberg correction. In addition, multivariate regression showed that the accuracy of a meta-analysis increased significantly with more included datasets even when controlling for sample size.
View details for DOI 10.1093/nar/gkw797
View details for PubMedID 27634930
Benchmarking Sepsis Gene Expression Diagnostics Using Public Data
CRITICAL CARE MEDICINE
2017; 45 (1): 1-10
In response to a need for better sepsis diagnostics, several new gene expression classifiers have been recently published, including the 11-gene "Sepsis MetaScore," the "FAIM3-to-PLAC8" ratio, and the Septicyte Lab. We performed a systematic search for publicly available gene expression data in sepsis and tested each gene expression classifier in all included datasets. We also created a public repository of sepsis gene expression data to encourage their future reuse.We searched National Institutes of Health Gene Expression Omnibus and EBI ArrayExpress for human gene expression microarray datasets. We also included the Glue Grant trauma gene expression cohorts.We selected clinical, time-matched, whole blood studies of sepsis and acute infections as compared to healthy and/or noninfectious inflammation patients. We identified 39 datasets composed of 3,241 samples from 2,604 patients.All data were renormalized from raw data, when available, using consistent methods.Mean validation areas under the receiver operating characteristic curve for discriminating septic patients from patients with noninfectious inflammation for the Sepsis MetaScore, the FAIM3-to-PLAC8 ratio, and the Septicyte Lab were 0.82 (range, 0.73-0.89), 0.78 (range, 0.49-0.96), and 0.73 (range, 0.44-0.90), respectively. Paired-sample t tests of validation datasets showed no significant differences in area under the receiver operating characteristic curves. Mean validation area under the receiver operating characteristic curves for discriminating infected patients from healthy controls for the Sepsis MetaScore, FAIM3-to-PLAC8 ratio, and Septicyte Lab were 0.97 (range, 0.85-1.0), 0.94 (range, 0.65-1.0), and 0.71 (range, 0.24-1.0), respectively. There were few significant differences in any diagnostics due to pathogen type.The three diagnostics do not show significant differences in overall ability to distinguish noninfectious systemic inflammatory response syndrome from sepsis, though the performance in some datasets was low (area under the receiver operating characteristic curve, < 0.7) for the FAIM3-to-PLAC8 ratio and Septicyte Lab. The Septicyte Lab also demonstrated significantly worse performance in discriminating infections as compared to healthy controls. Overall, public gene expression data are a useful tool for benchmarking gene expression diagnostics.
View details for DOI 10.1097/CCM.0000000000002021
View details for Web of Science ID 000390619000001
View details for PubMedID 27681387
Comprehensive Analysis of the Unfolded Protein Response in Breast Cancer Subtypes
JCO PRECISION ONCOLOGY
View details for Web of Science ID 000462058200023
Generalizable Biomarkers in Critical Care: Toward Precision Medicine.
Critical care medicine
2017; 45 (6): 934–39
View details for PubMedID 28509729
Comprehensive Analysis of the Unfolded Protein Response in Breast Cancer Subtypes.
JCO precision oncology
Purpose: Triple-negative breast cancers (TNBCs) are associated with a worse prognosis and patients with TNBC have fewer therapeutic options than patients with non-TNBC. Recently, the IRE1alpha-XBP1 branch of the unfolded protein response (UPR) was implicated in TNBC prognosis on the basis of a relatively small patient population, suggesting the diagnostic and therapeutic value of this pathway in TNBCs. In addition, the IRE1alpha-XBP1 and hypoxia-induced factor 1 alpha (HIF1alpha) pathways have been identified as interacting partners in TNBC, suggesting a novel mechanism of regulation. To comprehensively evaluate and validate these findings, we investigated the relative activities and relevance to patient survival of the UPR and HIF1alpha pathways in different breast cancer subtypes in large populations of patients.Materials and Methods: We performed a comprehensive analysis of gene expression and survival data from large cohorts of patients with breast cancer. The patients were stratified based on the average expression of the UPR or HIF1alpha gene signatures.Results: We identified a strong positive association between the XBP1 gene signature and estrogen receptor-positive status or the HIF1alpha gene signature, as well as the predictive value of the XBP1 gene signature for survival of patients who are estrogen receptor negative, or have TNBC or HER2+. In contrast, another important UPR branch, the ATF4/CHOP pathway, lacks prognostic value in breast cancer in general. Activity of the HIF1alpha pathway is correlated with patient survival in all the subtypes evaluated.Conclusion: These findings clarify the relevance of the UPR pathways in different breast cancer subtypes and underscore the potential therapeutic importance of the IRE1alpha-XBP1 branch in breast cancer treatment.
View details for PubMedID 29888341
Integrated, multicohort analysis of systemic sclerosis identifies robust transcriptional signature of disease severity.
2016; 1 (21)
Systemic sclerosis (SSc) is a rare autoimmune disease with the highest case-fatality rate of all connective tissue diseases. Current efforts to determine patient response to a given treatment using the modified Rodnan skin score (mRSS) are complicated by interclinician variability, confounding, and the time required between sequential mRSS measurements to observe meaningful change. There is an unmet critical need for an objective metric of SSc disease severity. Here, we performed an integrated, multicohort analysis of SSc transcriptome data across 7 datasets from 6 centers composed of 515 samples. Using 158 skin samples from SSc patients and healthy controls recruited at 2 centers as a discovery cohort, we identified a 415-gene expression signature specific for SSc, and validated its ability to distinguish SSc patients from healthy controls in an additional 357 skin samples from 5 independent cohorts. Next, we defined the SSc skin severity score (4S). In every SSc cohort of skin biopsy samples analyzed in our study, 4S correlated significantly with mRSS, allowing objective quantification of SSc disease severity. Using transcriptome data from the largest longitudinal trial of SSc patients to date, we showed that 4S allowed us to objectively monitor individual SSc patients over time, as (a) the change in 4S of a patient is significantly correlated with change in the mRSS, and (b) the change in 4S at 12 months of treatment could predict the change in mRSS at 24 months. Our results suggest that 4S could be used to distinguish treatment responders from nonresponders prior to mRSS change. Our results demonstrate the potential clinical utility of a novel robust molecular signature and a computational approach to SSc disease severity quantification.
View details for DOI 10.1172/jci.insight.89073
View details for PubMedID 28018971
View details for PubMedCentralID PMC5161207
Complement pathway amplifies caspase-11-dependent cell death and endotoxin-induced sepsis severity.
journal of experimental medicine
2016; 213 (11): 2365-2382
Cell death and release of proinflammatory mediators contribute to mortality during sepsis. Specifically, caspase-11-dependent cell death contributes to pathology and decreases in survival time in sepsis models. Priming of the host cell, through TLR4 and interferon receptors, induces caspase-11 expression, and cytosolic LPS directly stimulates caspase-11 activation, promoting the release of proinflammatory cytokines through pyroptosis and caspase-1 activation. Using a CRISPR-Cas9-mediated genome-wide screen, we identified novel mediators of caspase-11-dependent cell death. We found a complement-related peptidase, carboxypeptidase B1 (Cpb1), to be required for caspase-11 gene expression and subsequent caspase-11-dependent cell death. Cpb1 modifies a cleavage product of C3, which binds to and activates C3aR, and then modulates innate immune signaling. We find the Cpb1-C3-C3aR pathway induces caspase-11 expression through amplification of MAPK activity downstream of TLR4 and Ifnar activation, and mediates severity of LPS-induced sepsis (endotoxemia) and disease outcome in mice. We show C3aR is required for up-regulation of caspase-11 orthologues, caspase-4 and -5, in primary human macrophages during inflammation and that c3aR1 and caspase-5 transcripts are highly expressed in patients with severe sepsis; thus, suggesting that these pathways are important in human sepsis. Our results highlight a novel role for complement and the Cpb1-C3-C3aR pathway in proinflammatory signaling, caspase-11 cell death, and sepsis severity.
View details for PubMedID 27697835
Robust classification of bacterial and viral infections via integrated host gene expression diagnostics.
Science translational medicine
2016; 8 (346): 346ra91-?
Improved diagnostics for acute infections could decrease morbidity and mortality by increasing early antibiotics for patients with bacterial infections and reducing unnecessary antibiotics for patients without bacterial infections. Several groups have used gene expression microarrays to build classifiers for acute infections, but these have been hampered by the size of the gene sets, use of overfit models, or lack of independent validation. We used multicohort analysis to derive a set of seven genes for robust discrimination of bacterial and viral infections, which we then validated in 30 independent cohorts. We next used our previously published 11-gene Sepsis MetaScore together with the new bacterial/viral classifier to build an integrated antibiotics decision model. In a pooled analysis of 1057 samples from 20 cohorts (excluding infants), the integrated antibiotics decision model had a sensitivity and specificity for bacterial infections of 94.0 and 59.8%, respectively (negative likelihood ratio, 0.10). Prospective clinical validation will be needed before these findings are implemented for patient care.
View details for DOI 10.1126/scitranslmed.aaf7165
View details for PubMedID 27384347
Integrative, multi-cohort analysis of Epstein-Barr Virus (EBV)-positive and negative tumor samples to identify gene-signatures associated with EBV oncogenesis
LIPPINCOTT WILLIAMS & WILKINS. 2016: S263
View details for Web of Science ID 000436953200409
Coordination of stress signals by the lysine methyltransferase SMYD2 promotes pancreatic cancer.
Genes & development
2016; 30 (7): 772-785
Pancreatic ductal adenocarcinoma (PDAC) is a lethal form of cancer with few therapeutic options. We found that levels of the lysine methyltransferase SMYD2 (SET and MYND domain 2) are elevated in PDAC and that genetic and pharmacological inhibition of SMYD2 restricts PDAC growth. We further identified the stress response kinase MAPKAPK3 (MK3) as a new physiologic substrate of SMYD2 in PDAC cells. Inhibition of MAPKAPK3 impedes PDAC growth, identifying a potential new kinase target in PDAC. Finally, we show that inhibition of SMYD2 cooperates with standard chemotherapy to treat PDAC cells and tumors. These findings uncover a pivotal role for SMYD2 in promoting pancreatic cancer.
View details for DOI 10.1101/gad.275529.115
View details for PubMedID 26988419
View details for PubMedCentralID PMC4826394
Genome-wide expression for diagnosis of pulmonary tuberculosis: a multicohort analysis.
The Lancet. Respiratory medicine
2016; 4 (3): 213-224
Active pulmonary tuberculosis is difficult to diagnose and treatment response is difficult to effectively monitor. A WHO consensus statement has called for new non-sputum diagnostics. The aim of this study was to use an integrated multicohort analysis of samples from publically available datasets to derive a diagnostic gene set in the peripheral blood of patients with active tuberculosis.We searched two public gene expression microarray repositories and retained datasets that examined clinical cohorts of active pulmonary tuberculosis infection in whole blood. We compared gene expression in patients with either latent tuberculosis or other diseases versus patients with active tuberculosis using our validated multicohort analysis framework. Three datasets were used as discovery datasets and meta-analytical methods were used to assess gene effects in these cohorts. We then validated the diagnostic capacity of the three gene set in the remaining 11 datasets.A total of 14 datasets containing 2572 samples from 10 countries from both adult and paediatric patients were included in the analysis. Of these, three datasets (N=1023) were used to discover a set of three genes (GBP5, DUSP3, and KLF2) that are highly diagnostic for active tuberculosis. We validated the diagnostic power of the three gene set to separate active tuberculosis from healthy controls (global area under the ROC curve (AUC) 0·90 [95% CI 0·85-0·95]), latent tuberculosis (0·88 [0·84-0·92]), and other diseases (0·84 [0·80-0·95]) in eight independent datasets composed of both children and adults from ten countries. Expression of the three-gene set was not confounded by HIV infection status, bacterial drug resistance, or BCG vaccination. Furthermore, in four additional cohorts, we showed that the tuberculosis score declined during treatment of patients with active tuberculosis.Overall, our integrated multicohort analysis yielded a three-gene set in whole blood that is robustly diagnostic for active tuberculosis, that was validated in multiple independent cohorts, and that has potential clinical application for diagnosis and monitoring treatment response. Prospective laboratory validation will be required before it can be used in a clinical setting.National Institute of Allergy and Infectious Diseases, National Library of Medicine, the Stanford Child Health Research Institute, the Society for University Surgeons, and the Bill and Melinda Gates Foundation.
View details for DOI 10.1016/S2213-2600(16)00048-5
View details for PubMedID 26907218
META-ANALYSIS OF CONTINUOUS PHENOTYPES IDENTIFIES A GENE SIGNATURE THAT CORRELATES WITH COPD DISEASE STATUS.
Pacific Symposium on Biocomputing. Pacific Symposium on Biocomputing
2016; 22: 266-275
The utility of multi-cohort two-class meta-analysis to identify robust differentially expressed gene signatures has been well established. However, many biomedical applications, such as gene signatures of disease progression, require one-class analysis. Here we describe an R package, MetaCorrelator, that can identify a reproducible transcriptional signature that is correlated with a continuous disease phenotype across multiple datasets. We successfully applied this framework to extract a pattern of gene expression that can predict lung function in patients with chronic obstructive pulmonary disease (COPD) in both peripheral blood mononuclear cells (PBMCs) and tissue. Our results point to a disregulation in the oxidation state of the lungs of patients with COPD, as well as underscore the classically recognized inammatory state that underlies this disease.
View details for PubMedID 27896981
Hospital-acquired Pneumonia: A Host of Factors.
American journal of respiratory and critical care medicine
2016; 194 (11): 1309–11
View details for PubMedID 27905845
Blood transcriptional signatures for tuberculosis diagnosis: a glass half-empty perspective - Authors' reply.
The Lancet. Respiratory medicine
2016; 4 (6): e29
View details for PubMedID 27304800
EMPOWERING MULTI-COHORT GENE EXPRESSION ANALYSIS TO INCREASE REPRODUCIBILITY.
Pacific Symposium on Biocomputing. Pacific Symposium on Biocomputing
2016; 22: 144-153
A major contributor to the scientific reproducibility crisis has been that the results from homogeneous, single-center studies do not generalize to heterogeneous, real world populations. Multi-cohort gene expression analysis has helped to increase reproducibility by aggregating data from diverse populations into a single analysis. To make the multi-cohort analysis process more feasible, we have assembled an analysis pipeline which implements rigorously studied meta-analysis best practices. We have compiled and made publicly available the results of our own multi-cohort gene expression analysis of 103 diseases, spanning 615 studies and 36,915 samples, through a novel and interactive web application. As a result, we have made both the process of and the results from multi-cohort gene expression analysis more approachable for non-technical users.
View details for PubMedID 27896970
Integrated, Multi-cohort Analysis Identifies Conserved Transcriptional Signatures across Multiple Respiratory Viruses
2015; 43 (6): 1199-1211
Respiratory viral infections are a significant burden to healthcare worldwide. Many whole genome expression profiles have identified different respiratory viral infection signatures, but these have not translated to clinical practice. Here, we performed two integrated, multi-cohort analyses of publicly available transcriptional data of viral infections. First, we identified a common host signature across different respiratory viral infections that could distinguish (1) individuals with viral infections from healthy controls and from those with bacterial infections, and (2) symptomatic from asymptomatic subjects prior to symptom onset in challenge studies. Second, we identified an influenza-specific host response signature that (1) could distinguish influenza-infected samples from those with bacterial and other respiratory viral infections, (2) was a diagnostic and prognostic marker in influenza-pneumonia patients and influenza challenge studies, and (3) was predictive of response to influenza vaccine. Our results have applications in the diagnosis, prognosis, and identification of drug targets in viral infections.
View details for DOI 10.1016/j.immuni.2015.11.003
View details for Web of Science ID 000366846600022
View details for PubMedID 26682989
View details for PubMedCentralID PMC4684904
- Comprehensive Validation of the FAIM3:PLAC8 Ratio in Time-matched Public Gene Expression Data. American journal of respiratory and critical care medicine 2015; 192 (10): 1260-1261
The center for expanded data annotation and retrieval.
Journal of the American Medical Informatics Association
2015; 22 (6): 1148-1152
The Center for Expanded Data Annotation and Retrieval is studying the creation of comprehensive and expressive metadata for biomedical datasets to facilitate data discovery, data interpretation, and data reuse. We take advantage of emerging community-based standard templates for describing different kinds of biomedical datasets, and we investigate the use of computational techniques to help investigators to assemble templates and to fill in their values. We are creating a repository of metadata from which we plan to identify metadata patterns that will drive predictive data entry when filling in metadata templates. The metadata repository not only will capture annotations specified when experimental datasets are initially created, but also will incorporate links to the published literature, including secondary analyses and possible refinements or retractions of experimental interpretations. By working initially with the Human Immunology Project Consortium and the developers of the ImmPort data repository, we are developing and evaluating an end-to-end solution to the problems of metadata authoring and management that will generalize to other data-management environments.
View details for DOI 10.1093/jamia/ocv048
View details for PubMedID 26112029
Combined inhibition of BET family proteins and histone deacetylases as a potential epigenetics-based therapy for pancreatic ductal adenocarcinoma.
2015; 21 (10): 1163-1171
Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal human cancers and shows resistance to any therapeutic strategy used. Here we tested small-molecule inhibitors targeting chromatin regulators as possible therapeutic agents in PDAC. We show that JQ1, an inhibitor of the bromodomain and extraterminal (BET) family of proteins, suppresses PDAC development in mice by inhibiting both MYC activity and inflammatory signals. The histone deacetylase (HDAC) inhibitor SAHA synergizes with JQ1 to augment cell death and more potently suppress advanced PDAC. Finally, using a CRISPR-Cas9-based method for gene editing directly in the mouse adult pancreas, we show that de-repression of p57 (also known as KIP2 or CDKN1C) upon combined BET and HDAC inhibition is required for the induction of combination therapy-induced cell death in PDAC. SAHA is approved for human use, and molecules similar to JQ1 are being tested in clinical trials. Thus, these studies identify a promising epigenetic-based therapeutic strategy that may be rapidly implemented in fatal human tumors.
View details for DOI 10.1038/nm.3952
View details for PubMedID 26390243
- SMYD3 links methylation of MAP3K2 to Ras-driven tumors AMER ASSOC CANCER RESEARCH. 2015
A comprehensive time-course-based multicohort analysis of sepsis and sterile inflammation reveals a robust diagnostic gene set
SCIENCE TRANSLATIONAL MEDICINE
2015; 7 (287)
Although several dozen studies of gene expression in sepsis have been published, distinguishing sepsis from a sterile systemic inflammatory response syndrome (SIRS) is still largely up to clinical suspicion. We hypothesized that a multicohort analysis of the publicly available sepsis gene expression data sets would yield a robust set of genes for distinguishing patients with sepsis from patients with sterile inflammation. A comprehensive search for gene expression data sets in sepsis identified 27 data sets matching our inclusion criteria. Five data sets (n = 663 samples) compared patients with sterile inflammation (SIRS/trauma) to time-matched patients with infections. We applied our multicohort analysis framework that uses both effect sizes and P values in a leave-one-data set-out fashion to these data sets. We identified 11 genes that were differentially expressed (false discovery rate ≤1%, inter-data set heterogeneity P > 0.01, summary effect size >1.5-fold) across all discovery cohorts with excellent diagnostic power [mean area under the receiver operating characteristic curve (AUC), 0.87; range, 0.7 to 0.98]. We then validated these 11 genes in 15 independent cohorts comparing (i) time-matched infected versus noninfected trauma patients (4 cohorts), (ii) ICU/trauma patients with infections over the clinical time course (3 cohorts), and (iii) healthy subjects versus sepsis patients (8 cohorts). In the discovery Glue Grant cohort, SIRS plus the 11-gene set improved prediction of infection (compared to SIRS alone) with a continuous net reclassification index of 0.90. Overall, multicohort analysis of time-matched cohorts yielded 11 genes that robustly distinguish sterile inflammation from infectious inflammation.
View details for DOI 10.1126/scitranslmed.aaa5993
View details for Web of Science ID 000354433900010
View details for PubMedID 25972003
A COMPREHENSIVE TIME-BASED META-ANALYSIS OF SIRS AND SEPSIS REVEALS A ROBUST DISCRIMINATORY GENE SET
44th Critical Care Congress of the Society-of-Critical-Care-Medicine
LIPPINCOTT WILLIAMS & WILKINS. 2014
View details for Web of Science ID 000346211801253
- A drug repositioning approach identifies tricyclic antidepressants as inhibitors of small cell lung cancer and other neuroendocrine tumors AMER ASSOC CANCER RESEARCH. 2014
SMYD3 links lysine methylation of MAP3K2 to Ras-driven cancer.
2014; 510 (7504): 283-287
Deregulation of lysine methylation signalling has emerged as a common aetiological factor in cancer pathogenesis, with inhibitors of several histone lysine methyltransferases (KMTs) being developed as chemotherapeutics. The largely cytoplasmic KMT SMYD3 (SET and MYND domain containing protein 3) is overexpressed in numerous human tumours. However, the molecular mechanism by which SMYD3 regulates cancer pathways and its relationship to tumorigenesis in vivo are largely unknown. Here we show that methylation of MAP3K2 by SMYD3 increases MAP kinase signalling and promotes the formation of Ras-driven carcinomas. Using mouse models for pancreatic ductal adenocarcinoma and lung adenocarcinoma, we found that abrogating SMYD3 catalytic activity inhibits tumour development in response to oncogenic Ras. We used protein array technology to identify the MAP3K2 kinase as a target of SMYD3. In cancer cell lines, SMYD3-mediated methylation of MAP3K2 at lysine 260 potentiates activation of the Ras/Raf/MEK/ERK signalling module and SMYD3 depletion synergizes with a MEK inhibitor to block Ras-driven tumorigenesis. Finally, the PP2A phosphatase complex, a key negative regulator of the MAP kinase pathway, binds to MAP3K2 and this interaction is blocked by methylation. Together, our results elucidate a new role for lysine methylation in integrating cytoplasmic kinase-signalling cascades and establish a pivotal role for SMYD3 in the regulation of oncogenic Ras signalling.
View details for DOI 10.1038/nature13320
View details for PubMedID 24847881
A Meta-analysis of Lung Cancer Gene Expression Identifies PTK7 as a Survival Gene in Lung Adenocarcinoma.
2014; 74 (10): 2892-2902
Lung cancer remains the most common cause of cancer-related death worldwide and it continues to lack effective treatment. The increasingly large and diverse public databases of lung cancer gene expression constitute a rich source of candidate oncogenic drivers and therapeutic targets. To define novel targets for lung adenocarcinoma, we conducted a large-scale meta-analysis of genes specifically overexpressed in adenocarcinoma. We identified an 11-gene signature that was overexpressed consistently in adenocarcinoma specimens relative to normal lung tissue. Six genes in this signature were specifically overexpressed in adenocarcinoma relative to other subtypes of non-small cell lung cancer (NSCLC). Among these genes was the little studied protein tyrosine kinase PTK7. Immunohistochemical analysis confirmed that PTK7 is highly expressed in primary adenocarcinoma patient samples. RNA interference-mediated attenuation of PTK7 decreased cell viability and increased apoptosis in a subset of adenocarcinoma cell lines. Further, loss of PTK7 activated the MKK7-JNK stress response pathway and impaired tumor growth in xenotransplantation assays. Our work defines PTK7 as a highly and specifically expressed gene in adenocarcinoma and a potential therapeutic target in this subset of NSCLC. Cancer Res; 74(10); 2892-902. ©2014 AACR.
View details for DOI 10.1158/0008-5472.CAN-13-2775
View details for PubMedID 24654231
- A drug repositioning approach identifies tricyclic antidepressants as inhibitors of small cell lung cancer. AMER ASSOC CANCER RESEARCH. 2014
Multiplex meta-analysis of medulloblastoma expression studies with external controls.
Pacific Symposium on Biocomputing. Pacific Symposium on Biocomputing
We propose and discuss a method for doing gene expression meta-analysis (multiple datasets) across multiplex measurement modalities measuring the expression of many genes simultaneously (e.g. microarrays and RNAseq) using external control samples and a method of heterogeneity detection to identify and filter on comparable gene expression measurements. We demonstrate this approach on publicly available gene expression datasets from samples of medulloblastoma and normal cerebellar tissue and identify some potential new targets in the treatment of medulloblastoma.
View details for PubMedID 24297537
Integrated multi-cohort transcriptional meta-analysis of neurodegenerative diseases.
Acta neuropathologica communications
2014; 2: 93-?
Neurodegenerative diseases share common pathologic features including neuroinflammation, mitochondrial dysfunction and protein aggregation, suggesting common underlying mechanisms of neurodegeneration. We undertook a meta-analysis of public gene expression data for neurodegenerative diseases to identify a common transcriptional signature of neurodegeneration.Using 1,270 post-mortem central nervous system tissue samples from 13 patient cohorts covering four neurodegenerative diseases, we identified 243 differentially expressed genes, which were similarly dysregulated in 15 additional patient cohorts of 205 samples including seven neurodegenerative diseases. This gene signature correlated with histologic disease severity. Metallothioneins featured prominently among differentially expressed genes, and functional pathway analysis identified specific convergent themes of dysregulation. MetaCore network analyses revealed various novel candidate hub genes (e.g. STAU2). Genes associated with M1-polarized macrophages and reactive astrocytes were strongly enriched in the meta-analysis data. Evaluation of genes enriched in neurons revealed 70 down-regulated genes, over half not previously associated with neurodegeneration. Comparison with aging brain data (3 patient cohorts, 221 samples) revealed 53 of these to be unique to neurodegenerative disease, many of which are strong candidates to be important in neuropathogenesis (e.g. NDN, NAP1L2). ENCODE ChIP-seq analysis predicted common upstream transcriptional regulators not associated with normal aging (REST, RBBP5, SIN3A, SP2, YY1, ZNF143, IKZF1). Finally, we removed genes common to neurodegeneration from disease-specific gene signatures, revealing uniquely robust immune response and JAK-STAT signaling in amyotrophic lateral sclerosis.Our results implicate pervasive bioenergetic deficits, M1-type microglial activation and gliosis as unifying themes of neurodegeneration, and identify numerous novel genes associated with neurodegenerative processes.
View details for DOI 10.1186/s40478-014-0093-y
View details for PubMedID 25187168
View details for PubMedCentralID PMC4167139
A Drug Repositioning Approach Identifies Tricyclic Antidepressants as Inhibitors of Small Cell Lung Cancer and Other Neuroendocrine Tumors
2013; 3 (12): 1364-1377
Small cell lung cancer (SCLC) is an aggressive neuroendocrine subtype of lung cancer with high mortality. We used a systematic drug repositioning bioinformatics approach querying a large compendium of gene expression profiles to identify candidate U.S. Food and Drug Administration (FDA)-approved drugs to treat SCLC. We found that tricyclic antidepressants and related molecules potently induce apoptosis in both chemonaïve and chemoresistant SCLC cells in culture, in mouse and human SCLC tumors transplanted into immunocompromised mice, and in endogenous tumors from a mouse model for human SCLC. The candidate drugs activate stress pathways and induce cell death in SCLC cells, at least in part by disrupting autocrine survival signals involving neurotransmitters and their G protein-coupled receptors. The candidate drugs inhibit the growth of other neuroendocrine tumors, including pancreatic neuroendocrine tumors and Merkel cell carcinoma. These experiments identify novel targeted strategies that can be rapidly evaluated in patients with neuroendocrine tumors through the repurposing of approved drugs.Our work shows the power of bioinformatics-based drug approaches to rapidly repurpose FDA-approved drugs and identifies a novel class of molecules to treat patients with SCLC, a cancer for which no effective novel systemic treatments have been identified in several decades. In addition, our experiments highlight the importance of novel autocrine mechanisms in promoting the growth of neuroendocrine tumor cells.
View details for DOI 10.1158/2159-8290.CD-13-0183
View details for Web of Science ID 000328257500023
View details for PubMedID 24078773
View details for PubMedCentralID PMC3864571
A common rejection module (CRM) for acute rejection across multiple organs identifies novel therapeutics for organ transplantation.
journal of experimental medicine
2013; 210 (11): 2205-2221
Using meta-analysis of eight independent transplant datasets (236 graft biopsy samples) from four organs, we identified a common rejection module (CRM) consisting of 11 genes that were significantly overexpressed in acute rejection (AR) across all transplanted organs. The CRM genes could diagnose AR with high specificity and sensitivity in three additional independent cohorts (794 samples). In another two independent cohorts (151 renal transplant biopsies), the CRM genes correlated with the extent of graft injury and predicted future injury to a graft using protocol biopsies. Inferred drug mechanisms from the literature suggested that two FDA-approved drugs (atorvastatin and dasatinib), approved for nontransplant indications, could regulate specific CRM genes and reduce the number of graft-infiltrating cells during AR. We treated mice with HLA-mismatched mouse cardiac transplant with atorvastatin and dasatinib and showed reduction of the CRM genes, significant reduction of graft-infiltrating cells, and extended graft survival. We further validated the beneficial effect of atorvastatin on graft survival by retrospective analysis of electronic medical records of a single-center cohort of 2,515 renal transplant patients followed for up to 22 yr. In conclusion, we identified a CRM in transplantation that provides new opportunities for diagnosis, drug repositioning, and rational drug design.
View details for DOI 10.1084/jem.20122709
View details for PubMedID 24127489
View details for PubMedCentralID PMC3804941
- Identification of novel biomarkers for early detection of ovarian cancer AMER ASSOC CANCER RESEARCH. 2013
Cross-Species Functional Analysis of Cancer-Associated Fibroblasts Identifies a Critical Role for CLCF1 and IL-6 in Non-Small Cell Lung Cancer In Vivo
2012; 72 (22): 5744-5756
Cancer-associated fibroblasts (CAF) have been reported to support tumor progression by a variety of mechanisms. However, their role in the progression of non-small cell lung cancer (NSCLC) remains poorly defined. In addition, the extent to which specific proteins secreted by CAFs contribute directly to tumor growth is unclear. To study the role of CAFs in NSCLCs, a cross-species functional characterization of mouse and human lung CAFs was conducted. CAFs supported the growth of lung cancer cells in vivo by secretion of soluble factors that directly stimulate the growth of tumor cells. Gene expression analysis comparing normal mouse lung fibroblasts and mouse lung CAFs identified multiple genes that correlate with the CAF phenotype. A gene signature of secreted genes upregulated in CAFs was an independent marker of poor survival in patients with NSCLC. This secreted gene signature was upregulated in normal lung fibroblasts after long-term exposure to tumor cells, showing that lung fibroblasts are "educated" by tumor cells to acquire a CAF-like phenotype. Functional studies identified important roles for CLCF1-CNTFR and interleukin (IL)-6-IL-6R signaling in promoting growth of NSCLCs. This study identifies novel soluble factors contributing to the CAF protumorigenic phenotype in NSCLCs and suggests new avenues for the development of therapeutic strategies.
View details for DOI 10.1158/0008-5472.CAN-12-1097
View details for PubMedID 22962265
A Peripheral Blood Diagnostic Test for Acute Rejection in Renal Transplantation
AMERICAN JOURNAL OF TRANSPLANTATION
2012; 12 (10): 2710-2718
Monitoring of renal graft status through peripheral blood (PB) rather than invasive biopsy is important as it will lessen the risk of infection and other stresses, while reducing the costs of rejection diagnosis. Blood gene biomarker panels were discovered by microarrays at a single center and subsequently validated and cross-validated by QPCR in the NIH SNSO1 randomized study from 12 US pediatric transplant programs. A total of 367 unique human PB samples, each paired with a graft biopsy for centralized, blinded phenotype classification, were analyzed (115 acute rejection (AR), 180 stable and 72 other causes of graft injury). Of the differentially expressed genes by microarray, Q-PCR analysis of a five gene-set (DUSP1, PBEF1, PSEN1, MAPK9 and NKTR) classified AR with high accuracy. A logistic regression model was built on independent training-set (n = 47) and validated on independent test-set (n = 198)samples, discriminating AR from STA with 91% sensitivity and 94% specificity and AR from all other non-AR phenotypes with 91% sensitivity and 90% specificity. The 5-gene set can diagnose AR potentially avoiding the need for invasive renal biopsy. These data support the conduct of a prospective study to validate the clinical predictive utility of this diagnostic tool.
View details for DOI 10.1111/j.1600-6143.2012.04253.x
View details for Web of Science ID 000309180000018
View details for PubMedID 23009139
Non-HLA Antibodies to Immunogenic Epitopes Predict the Evolution of Chronic Renal Allograft Injury
JOURNAL OF THE AMERICAN SOCIETY OF NEPHROLOGY
2012; 23 (4): 750-763
Chronic allograft injury (CAI) results from a humoral response to mismatches in immunogenic epitopes between the donor and recipient. Although alloantibodies against HLA antigens contribute to the pathogenesis of CAI, alloantibodies against non-HLA antigens likely contribute as well. Here, we used high-density protein arrays to identify non-HLA antibodies in CAI and subsequently validated a subset in a cohort of 172 serum samples collected serially post-transplantation. There were 38 de novo non-HLA antibodies that significantly associated with the development of CAI (P<0.01) on protocol post-transplant biopsies, with enrichment of their corresponding antigens in the renal cortex. Baseline levels of preformed antibodies to MIG (also called CXCL9), ITAC (also called CXCL11), IFN-γ, and glial-derived neurotrophic factor positively correlated with histologic injury at 24 months. Measuring levels of these four antibodies could help clinicians predict the development of CAI with >80% sensitivity and 100% specificity. In conclusion, pretransplant serum levels of a defined panel of alloantibodies targeting non-HLA immunogenic antigens associate with histologic CAI in the post-transplant period. Validation in a larger, prospective transplant cohort may lead to a noninvasive method to predict and monitor for CAI.
View details for DOI 10.1681/ASN.2011060596
View details for Web of Science ID 000302333300022
View details for PubMedID 22302197
Ten Years of Pathway Analysis: Current Approaches and Outstanding Challenges
PLOS COMPUTATIONAL BIOLOGY
2012; 8 (2)
Pathway analysis has become the first choice for gaining insight into the underlying biology of differentially expressed genes and proteins, as it reduces complexity and has increased explanatory power. We discuss the evolution of knowledge base-driven pathway analysis over its first decade, distinctly divided into three generations. We also discuss the limitations that are specific to each generation, and how they are addressed by successive generations of methods. We identify a number of annotation challenges that must be addressed to enable development of the next generation of pathway analysis methods. Furthermore, we identify a number of methodological challenges that the next generation of methods must tackle to take advantage of the technological advances in genomics and proteomics in order to improve specificity, sensitivity, and relevance of pathway analysis.
View details for DOI 10.1371/journal.pcbi.1002375
View details for Web of Science ID 000300729900019
View details for PubMedID 22383865
View details for PubMedCentralID PMC3285573
Profiling of Autoantibodies in IgA Nephropathy, an Integrative Antibiomics Approach
CLINICAL JOURNAL OF THE AMERICAN SOCIETY OF NEPHROLOGY
2011; 6 (12): 2775-2784
IgG commonly co-exists with IgA in the glomerular mesangium of patients with IgA nephropathy (IgAN) with unclear clinical relevance. Autoantibody (autoAb) biomarkers to detect and track progression of IgAN are an unmet clinical need. The objective of the study was to identify IgA-specific autoAbs specific to IgAN.High-density protein microarrays were evaluated IgG autoAbs in the serum of IgAN patients (n = 22) and controls (n = 10). Clinical parameters, including annual GFR and urine protein measurements, were collected on all patients over 5 years. Bioinformatic data analysis was performed to select targets for further validation by immunohistochemistry (IHC).One hundred seventeen (1.4%) specific antibodies were increased in IgAN. Among the most significant were the autoAb to the Ig family of proteins. IgAN-specific autoAbs (approximately 50%) were mounted against proteins predominantly expressed in glomeruli and tubules, and selected candidates were verified by IHC. Receiver operating characteristic analysis of our study demonstrated that IgG autoAb levels (matriline 2, ubiquitin-conjugating enzyme E2W, DEAD box protein, and protein kinase D1) might be used in combination with 24-hour proteinuria to improve prediction of the progression of IgAN (area under the curve = 0.86, P = 0.02).IgAN is associated with elevated IgG autoAbs to multiple proteins in the kidney. This first analysis of the repertoire of autoAbs in IgAN identifies novel, immunogenic protein targets that are highly expressed in the kidney glomerulus and tubules that may bear relevance in the pathogenesis and progression of IgAN.
View details for DOI 10.2215/CJN.04600511
View details for Web of Science ID 000297948900009
View details for PubMedID 22157707
View details for PubMedCentralID PMC3255376
Progressive histological damage in renal allografts is associated with expression of innate and adaptive immunity genes
2011; 80 (12): 1364-1376
The degree of progressive chronic histological damage is associated with long-term renal allograft survival. In order to identify promising molecular targets for timely intervention, we examined renal allograft protocol and indication biopsies from 120 low-risk pediatric and adolescent recipients by whole-genome microarray expression profiling. In data-driven analysis, we found a highly regulated pattern of adaptive and innate immune gene expression that correlated with established or ongoing histological chronic injury, and also with development of future chronic histological damage, even in histologically pristine kidneys. Hence, histologically unrecognized immunological injury at a molecular level sets the stage for the development of chronic tissue injury, while the same molecular response is accentuated during established and worsening chronic allograft damage. Irrespective of the hypothesized immune or nonimmune trigger for chronic allograft injury, a highly orchestrated regulation of innate and adaptive immune responses was found in the graft at the molecular level. This occurred months before histologic lesions appear, and quantitatively below the diagnostic threshold of classic T-cell or antibody-mediated rejection. Thus, measurement of specific immune gene expression in protocol biopsies may be warranted to predict the development of subsequent chronic injury in histologically quiescent grafts and as a means to titrate immunosuppressive therapy.
View details for DOI 10.1038/ki.2011.245
View details for Web of Science ID 000297541900014
View details for PubMedID 21881554
- Applications of Translational Bioinformatics in Transplantation CLINICAL PHARMACOLOGY & THERAPEUTICS 2011; 90 (2): 323-327
Biomarkers in solid organ transplantation: establishing personalized transplantation medicine.
2011; 3 (6): 37-?
Technological advances in molecular and in silico research have enabled significant progress towards personalized transplantation medicine. It is now possible to conduct comprehensive biomarker development studies of transplant organ pathologies, correlating genomic, transcriptomic and proteomic information from donor and recipient with clinical and histological phenotypes. Translation of these advances to the clinical setting will allow assessment of an individual patient's risk of allograft damage or accommodation. Transplantation biomarkers are needed for active monitoring of immunosuppression, to reduce patient morbidity, and to improve long-term allograft function and life expectancy. Here, we highlight recent pre- and post-transplantation biomarkers of acute and chronic allograft damage or adaptation, focusing on peripheral blood-based methodologies for non-invasive application. We then critically discuss current findings with respect to their future application in routine clinical transplantation medicine. Complement-system-associated SNPs present potential biomarkers that may be used to indicate the baseline risk for allograft damage prior to transplantation. The detection of antibodies against novel, non-HLA, MICA antigens, and the expression of cytokine genes and proteins and cytotoxicity-related genes have been correlated with allograft damage and are potential post-transplantation biomarkers indicating allograft damage at the molecular level, although these do not have clinical relevance yet. Several multi-gene expression-based biomarker panels have been identified that accurately predicted graft accommodation in liver transplant recipients and may be developed into a predictive biomarker assay.
View details for DOI 10.1186/gm253
View details for PubMedID 21658299
View details for PubMedCentralID PMC3218811
- Functional Pathway Analysis for Understanding Immunologic Signature of Rejection: Current Approaches and Outstanding Challenges IMMUNOLOGIC SIGNATURES OF REJECTION 2011: 239-256
Comparison of multiplex meta analysis techniques for understanding the acute rejection of solid organ transplants
AMIA Summit on Translational Bioinformatics
BIOMED CENTRAL LTD. 2010
Combining the results of studies using highly parallelized measurements of gene expression such as microarrays and RNAseq offer unique challenges in meta analysis. Motivated by a need for a deeper understanding of organ transplant rejection, we combine the data from five separate studies to compare acute rejection versus stability after solid organ transplantation, and use this data to examine approaches to multiplex meta analysis.We demonstrate that a commonly used parametric effect size estimate approach and a commonly used non-parametric method give very different results in prioritizing genes. The parametric method providing a meta effect estimate was superior at ranking genes based on our gold-standard of identifying immune response genes in the transplant rejection datasets.Different methods of multiplex analysis can give substantially different results. The method which is best for any given application will likely depend on the particular domain, and it remains for future work to see if any one method is consistently better at identifying important biological signal across gene expression experiments.
View details for Web of Science ID 000290218700006
View details for PubMedID 21044364
View details for PubMedCentralID PMC2967747
A microarray analysis of the effects of moderate hypothermia and rewarming on gene expression by human hepatocytes (HepG2)
CELL STRESS & CHAPERONES
2010; 15 (5): 687-702
The gene expression changes produced by moderate hypothermia are not fully known, but appear to differ in important ways from those produced by heat shock. We examined the gene expression changes produced by moderate hypothermia and tested the hypothesis that rewarming after hypothermia approximates a heat-shock response. Six sets of human HepG2 hepatocytes were subjected to moderate hypothermia (31 degrees C for 16 h), a conventional in vitro heat shock (43 degrees C for 30 min) or control conditions (37 degrees C), then harvested immediately or allowed to recover for 3 h at 37 degrees C. Expression analysis was performed with Affymetrix U133A gene chips, using analysis of variance-based techniques. Moderate hypothermia led to distinct time-dependent expression changes, as did heat shock. Hypothermia initially caused statistically significant, greater than or equal to twofold changes in expression (relative to controls) of 409 sequences (143 increased and 266 decreased), whereas heat shock affected 71 (35 increased and 36 decreased). After 3 h of recovery, 192 sequences (83 increased, 109 decreased) were affected by hypothermia and 231 (146 increased, 85 decreased) by heat shock. Expression of many heat shock proteins was decreased by hypothermia but significantly increased after rewarming. A comparison of sequences affected by thermal stress without regard to the magnitude of change revealed that the overlap between heat and cold stress was greater after 3 h of recovery than immediately following thermal stress. Thus, while some overlap occurs (particularly after rewarming), moderate hypothermia produces extensive, time-dependent gene expression changes in HepG2 cells that differ in important ways from those induced by heat shock.
View details for DOI 10.1007/s12192-010-0181-2
View details for Web of Science ID 000280781800021
View details for PubMedID 20526826
View details for PubMedCentralID PMC3006613
Cell type-specific gene expression differences in complex tissues
2010; 7 (4): 287-289
We describe cell type-specific significance analysis of microarrays (csSAM) for analyzing differential gene expression for each cell type in a biological sample from microarray data and relative cell-type frequencies. First, we validated csSAM with predesigned mixtures and then applied it to whole-blood gene expression datasets from stable post-transplant kidney transplant recipients and those experiencing acute transplant rejection, which revealed hundreds of differentially expressed genes that were otherwise undetectable.
View details for DOI 10.1038/NMETH.1439
View details for Web of Science ID 000276150600017
View details for PubMedID 20208531
Extracting cell-type-specific gene expression differences from complex tissues
AMER ASSOC IMMUNOLOGISTS. 2010
View details for Web of Science ID 000209758304226
Meta-Analysis of Solid Organ Transplant Data Sets Identifies Differentially Expressed microRNAs Common in Heart, Kidney and Liver Allografts
10th American Transplant Congress
WILEY-BLACKWELL. 2010: 61–61
View details for Web of Science ID 000275921701073
NetPath: a public resource of curated signal transduction pathways
2010; 11 (1)
We have developed NetPath as a resource of curated human signaling pathways. As an initial step, NetPath provides detailed maps of a number of immune signaling pathways, which include approximately 1,600 reactions annotated from the literature and more than 2,800 instances of transcriptionally regulated genes - all linked to over 5,500 published articles. We anticipate NetPath to become a consolidated resource for human signaling pathways that should enable systems biology approaches.
View details for DOI 10.1186/gb-2010-11-1-r3
View details for Web of Science ID 000276433600011
View details for PubMedID 20067622
View details for PubMedCentralID PMC2847715
Predicting Novel Human Gene Ontology Annotations Using Semantic Analysis
IEEE-ACM TRANSACTIONS ON COMPUTATIONAL BIOLOGY AND BIOINFORMATICS
2010; 7 (1): 91-99
The correct interpretation of many molecular biology experiments depends in an essential way on the accuracy and consistency of the existing annotation databases. Such databases are meant to act as repositories for our biological knowledge as we acquire and refine it. Hence, by definition, they are incomplete at any given time. In this paper, we describe a technique that improves our previous method for predicting novel GO annotations by extracting implicit semantic relationships between genes and functions. In this work, we use a vector space model and a number of weighting schemes in addition to our previous latent semantic indexing approach. The technique described here is able to take into consideration the hierarchical structure of the Gene Ontology (GO) and can weight differently GO terms situated at different depths. The prediction abilities of 15 different weighting schemes are compared and evaluated. Nine such schemes were previously used in other problem domains, while six of them are introduced in this paper. The best weighting scheme was a novel scheme, n2tn. Out of the top 50 functional annotations predicted using this weighting scheme, we found support in the literature for 84 percent of them, while 6 percent of the predictions were contradicted by the existing literature. For the remaining 10 percent, we did not find any relevant publications to confirm or contradict the predictions. The n2tn weighting scheme also outperformed the simple binary scheme used in our previous approach.
View details for DOI 10.1109/TCBB.2008.29
View details for Web of Science ID 000274063600008
View details for PubMedID 20150671
View details for PubMedCentralID PMC3712327
Using gene arrays in diagnosis of rejection
CURRENT OPINION IN ORGAN TRANSPLANTATION
2009; 14 (1): 34-39
In the last decade, microarray technology has revolutionized biological research by allowing the screening of tens of thousands of genes simultaneously. This article reviews recent studies in organ transplantation using microarrays and highlights the issues that should be addressed in order to use microarrays in diagnosis of rejection.Microarrays have been useful in identifying potential biomarkers for chronic rejection in peripheral blood mononuclear cells, novel pathways for induction of tolerance, and genes involved in protecting the graft from the host immune system. Microarray analysis of peripheral blood mononuclear cells from chronic antibody-mediated rejection has identified potential noninvasive biomarkers. In a recent study, correlation of pathogenesis-based transcripts with histopathologic lesions is a promising step towards inclusion of microarrays in clinics for organ transplants.Despite promising results in diagnosis of histopathologic lesions using microarrays, the low dynamic range of microarrays and large measured expression changes within the probes for the same gene continue to cast doubts on their readiness for diagnosis of rejection. More studies must be performed to resolve these issues. Dominating expression of globin genes in whole blood poses another challenge for identification of noninvasive biomarkers. In addition, studies are also needed to demonstrate effects of different immunosuppression therapies and their outcomes.
View details for DOI 10.1097/MOT.0b013e32831e13d0
View details for Web of Science ID 000264312900007
View details for PubMedID 19337144
A novel signaling pathway impact analysis
2009; 25 (1): 75-82
Gene expression class comparison studies may identify hundreds or thousands of genes as differentially expressed (DE) between sample groups. Gaining biological insight from the result of such experiments can be approached, for instance, by identifying the signaling pathways impacted by the observed changes. Most of the existing pathway analysis methods focus on either the number of DE genes observed in a given pathway (enrichment analysis methods), or on the correlation between the pathway genes and the class of the samples (functional class scoring methods). Both approaches treat the pathways as simple sets of genes, disregarding the complex gene interactions that these pathways are built to describe.We describe a novel signaling pathway impact analysis (SPIA) that combines the evidence obtained from the classical enrichment analysis with a novel type of evidence, which measures the actual perturbation on a given pathway under a given condition. A bootstrap procedure is used to assess the significance of the observed total pathway perturbation. Using simulations we show that the evidence derived from perturbations is independent of the pathway enrichment evidence. This allows us to calculate a global pathway significance P-value, which combines the enrichment and perturbation P-values. We illustrate the capabilities of the novel method on four real datasets. The results obtained on these data show that SPIA has better specificity and more sensitivity than several widely used pathway analysis methods.SPIA was implemented as an R package available at http://vortex.cs.wayne.edu/ontoexpress/
View details for DOI 10.1093/bioinformatics/btn577
View details for Web of Science ID 000261996400012
View details for PubMedID 18990722
View details for PubMedCentralID PMC2732297
Identifying Uncertainty Regions in Support Vector Machines using Geometric Margin and Convex Hulls
2008 IEEE INTERNATIONAL JOINT CONFERENCE ON NEURAL NETWORKS, VOLS 1-8
View details for Web of Science ID 000263827202029
A systems biology approach for pathway level analysis
2007; 17 (10): 1537-1545
A common challenge in the analysis of genomics data is trying to understand the underlying phenomenon in the context of all complex interactions taking place on various signaling pathways. A statistical approach using various models is universally used to identify the most relevant pathways in a given experiment. Here, we show that the existing pathway analysis methods fail to take into consideration important biological aspects and may provide incorrect results in certain situations. By using a systems biology approach, we developed an impact analysis that includes the classical statistics but also considers other crucial factors such as the magnitude of each gene's expression change, their type and position in the given pathways, their interactions, etc. The impact analysis is an attempt to a deeper level of statistical analysis, informed by more pathway-specific biology than the existing techniques. On several illustrative data sets, the classical analysis produces both false positives and false negatives, while the impact analysis provides biologically meaningful results. This analysis method has been implemented as a Web-based tool, Pathway-Express, freely available as part of the Onto-Tools (http://vortex.cs.wayne.edu).
View details for DOI 10.1101/gr.6202607
View details for Web of Science ID 000249869200015
View details for PubMedID 17785539
View details for PubMedCentralID PMC1987343
Onto-Tools: new additions and improvements in 2006
NUCLEIC ACIDS RESEARCH
2007; 35: W206-W211
Onto-Tools is a freely available web-accessible software suite, composed of an annotation database and nine complementary data-mining tools. This article describes a new tool, Onto-Express-to-go (OE2GO), as well as some new features implemented in Pathway-Express and Onto-Miner over the past year. Pathway-Express (PE) has been enhanced to identify significantly perturbed pathways in a given condition using the differentially expressed genes in the input. OE2GO is a tool for functional profiling using custom annotations. The development of this tool was aimed at the researchers working with organisms for which annotations are not yet available in the public domain. OE2GO allows researchers to use either annotation data from the Onto-Tools database, or their own custom annotations. By removing the necessity to use any specific database, OE2GO makes the functional profiling available for all organisms, with annotations using any ontology. The Onto-Tools are freely available at http://vortex.cs.wayne.edu/projects.htm.
View details for DOI 10.1093/nar/gkm327
View details for Web of Science ID 000255311500039
View details for PubMedID 17584796
View details for PubMedCentralID PMC1933142
Semantic analysis of genome annotations using weighting schemes
2007 IEEE SYMPOSIUM ON COMPUTATIONAL INTELLIGENCE IN BIOINFORMATICS AND COMPUTATIONAL BIOLOGY
View details for Web of Science ID 000248516200030
A system biology approach for the steady-state analysis of gene signaling networks
PROGRESS IN PATTERN RECOGNITION, IMAGE ANALYSIS AND APPLICATIONS, PROCEEDINGS
2007; 4756: 32-41
View details for Web of Science ID 000252725900004
Babel's tower revisited: a universal resource for cross-referencing across annotation databases
2006; 22 (23): 2934-2939
Annotation databases are widely used as public repositories of biological knowledge. However, most of these resources have been developed by independent groups which used different designs and different identifiers for the same biological entities. As we show in this article, incoherent name spaces between various databases represent a serious impediment to using the existing annotations at their full potential. Navigating between various such name spaces by mapping IDs from one database to another is a very important issue which is not properly addressed at the moment.We have developed a web-based resource, Onto-Translate (OT), which effectively addresses this problem. OT is able to map onto each other different types of biological entities from the following annotation databases: Swiss-Prot, TrEMBL, NREF, PIR, Gene Ontology, KEGG, Entrez Gene, GenBank, GenPept, IMAGE, RefSeq, UniGene, OMIM, PDB, Eukaryotic Promoter Database, HUGO Gene Nomenclature Committee and NetAffx. Currently, OT is able to perform 462 types of mappings between 29 different types of IDs from 17 databases concerning 53 organisms. Among these, over 300 types of translations and 15 types of IDs are not currently supported by any other tool or resource. On average, OT is able to correctly map between 96 and 99% of the biological entities provided as input. In terms of speed, sets of approximately 20 000 IDs can be translated in <30 s, in most cases.OT is a part of Onto-Tools, which is freely available at http://vortex.cs.wayne.edu/Projects.html
View details for DOI 10.1093/bioinofrmatics/btl372
View details for Web of Science ID 000242246300015
View details for PubMedID 17068090
View details for PubMedCentralID PMC2435247
New Onto-Tools: Promoter-Express, nsSNPCounter and Onto-Translate
NUCLEIC ACIDS RESEARCH
2006; 34: W626-W631
The Onto-Tools suite is composed of an annotation database and eight complementary, web-accessible data mining tools: Onto-Express, Onto-Compare, Onto-Design, Onto-Translate, Onto-Miner, Pathway-Express, Promoter-Express and nsSNPCounter. Promoter-Express is a new tool added to the Onto-Tools ensemble that facilitates the identification of transcription factor binding sites active in specific conditions. nsSNPCounter is another new tool that allows computation and analysis of synonymous and non-synonymous codon substitutions for studying evolutionary rates of protein coding genes. Onto-Translate has also been enhanced to expand its scope and accuracy by fully utilizing the capabilities of the Onto-Tools database. Currently, Onto-Translate allows arbitrary mappings between 28 types of IDs for 53 organisms. Onto-Tools are freely available at http://vortex.cs.wayne.edu/Projects.html.
View details for DOI 10.1093/nar/gkl213
View details for Web of Science ID 000245650200126
View details for PubMedID 16845086
View details for PubMedCentralID PMC1538776
Reliability and reproducibility issues in DNA microarray measurements
TRENDS IN GENETICS
2006; 22 (2): 101-109
DNA microarrays enable researchers to monitor the expression of thousands of genes simultaneously. However, the current technology has several limitations. Here we discuss problems related to the sensitivity, accuracy, specificity and reproducibility of microarray results. The existing data suggest that for relatively abundant transcripts the existence and direction (but not the magnitude) of expression changes can be reliably detected. However, accurate measurements of absolute expression levels and the reliable detection of low abundance genes are difficult to achieve. The main problems seem to be the sub-optimal design or choice of probes and some incorrect probe annotations. Well-designed data-analysis approaches can rectify some of these problems.
View details for DOI 10.1016/j.tig.2005.12.005
View details for Web of Science ID 000235576900009
View details for PubMedID 16380191
View details for PubMedCentralID PMC2386979
Ontological analysis of gene expression data: current tools, limitations, and open problems
2005; 21 (18): 3587-3595
Independent of the platform and the analysis methods used, the result of a microarray experiment is, in most cases, a list of differentially expressed genes. An automatic ontological analysis approach has been recently proposed to help with the biological interpretation of such results. Currently, this approach is the de facto standard for the secondary analysis of high throughput experiments and a large number of tools have been developed for this purpose. We present a detailed comparison of 14 such tools using the following criteria: scope of the analysis, visualization capabilities, statistical model(s) used, correction for multiple comparisons, reference microarrays available, installation issues and sources of annotation data. This detailed analysis of the capabilities of these tools will help researchers choose the most appropriate tool for a given type of analysis. More importantly, in spite of the fact that this type of analysis has been generally adopted, this approach has several important intrinsic drawbacks. These drawbacks are associated with all tools discussed and represent conceptual limitations of the current state-of-the-art in ontological analysis. We propose these as challenges for the next generation of secondary data analysis tools.
View details for DOI 10.1093/bioinformatics/bti565
View details for Web of Science ID 000231694600001
View details for PubMedID 15994189
View details for PubMedCentralID PMC2435250
A semantic analysis of the annotations of the human genome
2005; 21 (16): 3416-3421
The correct interpretation of any biological experiment depends in an essential way on the accuracy and consistency of the existing annotation databases. Such databases are ubiquitous and used by all life scientists in most experiments. However, it is well known that such databases are incomplete and many annotations may also be incorrect. In this paper we describe a technique that can be used to analyze the semantic content of such annotation databases. Our approach is able to extract implicit semantic relationships between genes and functions. This ability allows us to discover novel functions for known genes. This approach is able to identify missing and inaccurate annotations in existing annotation databases, and thus help improve their accuracy. We used our technique to analyze the current annotations of the human genome. From this body of annotations, we were able to predict 212 additional gene-function assignments. A subsequent literature search found that 138 of these gene-functions assignments are supported by existing peer-reviewed papers. An additional 23 assignments have been confirmed in the meantime by the addition of the respective annotations in later releases of the Gene Ontology database. Overall, the 161 confirmed assignments represent 75.95% of the proposed gene-function assignments. Only one of our predictions (0.4%) was contradicted by the existing literature. We could not find any relevant articles for 50 of our predictions (23.58%). The method is independent of the organism and can be used to analyze and improve the quality of the data of any public or private annotation database.
View details for DOI 10.1093/bioinformatics/bti538
View details for Web of Science ID 000231360600012
View details for PubMedID 15955782
View details for PubMedCentralID PMC2435251
Recent additions and improvements to the Onto-Tools
NUCLEIC ACIDS RESEARCH
2005; 33: W762-W765
The Onto-Tools suite is composed of an annotation database and six seamlessly integrated, web-accessible data mining tools: Onto-Express, Onto-Compare, Onto-Design, Onto-Translate, Onto-Miner and Pathway-Express. The Onto-Tools database has been expanded to include various types of data from 12 new databases. Our database now integrates different types of genomic data from 19 sequence, gene, protein and annotation databases. Additionally, our database is also expanded to include complete Gene Ontology (GO) annotations. Using the enhanced database and GO annotations, Onto-Express now allows functional profiling for 24 organisms and supports 17 different types of input IDs. Onto-Translate is also enhanced to fully utilize the capabilities of the new Onto-Tools database with an ultimate goal of providing the users with a non-redundant and complete mapping from any type of identification system to any other type. Currently, Onto-Translate allows arbitrary mappings between 29 types of IDs. Pathway-Express is a new tool that helps the users find the most interesting pathways for their input list of genes. Onto-Tools are freely available at http://vortex.cs.wayne.edu/Projects.html.
View details for DOI 10.1093/nar/gki472
View details for Web of Science ID 000230271400156
View details for PubMedID 15980579
View details for PubMedCentralID PMC1160233
Identification of genomic signatures for the design of assays for the detection and monitoring of anthrax threats
PACIFIC SYMPOSIUM ON BIOCOMPUTING 2005
Sequences that are present in a given species or strain while absent from or different in any other organisms can be used to distinguish the target organism from other related or un-related species. Such DNA signatures are particularly important for the identification of genetic source of drug resistance of a strain or for the detection of organisms that can be used as biological agents in warfare or terrorism. Most approaches used to find DNA signatures are laboratory based, require a great deal of effort and can only distinguish between two organisms at a time. We propose a more efficient and cost-effective bioinformatics approach that allows identification of genomic fingerprints for a target organism. We validated our approach using a custom microarray, using sequences identified as DNA fingerprints of Bacillus anthracis. Hybridization results showed that the sequences found using our algorithm were truly unique to B. anthracis and were able to distinguish B. anthracis from its close relatives B. cereus and B. thuringiensis.
View details for Web of Science ID 000230169100021
View details for PubMedID 15759631
A novel bioinformatics technique for predicting condition-specific transcription factor binding sites
PROCEEDINGS OF THE 2005 IEEE SYMPOSIUM ON COMPUTATIONAL INTELLIGENCE IN BIOINFORMATICS AND COMPUTATIONAL BIOLOGY
View details for Web of Science ID 000235518600028
Onto-Tools: an ensemble of web-accessible, ontology-based tools for the functional design and interpretation of high-throughput gene expression experiments
NUCLEIC ACIDS RESEARCH
2004; 32: W449-W456
The Onto-Tools suite is composed of an annotation database and five seamlessly integrated web-accessible data mining tools: Onto-Express (OE), Onto-Compare (OC), Onto-Design (OD), Onto-Translate (OT) and Onto-Miner (OM). OM is a new tool that provides a unified access point and an application programming interface for most annotations available. Our database has been enhanced with more than 120 new commercial microarrays and annotations for Rattus norvegicus, Drosophila melanogaster and Carnorhabditis elegans. The Onto-Tools have been redesigned to provide better biological insight, improved performance and user convenience. The new features implemented in OE include support for gene names, LocusLink IDs and Gene Ontology (GO) IDs, ability to specify fold changes for the input genes, links to the KEGG pathway database and detailed output files. OC allows comparisons of the functional bias of more than 170 commercial microarrays. The latest version of OD allows the user to specify keywords if the exact GO term is not known as well as providing more details than the previous version. OE, OC and OD now have an integrated GO browser that allows the user to customize the level of abstraction for each GO category. The Onto-Tools are available online at http://vortex.cs.wayne.edu/Projects.html.
View details for DOI 10.1093/nar/gkh409
View details for Web of Science ID 000222273100090
View details for PubMedID 15215428
View details for PubMedCentralID PMC441547
Onto-Tools, the toolkit of the modern biologist: Onto-Express, Onto-Compare, Onto-Design and Onto-Translate
NUCLEIC ACIDS RESEARCH
2003; 31 (13): 3775-3781
Onto-Tools is a set of four seamlessly integrated databases: Onto-Express, Onto-Compare, Onto-Design and Onto-Translate. Onto-Express is able to automatically translate lists of genes found to be differentially regulated in a given condition into functional profiles characterizing the impact of the condition studied upon various biological processes and pathways. OE constructs functional profiles (using Gene Ontology terms) for the following categories: biochemical function, biological process, cellular role, cellular component, molecular function and chromosome location. Statistical significance values are calculated for each category. Once the initial exploratory analysis identified a number of relevant biological processes, specific mechanisms of interactions can be hypothesized for the conditions studied. Currently, many commercial arrays are available for the investigation of specific mechanisms. Each such array is characterized by a biological bias determined by the extent to which the genes present on the array represent specific pathways. Onto-Compare is a tool that allows efficient comparisons of any sets of commercial or custom arrays. Using Onto-Compare, a researcher can determine quickly which array, or set of arrays, covers best the hypotheses studied. In many situations, no commercial arrays are available for specific biological mechanisms. Onto-Design is a tool that allows the user to select genes that represent given functional categories. Onto-Translate allows the user to translate easily lists of accession numbers, UniGene clusters and Affymetrix probes into one another. All tools above are seamlessly integrated. The Onto-Tools are available online at http://vortex.cs.wayne.edu/Projects.html.
View details for DOI 10.1093/nar/gkg624
View details for Web of Science ID 000183832900108
View details for PubMedID 12824416
View details for PubMedCentralID PMC169030
Assessing the functional bias of commercial microarrays using the onto-compare database
Microarrays are at the center of a revolution in biotechnology, allowing researchers to screen tens of thousands of genes simultaneously. Typically, they have been used in exploratory research to help formulate hypotheses. In most cases, this phase is followed by a more focused, hypothesis-driven stage in which certain specific biological processes and pathways are thought to be involved. Since a single biological process can still involve hundreds of genes, microarrays are still the preferred approach as proven by the availability of focused arrays from several manufacturers. Because focused arrays from different manufacturers use different sets of genes, each array will represent any given regulatory pathway to a different extent. We argue that a functional analysis of the arrays available should be the most important criterion used in the array selection. We developed Onto-Compare as a database that can provide this functionality, based on the Gene Ontology Consortium nomenclature. We used this tool to compare several arrays focused on apoptosis, oncogenes, and tumor suppressors. We considered arrays from BD Biosciences Clontech, PerkinElmer, Sigma-Genosys, and SuperArray. We showed that among the oncogene arrays, the PerkinElmer MICROMAX oncogene microarray has a better representation of oncogenesis, protein phosphorylation, and negative control of cell proliferation. The comparison of the apoptosis arrays showed that most apoptosis-related biological processes are equally well represented on the arrays considered. However, functional categories such as immune response, cell-cell signaling, cell-surface receptor linked signal transduction, and interleukins are better represented on the Sigma-Genoys Panorama human apoptosis array. At the same time, processes such as cell cycle control, oncogenesis, and negative control of cell proliferation are better represented on the BD Biosciences Clontech Atlas Select human apoptosis array.
View details for Web of Science ID 000181595900009
View details for PubMedID 12664686
Global functional profiling of gene expression
2003; 81 (2): 98-104
The typical result of a microarray experiment is a list of tens or hundreds of genes found to be differentially regulated in the condition under study. Independent of the methods used to select these genes, the common task faced by any researcher is to translate these lists of genes into a better understanding of the biological phenomena involved. Currently, this is done through a tedious combination of searches through the literature and a number of public databases. We developed Onto-Express (OE) as a novel tool able to automatically translate such lists of differentially regulated genes into functional profiles characterizing the impact of the condition studied. OE constructs functional profiles (using Gene Ontology terms) for the following categories: biochemical function, biological process, cellular role, cellular component, molecular function, and chromosome location. Statistical significance values are calculated for each category. We demonstrate the validity and the utility of this comprehensive global analysis of gene function by analyzing two breast cancer datasets from two separate laboratories. OE was able to identify correctly all biological processes postulated by the original authors, as well as discover novel relevant mechanisms.
View details for DOI 10.1016/S0888-7543(02)00021-6
View details for Web of Science ID 000181532700002
View details for PubMedID 12620386
Spermatozoal RNA profiles of normal fertile men
2002; 360 (9335): 772-777
Findings from several studies support the conclusion that spermatozoa contain a complex repertoire of mRNAs. Even though these mRNAs are thought to provide an insight into past events of spermatogenesis, their complexity and function have yet to be established. Our aim was to determine whether we could use spermatozoal mRNAs to generate a genetic fingerprint of normal fertile men.We used a suite of microarrays containing 27016 unique expressed sequence tags (ESTs) to investigate cDNAs from a pool of 19 testes, cDNAs from a pool of nine individual ejaculate spermatozoal mRNAs, and cDNAs constructed from spermatozoal mRNAs from a single ejaculate. We also used ontological data mining to determine the function of the genes identified in each EST profile.The cDNAs from the testes, pooled ejaculate, and single ejaculate hybridised to 7157, 3281, and 2780 ESTs, respectively. The testicular population contained all of the ESTs identified by the cDNAs from the pooled and individual ejaculate. The pooled ejaculate population contained all but four ESTs identified from the individual ejaculate. A subset of the spermatozoal mRNAs was associated with embryo development.The microarray data from testes and spermatozoa (pooled and individual) were concordant, supporting the view that a spermatozoal mRNA fingerprint can be obtained from normal fertile men. Thus, profiling can be used to monitor past events-ie, gene expression of spermatogenesis. Moreover, the data suggest that, in addition to delivering the paternal genome, spermatozoa provide the zygote with a unique suite of paternal mRNAs. Ejaculate spermatozoa can now be used as a non-invasive proxy for investigations of testis-specific infertility.
View details for Web of Science ID 000177933000019
View details for PubMedID 12241836
Profiling gene expression using onto-express
2002; 79 (2): 266-270
Gene expression profiles obtained through microarray or data mining analyses often exist as vast data strings. To interpret the biology of these genetic profiles, investigators must analyze this data in the context of other information such as the biological, biochemical, or molecular function of the translated proteins. This is particularly challenging for a human analyst because large quantities of less than relevant data often bury such information. To address this need we implemented an automated routine, called Onto-Express (http://vortex.cs.wayne.edu:8080), to systematically translate genetic fingerprints into functional profiles. Using strings of accession or cluster identification numbers, Onto-Express searches the public databases and returns tables that correlate expression profiles with the cytogenetic locations, biochemical and molecular functions, biological processes, cellular components, and cellular roles of the translated proteins. The profiles created by Onto-Express fundamentally increase the value of gene expression analyses by facilitating the translation of quantitative value sets to records that contain biological implications.
View details for DOI 10.1006/geno.2002.6698
View details for Web of Science ID 000173628100016
View details for PubMedID 11829497