Early Gene Signature Research Articles

Identification of early gene expression profiles associated with long-lasting antibody responses to the Ebola vaccine Ad26.ZEBOV/MVA-BN-Filo

Ebola virus disease is a severe hemorrhagic fever with a high fatality rate. We investigate transcriptome profiles at 3 h, 1day, and 7days after vaccination with Ad26.ZEBOV and MVA-BN-Filo. 3h after Ad26.ZEBOV injection, we observe an increase in genes related to antigen presentation, sensing, and T and B cell receptors. The highest response occurs 1day after Ad26.ZEBOV injection, with an increase of the gene expression of interferon-induced antiviral molecules, monocyte activation, and sensing receptors. This response is regulated by the HESX1, ATF3, ANKRD22, and ETV7 transcription factors. A plasma cell signature is observed on day 7 post-Ad26.ZEBOV vaccination, with an increase of CD138, MZB1, CD38, CD79A, and immunoglobulin genes. We have identified early expressed genes correlated with the magnitude of the antibody response 21days after the MVA-BN-Filo and 364days after Ad26.ZEBOV vaccinations. Our results provide early gene signatures that correlate with vaccine-induced Ebola virus glycoprotein-specific antibodies.

Transcriptome-based prediction of drugs, inhibiting cardiomyogenesis in human induced pluripotent stem cells

SummaryAnimal studies for embryotoxicity evaluation of potential therapeutics and environmental factors are complex, costly, and time-consuming. Often, studies are not of human relevance because of species differences. In the present study, we recapitulated the process of cardiomyogenesis in human induced pluripotent stem cells (hiPSCs) by modulation of the Wnt signaling pathway to identify a key cardiomyogenesis gene signature that can be applied to identify compounds and/or stress factors compromising the cardiomyogenesis process. Among the 23 tested teratogens and 16 non-teratogens, we identified three retinoids including 13-cis-retinoic acid that completely block the process of cardiomyogenesis in hiPSCs. Moreover, we have identified an early gene signature consisting of 31 genes and associated biological processes that are severely affected by the retinoids. To predict the inhibitory potential of teratogens and non-teratogens in the process of cardiomyogenesis we established the “Developmental Cardiotoxicity Index” (CDI31g) that accurately differentiates teratogens and non-teratogens to do or do not affect the differentiation of hiPSCs to functional cardiomyocytes.

Onset of hippocampal network aberration and memory deficits in P301S tau mice are associated with an early gene signature.

Hyperphosphorylation and deposition of tau in the brain characterizes frontotemporal dementia and Alzheimer's disease. Disease-associated mutations in the tau-encoding MAPT gene have enabled the generation of transgenic mouse models that recapitulate aspects of human neurodegenerative diseases, including tau hyperphosphorylation and neurofibrillary tangle formation. Here, we characterized the effects of transgenic P301S mutant human tau expression on neuronal network function in the murine hippocampus. Onset of progressive spatial learning deficits in P301S tau transgenic TAU58/2 mice were paralleled by long-term potentiation deficits and neuronal network aberrations during electrophysiological and EEG recordings. Gene-expression profiling just prior to onset of apparent deficits in TAU58/2 mice revealed a signature of immediate early genes that is consistent with neuronal network hypersynchronicity. We found that the increased immediate early gene activity was confined to neurons harbouring tau pathology, providing a cellular link between aberrant tau and network dysfunction. Taken together, our data suggest that tau pathology drives neuronal network dysfunction through hyperexcitation of individual, pathology-harbouring neurons, thereby contributing to memory deficits.

Predictive early gene signature during mouse Bhas 42 cell transformation induced by synthetic amorphous silica nanoparticles

Synthetic amorphous silica nanoparticles (SAS) are used widely in industrial applications. These nanoparticles are not classified for their carcinogenicity in humans. However, some data still demonstrate a potential carcinogenic risk of these compounds in humans. The Bhas 42 cell line was developed to screen chemicals, as tumor-initiators or -promoters according to their ability to trigger cell-to-cell transformation, in a cell transformation assay. In the present study, we performed unsupervised transcriptomic analysis after exposure of Bhas 42 cells to NM-203 SAS as well as to positive (Min-U-Sil 5® crystalline silica microparticles, and 12-O-tetradecanoylphorbol-13-acetate) and negative (diatomaceous earth) control compounds. We identified a common gene signature for 21 genes involved in the early stage of the SAS- Min-U-Sil 5®- or TPA-induced cell transformation. These genes were related to cell proliferation (over expression) and cell adhesion (under expression). Among them, 12 were selected on the basis of their potential impact on cell transformation. RT-qPCR and western blotting were used to confirm the transcriptomic data. Moreover, similar gene alterations were found when Bhas 42 cells were treated with two other transforming SAS. In conclusion, the results obtained in the current study highlight a 12-gene signature that could be considered as a potential early “bio-marker” of cell transformation induced by SAS and perhaps other chemicals.

Essential Role of CARD14 in Murine Experimental Psoriasis.

Caspase recruitment domain family member 14 (CARD14) was recently identified as a psoriasis-susceptibility gene, but its immunological role in the pathogenesis of psoriasis in vivo remains unclear. In this study, we examined the role of CARD14 in murine experimental models of psoriasis induced by either imiquimod (IMQ) cream or recombinant IL-23 injection. In all models tested, the psoriasiform skin inflammation was abrogated in Card14-/- mice. Comparison of the early gene signature of the skin between IMQ-cream-treated Card14-/- mice and Tlr7-/-Tlr9-/- mice revealed not only their similarity, but also distinct gene sets targeted by IL-23. Cell type-specific analysis of these mice identified skin Langerinhigh Langerhans cells as a potent producer of IL-23, which was dependent on both TLR7 and TLR9 but independent of CARD14, suggesting that CARD14 is acting downstream of IL-23, not TLR7 or TLR9. Instead, a bone marrow chimera study suggested that CARD14 in radio-sensitive hematopoietic cells was required for IMQ-induced psoriasiform skin inflammation, controlling the number of Vγ4+ T cells producing IL-17 or IL-22 infiltrating through the dermis to the inflamed epidermis. These data indicate that CARD14 is essential and a potential therapeutic target for psoriasis.

Transcriptomics in Human Challenge Models

Human challenge models, in which volunteers are experimentally infected with a pathogen of interest, provide the opportunity to directly identify both natural and vaccine-induced correlates of protection. In this review, we highlight how the application of transcriptomics to human challenge studies allows for the identification of novel correlates and gives insight into the immunological pathways required to develop functional immunity. In malaria challenge trials for example, innate immune pathways appear to play a previously underappreciated role in conferring protective immunity. Transcriptomic analyses of samples obtained in human challenge studies can also deepen our understanding of the immune responses preceding symptom onset, allowing characterization of innate immunity and early gene signatures, which may influence disease outcome. Influenza challenge studies demonstrate that these gene signatures have diagnostic potential in the context of pandemics, in which presymptomatic diagnosis of at-risk individuals could allow early initiation of antiviral treatment and help limit transmission. Furthermore, gene expression analysis facilitates the identification of host factors contributing to disease susceptibility, such as C4BPA expression in enterotoxigenic Escherichia coli infection. Overall, these studies highlight the exceptional value of transcriptional data generated in human challenge trials and illustrate the broad impact molecular data analysis may have on global health through rational vaccine design and biomarker discovery.

AS03- and MF59-Adjuvanted Influenza Vaccines in Children.

Influenza is a major cause of respiratory disease leading to hospitalization in young children. However, seasonal trivalent influenza vaccines (TIVs) have been shown to be ineffective and poorly immunogenic in this population. The development of live-attenuated influenza vaccines and adjuvanted vaccines are important advances in the prevention of influenza in young children. The oil-in-water emulsions MF59 and adjuvant systems 03 (AS03) have been used as adjuvants in both seasonal adjuvanted trivalent influenza vaccines (ATIVs) and pandemic monovalent influenza vaccines. Compared with non-adjuvanted vaccine responses, these vaccines induce a more robust and persistent antibody response for both homologous and heterologous influenza strains in infants and young children. Evidence of a significant improvement in vaccine efficacy with these adjuvanted vaccines resulted in the use of the monovalent (A/H1N1) AS03-adjuvanted vaccine in children in the 2009 influenza pandemic and the licensure of the seasonal MF59 ATIV for children aged 6 months to 2 years in Canada. The mechanism of action of MF59 and AS03 remains unclear. Adjuvants such as MF59 induce proinflammatory cytokines and chemokines, including CXCL10, but independently of type-1 interferon. This proinflammatory response is associated with improved recruitment, activation and maturation of antigen presenting cells at the injection site. In young children MF59 ATIV produced more homogenous and robust transcriptional responses, more similar to adult-like patterns, than did TIV. Early gene signatures characteristic of the innate immune response, which correlated with antibody titers were also identified. Differences were detected when comparing child and adult responses including opposite trends in gene set enrichment at day 3 postvaccination and, unlike adult data, a lack of correlation between magnitude of plasmablast response at day 7 and antibody titers at day 28 in children. These insights show the utility of novel approaches in understanding new adjuvants and their importance for developing improved influenza vaccines for children.

PO and ID BCG vaccination in humans induce distinct mucosal and systemic immune responses and CD4+ T cell transcriptomal molecular signatures

Protective efficacy of Bacillus Calmette-Guérin (BCG) may be affected by the methods and routes of vaccine administration. We have studied the safety and immunogenicity of oral (PO) and/or intradermal (ID) administration of BCG in healthy human subjects. No major safety concerns were detected in the 68 healthy adults vaccinated with PO and/or ID BCG. Although both PO and ID BCG could induce systemic Th1 responses capable of IFN-γ production, ID BCG more strongly induced systemic Th1 responses. In contrast, stronger mucosal responses (TB-specific secretory IgA and bronchoalveolar lavage T cells) were induced by PO BCG vaccination. To generate preliminary data comparing the early gene signatures induced by mucosal and systemic BCG vaccination, CD4+ memory T cells were isolated from subsets of BCG vaccinated subjects pre- (Day 0) and post-vaccination (Days 7 and 56), rested or stimulated with BCG infected dendritic cells, and then studied by Illumina BeadArray transcriptomal analysis. Notably, distinct gene expression profiles were identified both on Day 7 and Day 56 comparing the PO and ID BCG vaccinated groups by GSEA analysis. Future correlation analyses between specific gene expression patterns and distinct mucosal and systemic immune responses induced will be highly informative for TB vaccine development.

Molecular signatures of the evolving immune response in mice following a Bordetella pertussis infection.

Worldwide resurgence of pertussis necessitates the need for improvement of pertussis vaccines and vaccination strategies. Since natural infections induce a longer-lasting immunity than vaccinations, detailed knowledge of the immune responses following natural infection can provide important clues for such improvement. The purpose was to elucidate the kinetics of the protective immune response evolving after experimental Bordetella pertussis (B. pertussis) infection in mice. Data were collected from (i) individual analyses, i.e. microarray, flow cytometry, multiplex immunoassays, and bacterial clearance; (ii) twelve time points during the infection; and (iii) different tissues involved in the immune responses, i.e. lungs, spleen and blood. Combined data revealed detailed insight in molecular and cellular sequence of events connecting different phases (innate, bridging and adaptive) of the immune response following the infection. We detected a prolonged acute phase response, broad pathogen recognition, and early gene signatures of subsequent T-cell recruitment in the lungs. Activation of particular transcription factors and specific cell markers provided insight into the time course of the transition from innate towards adaptive immune responses, which resulted in a broad spectrum of systemic antibody subclasses and splenic Th1/Th17 memory cells against B. pertussis. In addition, signatures preceding the local generation of Th1 and Th17 cells as well as IgA in the lungs, considered key elements in protection against B. pertussis, were established. In conclusion, molecular and cellular immunological processes in response to live B. pertussis infection were unraveled, which may provide guidance in selecting new vaccine candidates that should evoke local and prolonged protective immune responses.

A systems biology approach demonstrates that PO and ID BCG induce distinct CD4+ T cell molecular signatures. (VAC7P.970)

Abstract Vaccination with Bacillus Calmette-Guérin (BCG) is and constitutes the most widely used tuberculosis (TB) control strategy in the world. Protective efficacy of BCG may be affected by the methods and routes of vaccine administration. Here we have used a systems biology approach to study innate and adaptive responses to oral (PO) and intradermal (ID) BCG vaccination in humans. 60 healthy adults vaccinated with PO and/or ID BCG was studied. Although both PO and ID BCG could induce systemic Th1 responses capable of IFN-γ production, ID BCG more strongly induced systemic Th1 responses, while significantly higher mucosal responses (TB-specific sIgA and bronchoalveolar lavage T cells) were induced by PO BCG vaccination. In order to identify early gene signatures that predict optimal mucosal and systemic immune responses, CD4+ memory T cells were isolated from both PO and ID BCG vaccinated volunteers pre- (Day 0) and post-vaccination (Day 7 and 56), and studied by Illumina BeadArray transcriptomal analysis. Notably, distinct gene expression profiles were identified both on day 7 and day 56 comparing the PO and ID BCG vaccinated groups by GSEA analysis. Correlations between specific expression patterns and mucosal vs. systemic immune responses are being studied. These data highlight the utility of systems biology approaches in studying differential mucosal and systemic immune responses.

Impact of supervised gene signatures of early hypoxia on patient survival

Background and purpose Hypoxia is a common feature of solid tumors associated with therapy resistance, increased malignancy and poor prognosis. Several approaches have been developed with the hope of identifying patients harboring hypoxic tumors including the use of microarray based gene signatures. However, studies to date have largely ignored the strong time dependency of hypoxia-regulated gene expression. We hypothesized that use of time-dependent patterns of gene expression during hypoxia would enable development of superior prognostic expression signatures. Materials and methods Using published data from the microarray study of Chi et al., we extracted gene signatures correlating with induction during either early or late hypoxic exposure. Gene signatures were derived from in vitro exposed human mammary epithelial cell line (HMEC) under 0% or 2% oxygen. Gene signatures correlating with early and late up-regulation were tested by means of Kaplan–Meier survival, univariate, and multivariate analysis on a patient data set with primary breast cancer treated conventionally (surgery plus on indication radiotherapy and systemic therapy). Results We found that the two early hypoxia gene signatures extracted from 0% and 2% hypoxia showed significant prognostic power (log-rank test: p = 0.004 at 0%, p = 0.034 at 2%) in contrast to the late hypoxia signatures. Both early gene signatures were linked to the insulin pathway. From the multivariate Cox-regression analysis, the early hypoxia signature ( p = 0.254) was found to be the 4th best prognostic factor after lymph node status ( p = 0.002), tumor size ( p = 0.016) and Elston grade ( p = 0.111). On this data set it indeed provided more information than ER status or p53 status. Conclusions The hypoxic stress elicits a wide panel of temporal responses corresponding to different biological pathways. Early hypoxia signatures were shown to have a significant prognostic power. These data suggest that gene signatures identified from in vitro experiments could contribute to individualized medicine.