Researchers in metabolomics have long recognized that an individual’s metabolic profile is very sensitive to multiple factors such as age, gender, diet, etc. It is natural, sometimes necessary, to include these additional factors during metabolomic data analysis. MetATT (http://metatt.metabolomics.ca) is a user-friendly, web-based tool designed for analyzing two-factor (including time-series) metabolomic data, e.g., to consider the gender effect in a case-control study, or to investigate temporal profiles under different experimental conditions.

To use MetATT, users first need to convert their data into the proper format. MetATT accepts a data table with samples in rows and features (spectral bins, peaks, compounds) in column (or in transposed format). The first column must contain sample names, while the next two columns must contain group labels of the two factors. For time-series data, the column of different time points must be labeled as Time, and samples from the same subjects must appear as blocks. More detailed instructions can be obtained on the website (Fig.1).

 

Figure 1. MetATT home page. This page gives an overview of the methods implemented in MetATT. Experienced users can directly click the "Click here to start" link to start analysis. First-time users should follow the "Data Formats" link or the "FAQs" link to become familiar with the basic requirements and features of the program. The "Resources" link provides the R scripts for users who want to run the analysis on their local computers from the command line of R.


MetATT contains five carefully selected methods to help researchers understand their data. In particular, MetATT provides two visualization techniques to facilitate data exploration and pattern discovery – interactive 3D principal component analysis (PCA) and two-way clustered heat map. The PCA visualization supports rotation, zooming, and highlighting. Pointing and clicking on any data point will display the characteristic variables of the corresponding sample (Fig. 2). The two-way clustered heat map presents all the data points in the form of colored heat maps based on their (usually normalized) values. Users can highlight and re-arrange the heat map data any way they wish (Fig. 3). MetATT also supports the classical univariate two-way ANOVA method, along with two recent multivariate extensions – ANOVA-simultaneous component analysis (ASCA) and multivariate empirical Bayes time-series data analysis (MEBA). The two-way ANOVA supports within- and between- subjects analysis for time-series/repeated samples and two-factor independent samples, respectively. The results are summarized as a Venn diagram (Fig. 4) in addition to a detailed data table. The ASCA method first decomposes the variation in the entire data set into individual variations induced by major experimental factors (and their interactions), and then applies PCA to each variation to obtain a simple and interpretable result. MetATT’s implementation of ASCA also incorporates two recent improvements for variable selection and model validation. The MEBA method compares the average temporal changes of individual variables among different biological conditions. The result is a ranked list of variables with distinctive temporal profiles (Fig. 5).



Figure 2. Interactive 3D PCA visualization. The colors of these data points are based on the factor labels. Users can change the colors according to any of the two factor labels. Users can also rotate or zoom the 3D plot. Clicking on any data point will display the top 20 characteristic variables of the corresponding sample on the right hand side of the screen.




Figure 3. Two-way clustered heat map. The samples are sorted by their factor labels. Users can change the hierarchical clustering algorithms, the color schema, as well as the order of the samples to suit their preferences.




Figure 4. Venn diagram summary of two-way ANOVA results.




Figure 5. The top four variables with distinctive temporal profiles. The temporal profiles are presented as line graphs coloured based on the two conditions (WT and MT) under study.


The complex procedures are implemented in a simple, stepwise manner through a user-friendly web-interface. At the end of session, a detailed analysis report will be generated which integrates the background introduction of each method used together with the corresponding graphical and text outputs.

 

Metabolomics is an emerging field that is complementary to other omics sciences and that is gaining increasing interest across all disciplines. Because of metabolomics' unique advantages, it is now being applied in functional genomics, integrative and systems biology, pharmacogenomics, and biomarker discovery for drug development and therapy monitoring.  More than 95% of today's biomarkers are small molecules or metabolites (MW <1500 Da), which can be used for disease testing, drug testing, toxic exposure testing, and food consumption tracking. While standard clinical assays are limited in the number and type of compounds that can be detected, metabolomics measures many more compounds. Since a single compound is not always the best biomarker (diagnostic, prognostic or predictive), the use of metabolomics allows information about multiple compounds to be used together to make better medical decisions. Global metabolic profiling has now been used to determine clinical biomarkers in assessing the pathophysiological health status of patients.

In the following two recent clinical studies, metabolomics approaches were used to predict the risk for cardiovascular disease (CVD) and the development of diabetes, respectively.

1. Z. Wang, E. Klipfell, B.J. Bennett, R. Koeth, B.S. Levison, B. DuGar et al., Gut Flora Metabolism of Phosphatidylcholine Promotes Cardiovascular Disease, Nature, 2011, vol 472, pp. 57-63.

Metabolic profiles of plasma in an independent large clinical cohort were used to predict risk for cardiovascular disease (CVD). Three plasma metabolites of the dietary lipid phosphatidylcholine (PC) - choline, trimethylamine N-oxide (TMAO) and betaine - were identified as predictive biomarkers for CVD. Stable isotope dilution LC/MS/MS was used for quantification of those metabolites. This is the first study to identify a direct link between intestinal microflora, dietary PC and CVD risk. This knowledge could be used for the development of new diagnostic tests and may help to inform therapeutic strategies for CVD.


2. T.J. Wang, M.G. Larson, R.S. Vasan, S. Cheng, E.P. Rhee, E. McCabe et al., Metabolite Profiles and The Risk of Developing Diabetes, Nature Medicine, 2011, vol 17, pp. 448-454

Plasma metabolic profiles of 2,422 subjects from the Framingham Offspring Study were performed using an MS-based metabolomics platform (LC-MS/MS). The objective was to investigate whether metabolite profiles could predict the development of diabetes. Amino acids, amines and other polar metabolites were profiled. From these groups, five metabolites of interest were identified: isoleucine, leucine, valine, tyrosine and phenylalanine. These branched-chain and aromatic amino acids were identified as predictors of the future development of diabetes. This study suggests that amino acid profiles could be used in diabetes risk assessment.

3) Metabolomics Current Contents

Recently published papers in metabolomics:

• Metabolomics in early Alzheimer's disease: identification of altered plasma sphingolipidome using shotgun lipidomics.
  
• Serum metabolomics as a novel diagnostic approach for gastrointestinal cancer.
• An overview of NMR-based metabolomics to identify secondary plant compounds involved in host plant resistance.
• Metabolomics reveals phospholipids as important nutrient sources during Salmonella growth in bile in vitro and in vivo.
• Metabolomics study of alcohol-induced liver injury and hepatocellular carcinoma xenografts in mice.
• Metabolomics of faecal extracts detects altered metabolic activity of gut microbiota in ulcerative colitis and irritable bowel syndrome.
• Metabolomics reveals differential levels of oral metabolites in HIV-infected patients: Toward novel diagnostic targets.
• UPLC-MS-based urine metabolomics reveals indole-3-lactic acid and phenyllactic acid as conserved biomarkers for alcohol-induced liver disease in the Ppara-null mouse model.
• Identifying static and kinetic lipid phenotypes by high resolution UPLC/MS: Unraveling diet-induced changes in lipid homeostasis by coupling metabolomics and fluxomics.
  

4) MetaboNews

Jul 2011	KEGG has gone commercial - Due to a lack of funding, one of Biology's most popular databases has moved to a subscription model. Minoru Kanehisa, a bioinformatics professor at Kyoto University, has led this project since its beginning in 1995. The KEGG website will continue to be freely available. However, academic and commercial users wanting access to KEGG's FTP site will now have to pay for access. Kanehisa and his colleagues have started a non-profit organization, NPO Bioinformatics Japan, as a means for raising funds to pay for KEGG's operations. Academic users will pay $2,000 per year for individual users and $5,000 per year for organizations. Kanehisa estimates that if they can get just 10% of the current FTP users (totalling 3,000 users per month), then they will have enough money to survive. A lot of work goes into making KEGG the high quality database that academic and commercial users have come to rely on during the last 15 plus years. As a result, KEGG employs a large team of 25 full-time staff and 5-10 part-time employees. To support KEGG and all of its efforts to date, consider a KEGG FTP subscription. Sources: KEGG and GenomeWeb
Jul 2011	Metabolomics Society meeting summary - The Seventh International Conference of the Metabolomics Society was held in Cairns, Australia, June 27-30, 2011, following relocation from Japan due to possible power shortages following tsunami.  Despite the logistic challenges of such a last-minute move, the Australian hosts put together a great event with more than 140 posters and 80 speakers from around the world. The program was packed with presentations covering a wide variety of topics in metabolomics: everything from biomarker discovery, pathways and disease, plants, microbes and the environment, to pharmacology and nutrition. New technologies, fluxomics, bioinformatics, and systems biology were also hot topics. The emphasis on new capabilities in quantitative lipidomics was especially noteworthy.  Another highlight was the presentation from Dr. Marasu Tomita, who is the director of the Institute for Advanced Biosciences, Keio University and was one of the original organizers for the conference. His talk emphasized the growing prominence of metabolomics in Japan, as reflected by the impressive metabolomics facilities in Tsuruoka city and their remarkable analytical capabilities. The south pacific, including Japan, Australia and New Zealand was generally very well represented at the conference, showing the growing global importance of metabolomics as an omics science. Japan will host the conference in 2014. Conference proceedings are available on the Metabolomics Society website at http://www.metabolomicssociety.org/news.html.7

Please note: If you know of any metabolomics news that we should feature in future issues of this newsletter, please email Ian Forsythe (metabolomics.innovation@gmail.com).

5) Metabolomics Events

12-14 Sep 2011	CIC bioGUNE International Workshop on Metabolomics Venue: Bilbao, Spain In the "Global Systems Biology" era, metabolomics constitutes one of the most powerful technologies to integrate the biological pathways and understand how a living organism interacts with its environment. Metabolomics can be defined as the quantitative and qualitative analysis of all metabolites (molecules with a molecular weight of less than 1,500 Da) in a given organism. This results in the construction of a metabolome or metabolic fingerprint, analogous to the genome or the proteome. Since the set of all metabolites is directly linked to the actual state of a cell, tissue or an organism and thus to the phenotype, the metabolome is optimally suited for the determination of biomarkers that are typical for certain genotypes or pathologies, and to identify key cellular pathways involved in the development and progression of diseases. CIC bioGUNE has prepared a very interesting scientific program in which leading academics speakers will tell us about the state-of-the-art in metabolomics including technological aspects related to the different platforms (RMN, GC and LC). The workshop will also cover recent breakthroughs in single-cell and high-throughput metabolomics approaches along with the applications that this technology has found in environmental and disease biomarker discovery areas. Importantly, the scientific program will be completed with poster sessions and presentations from equipment manufacturers. In summary the participants will have the opportunity to learn, meet and interact with experts in different metabolomics-related areas obtaining a deep and broad vision on this powerful technology. For more information, please visit http://metabolomicsworkshop2011.cicbiogune.es/.
25-28 Sep 2011	Metabomeeting 2011 Venue: Helsinki, Finland The Metabolic Profiling Forum (MPF) is pleased to announce Metabomeeting 2011, to be held in collaboration with the VTT Research Centre of Finland at the Helsinki Congress Paasitorni. The conference centre is located in Siltasaari, in the heart of Helsinki, on the shore-front and next to the Hakaniemi market   The MPF is also pleased to be organising Metabomeeting 2011 in conjunction with the inaugural Nordic Metabolomics meeting, which for this first meeting will be focusing on the latest developments/enhancements in data analysis and modelling. For more information, please visit http://thempf.org/mpf_cms/index.php/conferencesworkshops/forthcoming-mpf-events/33-mm11.
30 Sep-4 Oct 2011	ASMS Asilomar Metabolomics meeting Venue: Pacific Grove, California, USA The upcoming ASMS Asilomar Metabolomics meeting will promote an in-depth discussion of current problems and perspectives of mass spectrometry-based metabolomics. The meeting is endorsed by the Metabolomics Society. We will challenge ideas and concepts throughout the conference and have allocated ample time for discussions, e.g., during evening receptions. In addition to invited speakers, there will be poster sessions and oral contributions based on abstract submissions. We will focus on a range of hot  topics, including:  databases; metabolite/gene annotations; fluxes; compound identifications; metabolite imaging; secondary metabolism; metabolomics in human diseases; and quality control for high-throughput metabolomic quantifications. Download conference brochure For more information, please visit http://www.asms.org/Conferences/AsilomarConference/GeneralInformation/tabid/99/Default.aspx.
8-9 Nov 2011	7th annual Advances in Metabolic Profiling Venue: Dublin, Ireland Welcome to the 7th annual Advances in Metabolic Profiling conference and exhibition. This year's event will be held in the dynamic city of Dublin, economic and cultural centre of Ireland. The conference will be co-located with Mass Spec Europe. Registered delegates will have access to both meetings ensuring a very cost-effective trip. For more information, please visit http://www.selectbiosciences.com/conferences/AMP2011/.
13-16 Nov 2011	International Conference on Natural Products (ICNP2011) - Metabolomics A New Frontier in Natural Products Science Venue: Palm Garden Hotel, Putrajaya, Malaysia On behalf of the Organising Committee you are invited to participate in the next International Conference on Natural Products (ICNP2011) - Metabolomics A New Frontier in Natural Products Science. The conference is being organised by the Universiti Putra Malaysia and Malaysian Natural Products Society. The conference will be held at Palm Garden Hotel, IOI Resort, PUTRAJAYA from 13th -16th November 2011. For more information, please visit http://www.icnp2011.upm.edu.my/index.html.
25-28 June 2012	METABOLOMICS 2012: Breakthroughs in plant, microbial and human biology, clinical and nutritional research, and biomarker discovery Venue: Washington Marriott Wardman Park Hotel, Washington, DC, USA The Metabolomics Society is pleased to announce the location and dates for our next annual meeting 'METABOLOMICS 2012'. We will host a program full of practical workshops and parallel sessions covering the broad range of biological and technological metabolomics topics as well as provide rich opportunities for networking. Prominent scientists will speak on the state-of-the-art in a number of leading disciplines to kick off each session, after which, we will have a full agenda of innovative speakers with specific oral presentation opportunities provided for younger researchers. We invite you to reserve the above dates in your calendar and follow our website for further details, www.metabolomics2012.org. Local Organisers: Dan Bearden, Rick Beger, Rima Kaddurah-Daouk, Lloyd W. Sumner, Don Robertson, Padma Maruvada and Donna Kimball. For more information, please visit http://www.metabolomics2012.org.

Please note: If you know of any metabolomics lectures, meetings, workshops, or training sessions that we should feature in future issues of this newsletter, please email Ian Forsythe (metabolomics.innovation@gmail.com).

6) Metabolomics Jobs

This is a resource for advertising positions in metabolomics. If you have a job you would like posted in this newsletter, please email Ian Forsythe (metabolomics.innovation@gmail.com). Job postings will be carried for a maximum of 4 issues (8 weeks) unless the position is filled prior to that date.

Jobs Offered

Job Title	Employer	Location	Date Posted	Source
Postdoctoral Research Associate position to analyze plant composition	Iowa State University	Ames, Iowa	25-Aug-2011	Metabolomics Society Jobs
Postdoctoral Fellow in Metabolomics	Duke University	Durham, NC	20-Aug-2011	Metabolomics Society Jobs
Scientist, Proteomics/Metabolomics	Merck Sharp & Dohme (MSD) Singapore	Singapore	20-Aug-2011	JobsDB.com
Post-Doctoral Fellow: Biomarkers of Disease in Dairy Cows	University of Alberta	Edmonton, AB	3-Aug-2011	University of Alberta

Jobs Wanted
Very highly qualified individuals (e.g., lab managers, professors, directors, executives with extensive experience) seeking employment in metabolomics are encouraged to submit their position requests to Ian Forsythe (metabolomics.innovation@gmail.com). Upon review, a limited number of job submissions will be selected for publication in the Jobs Wanted section.

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Ian J. Forsythe, M.Sc. Editor MetaboNews Department of Computing Science University of Alberta 221 Athabasca Hall Edmonton, AB, T6G 2E8, Canada Email: metabolomics.innovation@gmail.com Website: http://www.metabonews.ca

This newsletter is produced by The Metabolomics Innovation Centre (TMIC, http://www.metabolomicscentre.ca/) for the benefit of the worldwide metabolomics community. MetaboNews is currently sponsored by TMIC: