title: Biomarker discovery in heterogeneous tissue samples -taking the in-silico deconfounding approach.
abstract: BACKGROUND
  For heterogeneous tissues, such as blood, measurements of gene expression are confounded by relative proportions of cell types involved. Conclusions have to rely on estimation of gene expression signals for homogeneous cell populations, e.g. by applying micro-dissection, fluorescence activated cell sorting, or in-silico deconfounding. We studied feasibility and validity of a non-negative matrix decomposition algorithm using experimental gene expression data for blood and sorted cells from the same donor samples. Our objective was to optimize the algorithm regarding detection of differentially expressed genes and to enable its use for classification in the difficult scenario of reversely regulated genes. This would be of importance for the identification of candidate biomarkers in heterogeneous tissues.
  
  
  RESULTS
  Experimental data and simulation studies involving noise parameters estimated from these data revealed that for valid detection of differential gene expression, quantile normalization and use of non-log data are optimal. We demonstrate the feasibility of predicting proportions of constituting cell types from gene expression data of single samples, as a prerequisite for a deconfounding-based classification approach.Classification cross-validation errors with and without using deconfounding results are reported as well as sample-size dependencies. Implementation of the algorithm, simulation and analysis scripts are available.
  
  
  CONCLUSIONS
  The deconfounding algorithm without decorrelation using quantile normalization on non-log data is proposed for biomarkers that are difficult to detect, and for cases where confounding by varying proportions of cell types is the suspected reason. In this case, a deconfounding ranking approach can be used as a powerful alternative to, or complement of, other statistical learning approaches to define candidate biomarkers for molecular diagnosis and prediction in biomedicine, in realistically noisy conditions and with moderate sample sizes.
authors.affiliation: Department of Genetics and Biometry, Research Institute for Biology of Farm Animals, Wilhelm-Stahl Allee 2, D 18196 Dummerstorf, Germany. repsilber@fbn-dummerstorf.de
authors.forename: Dirk
authors.initials: D
authors.lastname: Repsilber
authors.forename: Sabine
authors.initials: S
authors.lastname: Kern
authors.forename: Anna
authors.initials: A
authors.lastname: Telaar
authors.forename: Gerhard
authors.initials: G
authors.lastname: Walzl
authors.forename: Gillian F
authors.initials: GF
authors.lastname: Black
authors.forename: Joachim
authors.initials: J
authors.lastname: Selbig
authors.forename: Shreemanta K
authors.initials: SK
authors.lastname: Parida
authors.forename: Stefan H E
authors.initials: SH
authors.lastname: Kaufmann
authors.forename: Marc
authors.initials: M
authors.lastname: Jacobsen
mesh_terms: D000465:Algorithms; D015415:Biomarkers / Q000737:chemistry*; D019295:Computational Biology / Q000379:methods*; D020869:Gene Expression Profiling; D020411:Oligonucleotide Array Sequence Analysis
chemical_list: D015415:Biomarkers