IsoGeneGUI: a graphical user interface for analyzing dose-response - PowerPoint PPT Presentation

Background Test for Monotone Trend Software IsoGeneGUI Package IsoGeneGUI: a graphical user interface for analyzing dose-response studies in microarray experiments Research team: Setia Pramana 1 , Dan Lin 1 , Ziv Shkedy 1 , Tobias Verbeke 2 , Philippe Haldermans 1 ,Willem Talloen 3 . 1. IBioStat, Universiteit Hasselt, Belgium 2. OpenAnalytics, Belgium 3. Johnson & Johnson Pharmaceutical R&D, Belgium useR! 2010 Conference, 21 July 2010, Gaithersburg, Maryland, USA

Background Test for Monotone Trend Software IsoGeneGUI Package Overview Background: Dose-response Studies Test for Monotone Trend The Software Analysis using IsoGeneGUI Package Data exploration Likelihood Ratio Test E 2 analysis Resampling-based analysis Significance Analysis of Microarrays (SAM) User defined graphical display

Background Test for Monotone Trend Software IsoGeneGUI Package Dose-response studies The fundamental objective of drug development is to find a dose or a range of dose that 10 efficacious and safe. Dose-response study 9 gene expression investigates the dependence of 8 the response on doses. 7 Within a microarray setting the response of interest is gene 6 expression. 0 0.01 0.04 0.16 0.63 2.5 dose

Background Test for Monotone Trend Software IsoGeneGUI Package Dose-response studies in Microarray Experiments The aim is to identify a subset of genes with a monotone trend. PAVA In general, increasing the dose 3.5 will result in a proportional 3.0 increase in the response. 2.5 + * gene expression 2.0 Estimation: Isotonic regression with 1.5 * + + * * Pool-Adjacent-Violator-Algorithm 1.0 + * * (PAVA). 0.5 1 2 3 4 doses

Background Test for Monotone Trend Software IsoGeneGUI Package Testing for Monotonic Trend Hypotheses: For gene i ( i = 1 , · · · , m ) with K + 1 doses ( j = 0 , · · · , K ) H 0 : µ ( d 0 ) = µ ( d 1 ) = · · · = µ ( d K ) ( No Dose Effect ) H Up µ ( d 0 ) ≤ µ ( d 1 ) ≤ · · · ≤ µ ( d K ) ( Ordered alternative ) : 1 E 2 The likelihood ratio test ( ¯ 01 ). The t -type tests: Williams, Marcus, M and M ′ Lin et.al 2007

Background Test for Monotone Trend Software IsoGeneGUI Package Testing for Monotonic Trend: Inference Inference Exact Distribution Resampling-based Inference 2 LRT ( E ) Permutations Significance Analysis 01 Williams of Microarrays (SAM) Marcus 2 LRT ( E ) 01 2 SAM LRT ( E ) 01 Williams Williams SAM Marcus Marcus SAM M M SAM Modified M SAM Modified M ( M ' ) Both approaches will be discussed in the next slides

Background Test for Monotone Trend Software IsoGeneGUI Package Resampling-based Inference: Permutations Observed test statistics E 2 : E 2 ¯ 1 , ¯ E 2 2 , . . . , ¯ E 2 m Resampling-based Inference, the permutation matrix T : E 2 E 2 E 2 ¯ ¯ ¯   . . . 1 B 11 11 E 2 E 2 E 2 ¯ ¯ ¯ . . . 2 B   21 22 T =  . .  . .   . .   E 2 ¯ E 2 ¯ E 2 ¯ m 1 m 2 . . . mB , B is the number of permutations. Raw p -values: P i = #( b :¯ E 2 ib ≥ E 2 i ) . B

Background Test for Monotone Trend Software IsoGeneGUI Package Significance Analysis of Microarrays (SAM, Tusher et al., 2001) for microarrays under order restriction A modified score is assigned to each gene. For the t -type test statistics (i.e., Williams, Marcus, the M , and the M ′ ): Original: Modified: M ′ SAM = ˆ µ ⋆ K − ˆ µ ⋆ M ′ = ˆ µ ⋆ K − ˆ µ ⋆ 0 0 s ′ +ˆ s 0 s ′ ˆ ˆ s 0 is the fudge factor added to protect against inflated ˆ statistic due to small standard errors.

Background Test for Monotone Trend Software IsoGeneGUI Package Software Software for dose-response in microarrays analysis: R package: IsoGene, available in CRAN: http://cran.ii.uib.no/web/packages/IsoGene/ Pramana et.al 2010, The R Journal R GUI package: IsoGeneGUI, available in the Bioconductor project: http://bioconductor.org/packages/2.7/bioc/html/IsoGeneGUI.html

Background Test for Monotone Trend Software IsoGeneGUI Package IsoGeneGUI: Main Window Load Data Data Exploration Analysis: E 2 ¯ 01 exact distribution. Resampling-based Inference Multiplicity adjustment (FWER and FDR) Significant Analysis of Microarrays (SAM) Output & Graphical Displays

Background Test for Monotone Trend Software IsoGeneGUI Package IsoGeneGUI: Data Format The package can read data in the following formats: R workspace (*.RData files) Excel file (*.xls files) text file (*.txt files)

Background Test for Monotone Trend Software IsoGeneGUI Package IsoGeneGUI: Data Exploration Exploratory Data Analysis The summary statistic The Isotonic regression plots

Background Test for Monotone Trend Software IsoGeneGUI Package E 2 IsoGeneGUI: The ¯ 01 Analysis The ¯ E 2 Antipsychotic study. 01 test and BH-FDR (0.05) 6 dose levels are specified: 4-5 samples at each dose level. 11,565 genes. The dataset is available inside the package. 10 9 gene expression 8 7 6 0 0.01 0.04 0.16 0.63 2.5 dose

Background Test for Monotone Trend Software IsoGeneGUI Package E 2 IsoGeneGUI: ¯ 01 Result E 2 ¯ 01 with BH-FDR (0.05). 298 genes with significant monotone trend

Background Test for Monotone Trend Software IsoGeneGUI Package IsoGeneGUI: List of Significant Genes List significant genes. The list be saved as an R object or/and an excel file

Background Test for Monotone Trend Software IsoGeneGUI Package IsoGeneGUI: Volcano Plot E 2 ¯ 01 vs. Fold change The graph can be copied into clipboard and be saved into different image types.

Background Test for Monotone Trend Software IsoGeneGUI Package IsoGeneGUI: Resampling-based Inference Required specification: number of permutations, test statistic and multiplicity adjustment. The permutation result can be saved and used later. List of significant genes using the selected statistic(s). Graphical displays.

Background Test for Monotone Trend Software IsoGeneGUI Package IsoGeneGUI: SAM (specification) SAM permutation dialog box Required : Number of permutations and the fudge factor. SAM regularized test statistic, e.g.,: M ′ SAM = ˆ µ ⋆ K − ˆ µ ⋆ 0 s ′ +ˆ s 0 ˆ Delta ( ∆ ) is chosen with estimation of the FDR.

Background Test for Monotone Trend Software IsoGeneGUI Package IsoGeneGUI: SAM (Results) Threshold/Delta: 0.68 FDR: 0.0436 Number of significant genes: 164 Delta table:

Background Test for Monotone Trend Software IsoGeneGUI Package User Defined Graphical Display User can define their own figures:

Background Test for Monotone Trend Software IsoGeneGUI Package Help Html help with screen shots

Background Test for Monotone Trend Software IsoGeneGUI Package More detail... Book of Lin, D., Shkedy, Z., Yekutieli, D., Amaratunga, D., and Bijnens., L., editors. Modeling Dose-response Microarray Data in Early Drug Development Experiments Using R . Springer. To be published in 2010. IsoGene Project site: http://www.ibiostat.be/software/IsoGeneGUI/index.html Thank you for your attention...