| Title: | BLMA: A package for bi-level meta-analysis |
|---|---|
| Description: | Suit of tools for bi-level meta-analysis. The package can be used in a wide range of applications, including general hypothesis testings, differential expression analysis, functional analysis, and pathway analysis. |
| Authors: | Tin Nguyen <[email protected]>, Hung Nguyen <[email protected]>, and Sorin Draghici <[email protected]> |
| Maintainer: | Van-Dung Pham <[email protected]> |
| License: | GPL (>=2) |
| Version: | 1.31.0 |
| Built: | 2024-10-30 04:28:31 UTC |
| Source: | https://github.com/bioc/BLMA |
Combine independent studies using the average of p-values
addCLT(x)addCLT(x)
x |
is an array of independent p-values |
This method is based on the fact that sum of independent uniform variables follow the Irwin-Hall distribution [1a,1b]. When the number of p-values is small (n<20), the distribution of the average of p-values can be calculated using a linear transformation of the Irwin-Hall distribution. When n is large, the distribution is approximated using the Central Limit Theorem to avoid underflow/overflow problems [2,3,4,5].
combined p-value
Tin Nguyen and Sorin Draghici
[1a] P. Hall. The distribution of means for samples of size n drawn from a population in which the variate takes values between 0 and 1, all such values being equally probable. Biometrika, 19(3-4):240-244, 1927.
[1b] J. O. Irwin. On the frequency distribution of the means of samples from a population having any law of frequency with finite moments, with special reference to Pearson's Type II. Biometrika, 19(3-4):225-239, 1927.
[2] T. Nguyen, R. Tagett, M. Donato, C. Mitrea, and S. Draghici. A novel bi-level meta-analysis approach – applied to biological pathway analysis. Bioinformatics, 32(3):409-416, 2016.
[3] T. Nguyen, C. Mitrea, R. Tagett, and S. Draghici. DANUBE: Data-driven meta-ANalysis using UnBiased Empirical distributions – applied to biological pathway analysis. Proceedings of the IEEE, PP(99):1-20, 2016.
[4] T. Nguyen, D. Diaz, R. Tagett, and S. Draghici. Overcoming the matched-sample bottleneck: an orthogonal approach to integrate omic data. Scientific Reports, 6:29251, 2016.
[5] T. Nguyen, D. Diaz, and S. Draghici. TOMAS: A novel TOpology-aware Meta-Analysis approach applied to System biology. In Proceedings of the 7th ACM International Conference on Bioin- formatics, Computational Biology, and Health Informatics, pages 13-22. ACM, 2016.
x <- rep(0,10) addCLT(x) x <- runif(10) addCLT(x)x <- rep(0,10) addCLT(x) x <- runif(10) addCLT(x)
Perform a bi-level meta-analysis in conjunction with any of the classical hypothesis testing methods, such as t-test, Wilcoxon test, etc.
bilevelAnalysisClassic(x, y = NULL, splitSize = 5, metaMethod = addCLT, func = t.test, p.value = "p.value", ...)bilevelAnalysisClassic(x, y = NULL, splitSize = 5, metaMethod = addCLT, func = t.test, p.value = "p.value", ...)
x |
a list of numeric vectors |
y |
an optional list of numeric vectors |
splitSize |
the minimum number of size in each split sample. splitSize should be at least 3. By default, splitSize=5 |
metaMethod |
the method used to combine p-values. This should be one of addCLT (additive method [1]), fishersMethod (Fisher's method [5]), stoufferMethod (Stouffer's method [6]), max (maxP method [7]), or min (minP method [8]) |
func |
the name of the hypothesis test. By default func=t.test |
p.value |
the component that returns the p-value after performing the test provided by the func parameter. For example, the function t-test returns the class "htest" where the component "p.value" is the p-value of the test. By default, p.value="p.value" |
... |
additional parameters for func |
This function performs a bi-level meta-analysis for the lists
of samples [1]. It performs intra-experiment analyses to compare the
vectors in x agains the corresponding vectors in y using the function
intraAnalysisClassic in conjunction with the test provided
in func. For example, it compares the first vector in x with the
first vector in y, the second vector in x with the second vector in y, etc.
When y is null, then the comparisons are reduced to one-sample tests. After
these comparisons, we have a list of p-values, one for each comparision.
The function then combines these p-values to obtain a single p-value
using metaMethod.
the combined p-value
Tin Nguyen and Sorin Draghici
[1] T. Nguyen, R. Tagett, M. Donato, C. Mitrea, and S. Draghici. A novel bi-level meta-analysis approach – applied to biological pathway analysis. Bioinformatics, 32(3):409-416, 2016.
intraAnalysisClassic, intraAnalysisGene, bilevelAnalysisGene
set.seed(1) l1 <- lapply(as.list(seq(3)),FUN=function (x) rnorm(n=10, mean=1)) l1 # one-sample t-test lapply(l1, FUN=function(x) t.test(x, alternative="greater")$p.value) # combining the p-values of one-sample t-tests: addCLT(unlist(lapply(l1, FUN=function(x) t.test(x, alter="g")$p.value))) #Bi-level meta-analysis bilevelAnalysisClassic(x=l1, alternative="greater")set.seed(1) l1 <- lapply(as.list(seq(3)),FUN=function (x) rnorm(n=10, mean=1)) l1 # one-sample t-test lapply(l1, FUN=function(x) t.test(x, alternative="greater")$p.value) # combining the p-values of one-sample t-tests: addCLT(unlist(lapply(l1, FUN=function(x) t.test(x, alter="g")$p.value))) #Bi-level meta-analysis bilevelAnalysisClassic(x=l1, alternative="greater")
Perform a bi-level meta-analysis in conjunction with the moderate t-test (limma package) for the purpose of differential expression analysis of multiple gene expression datasets
bilevelAnalysisGene(dataList, groupList, splitSize = 5, metaMethod = addCLT)bilevelAnalysisGene(dataList, groupList, splitSize = 5, metaMethod = addCLT)
dataList |
a list of datasets. Each dataset is a data frame where the rows are the gene IDs and the columns are the samples |
groupList |
a list of vectors. Each vector represents the phenotypes of the corresponding dataset in dataList, which are either 'c' (control) or 'd' (disease). |
splitSize |
the minimum number of disease samples in each split dataset. splitSize should be at least 3. By default, splitSize=5 |
metaMethod |
the method used to combine p-values. This should be one of addCLT (additive method [1]), fishersMethod (Fisher's method [5]), stoufferMethod (Stouffer's method [6]), max (maxP method [7]), or min (minP method [8]) |
The bi-level framework combines the datasets at two levels: an intra- experiment analysis, and an inter-experiment analysis [1]. At the intra-experiment analysis, the framework splits a dataset into smaller datasets, performs a moderated t-test (limma package) on split datasets, and then combines p-values of individual genes using metaMethod. At the inter-experiment analysis, the p-values obtained for each individual datasets are combined using metaMethod
A data frame containing the following components:
rownames: gene IDs that are common in all datasets
pLimma: the p-values obtained by combining pLimma values of individual datasets
pLimma.fdr: FDR-corrected p-values of pLimma
pBilevel: the p-values obtained from combining pIntra values of individual datasets
pBilevel.fdr: FDR-corrected p-values of pBilevel
Tin Nguyen and Sorin Draghici
[1] T. Nguyen, R. Tagett, M. Donato, C. Mitrea, and S. Draghici. A novel bi-level meta-analysis approach – applied to biological pathway analysis. Bioinformatics, 32(3):409-416, 2016.
bilevelAnalysisGene, intraAnalysisClassic
dataSets <- c("GSE17054", "GSE57194", "GSE33223", "GSE42140") data(list=dataSets, package="BLMA") names(dataSets) <- dataSets dataList <- lapply(dataSets, function(dataset) get(paste0("data_", dataset))) groupList <- lapply(dataSets, function(dataset) get(paste0("group_", dataset))) Z <- bilevelAnalysisGene(dataList = dataList, groupList = groupList) head(Z)dataSets <- c("GSE17054", "GSE57194", "GSE33223", "GSE42140") data(list=dataSets, package="BLMA") names(dataSets) <- dataSets dataList <- lapply(dataSets, function(dataset) get(paste0("data_", dataset))) groupList <- lapply(dataSets, function(dataset) get(paste0("group_", dataset))) Z <- bilevelAnalysisGene(dataList = dataList, groupList = groupList) head(Z)
Perform a bi-level meta-analysis in conjunction with geneset enrichment methods (ORA/GSA/PADOG) to integrate multiple gene expression datasets.
bilevelAnalysisGeneset(gslist, gs.names, dataList, groupList, splitSize = 5, metaMethod = addCLT, enrichment = "ORA", pCutoff = 0.05, percent = 0.05, mc.cores = 1, ...)bilevelAnalysisGeneset(gslist, gs.names, dataList, groupList, splitSize = 5, metaMethod = addCLT, enrichment = "ORA", pCutoff = 0.05, percent = 0.05, mc.cores = 1, ...)
gslist |
a list of gene sets. |
gs.names |
names of the gene sets. |
dataList |
a list of datasets to be combined. Each dataset is a data frame where the rows are the gene IDs and the columns are the samples. |
groupList |
a list of vectors. Each vector represents the phenotypes of the corresponding dataset in dataList. The elements of each vector are either 'c' (control) or 'd' (disease). |
splitSize |
the minimum number of disease samples in each split dataset. splitSize should be at least 3. By default, splitSize=5 |
metaMethod |
the method used to combine p-values. This should be one of addCLT (additive method [1]), fisherMethod (Fisher's method [5]), stoufferMethod (Stouffer's method [6]), max (maxP method [7]), or min (minP method [8]) |
enrichment |
the method used for enrichment analysis. This should be one of "ORA", "GSA", or "PADOG". By default, enrichment is set to "ORA". |
pCutoff |
cutoff p-value used to identify differentially expressed (DE) genes. This parameter is used only when the enrichment method is "ORA". By default, pCutoff=0.05 (five percent) |
percent |
percentage of genes with highest foldchange to be considered as differentially expressed (DE). This parameter is used when the enrichment method is "ORA". By default percent=0.05 (five percent). Please note that only genes with p-value less than pCutoff will be considered |
mc.cores |
the number of cores to be used in parallel computing. By default, mc.cores=1 |
... |
additional parameters of the GSA/PADOG functions |
The bi-level framework combines the datasets at two levels: an intra- experiment analysis, and an inter-experiment analysis [1]. At the intra-level analysis, the framework splits a dataset into smaller datasets, performs enrichment analysis for each split dataset (using ORA [2], GSA [3], or PADOG [4]), and then combines the results of these split datasets using metaMethod. At the inter-level analysis, the results obtained for individual datasets are combined using metaMethod
A data frame (rownames are geneset/pathway IDs) that consists of the following information:
Name: name/description of the corresponding pathway/geneset
Columns that include the pvalues obtained from the intra-experiment analysis of individual datasets
pBLMA: p-value obtained from the inter-experiment analysis using addCLT
rBLMA: ranking of the geneset/pathway using addCLT
pBLMA.fdr: FDR-corrected p-values
Tin Nguyen and Sorin Draghici
[1] T. Nguyen, R. Tagett, M. Donato, C. Mitrea, and S. Draghici. A novel bi-level meta-analysis approach – applied to biological pathway analysis. Bioinformatics, 32(3):409-416, 2016.
[2] S. Draghici, P. Khatri, R. P. Martin, G. C. Ostermeier, and S. A. Krawetz. Global functional profiling of gene expression. Genomics, 81(2):98-104, 2003.
[3] B. Efron and R. Tibshirani. On testing the significance of sets of genes. The Annals of Applied Statistics, 1(1):107-129, 2007.
[4] A. L. Tarca, S. Draghici, G. Bhatti, and R. Romero. Down-weighting overlapping genes improves gene set analysis. BMC Bioinformatics, 13(1):136, 2012.
[5] R. A. Fisher. Statistical methods for research workers. Oliver & Boyd, Edinburgh, 1925.
[6] S. Stouffer, E. Suchman, L. DeVinney, S. Star, and J. Williams, RM. The American Soldier: Adjustment during army life, volume 1. Princeton University Press, Princeton, 1949.
[7] L. H. C. Tippett. The methods of statistics. The Methods of Statistics, 1931.
[8] B. Wilkinson. A statistical consideration in psychological research. Psychological Bulletin, 48(2):156, 1951.
bilevelAnalysisPathway, phyper, GSA, padog
# load KEGG pathways and create gene sets x <- loadKEGGPathways() gslist <- lapply(x$kpg,FUN=function(y){return (nodes(y));}) gs.names <- x$kpn[names(gslist)] # load example data dataSets <- c("GSE17054", "GSE57194", "GSE33223", "GSE42140") data(list=dataSets, package="BLMA") names(dataSets) <- dataSets dataList <- lapply(dataSets, function(dataset) get(paste0("data_", dataset))) groupList <- lapply(dataSets, function(dataset) get(paste0("group_", dataset))) # perform bi-level meta-analysis in conjunction with ORA ORAComb <- bilevelAnalysisGeneset(gslist, gs.names, dataList, groupList, enrichment = "ORA") head(ORAComb[, c("Name", "pBLMA", "pBLMA.fdr", "rBLMA")]) # perform bi-level meta-analysis in conjunction with GSA GSAComb <- bilevelAnalysisGeneset(gslist, gs.names, dataList, groupList, enrichment = "GSA", nperms = 200, random.seed = 1) head(GSAComb[, c("Name", "pBLMA", "pBLMA.fdr", "rBLMA")]) # perform bi-level meta-analysi in conjunction with PADOG set.seed(1) PADOGComb <- bilevelAnalysisGeneset(gslist, gs.names, dataList, groupList, enrichment = "PADOG", NI=200) head(PADOGComb[, c("Name", "pBLMA", "pBLMA.fdr", "rBLMA")])# load KEGG pathways and create gene sets x <- loadKEGGPathways() gslist <- lapply(x$kpg,FUN=function(y){return (nodes(y));}) gs.names <- x$kpn[names(gslist)] # load example data dataSets <- c("GSE17054", "GSE57194", "GSE33223", "GSE42140") data(list=dataSets, package="BLMA") names(dataSets) <- dataSets dataList <- lapply(dataSets, function(dataset) get(paste0("data_", dataset))) groupList <- lapply(dataSets, function(dataset) get(paste0("group_", dataset))) # perform bi-level meta-analysis in conjunction with ORA ORAComb <- bilevelAnalysisGeneset(gslist, gs.names, dataList, groupList, enrichment = "ORA") head(ORAComb[, c("Name", "pBLMA", "pBLMA.fdr", "rBLMA")]) # perform bi-level meta-analysis in conjunction with GSA GSAComb <- bilevelAnalysisGeneset(gslist, gs.names, dataList, groupList, enrichment = "GSA", nperms = 200, random.seed = 1) head(GSAComb[, c("Name", "pBLMA", "pBLMA.fdr", "rBLMA")]) # perform bi-level meta-analysi in conjunction with PADOG set.seed(1) PADOGComb <- bilevelAnalysisGeneset(gslist, gs.names, dataList, groupList, enrichment = "PADOG", NI=200) head(PADOGComb[, c("Name", "pBLMA", "pBLMA.fdr", "rBLMA")])
Perform a bi-level meta-analysis conjunction with Impact Analysis to integrate multiple gene expression datasets
bilevelAnalysisPathway(kpg, kpn, dataList, groupList, splitSize = 5, metaMethod = addCLT, pCutoff = 0.05, percent = 0.05, mc.cores = 1, nboot = 200, seed = 1)bilevelAnalysisPathway(kpg, kpn, dataList, groupList, splitSize = 5, metaMethod = addCLT, pCutoff = 0.05, percent = 0.05, mc.cores = 1, nboot = 200, seed = 1)
kpg |
list of pathway graphs as objects of type graph
(e.g., |
kpn |
names of the pathways. |
dataList |
a list of datasets to be combined. Each dataset is a data frame where the rows are the gene IDs and the columns are the samples. |
groupList |
a list of vectors. Each vector represents the phenotypes of the corresponding dataset in dataList, which are either 'c' (control) or 'd' (disease). |
splitSize |
the minimum number of disease samples in each split dataset. splitSize should be at least 3. By default, splitSize=5 |
metaMethod |
the method used to combine p-values. This should be one of addCLT (additive method [1]), fisherMethod (Fisher's method [5]), stoufferMethod (Stouffer's method [6]), max (maxP method [7]), or min (minP method [8]) |
pCutoff |
cutoff p-value used to identify differentially expressed (DE) genes. This parameter is used only when the enrichment method is "ORA". By default, pCutoff=0.05 (five percent) |
percent |
percentage of genes with highest foldchange to be considered as differentially expressed (DE). This parameter is used when the enrichment method is "ORA". By default percent=0.05 (five percent). Please note that only genes with p-value less than pCutoff will be considered |
mc.cores |
the number of cores to be used in parallel computing. By default, mc.cores=1 |
nboot |
number of bootstrap iterations. By default, nboot=200 |
seed |
seed. By default, seed=1. |
The bi-level framework combines the datasets at two levels: an intra-experiment analysis, and an inter-experiment analysis [1]. At the intra-level analysis, the framework splits a dataset into smaller datasets, performs pathway analysis for each split dataset using Impact Analysis [2,3], and then combines the results of these split datasets using metaMethod. At the inter-level analysis, the results obtained for individual datasets are combined using metaMethod
A data frame (rownames are geneset/pathway IDs) that consists of the following information:
Name: name/description of the corresponding pathway/geneset
Columns that include the pvalues obtained from the intra-experiment analysis of individual datasets
pBLMA: p-value obtained from the inter-experiment analysis using addCLT
rBLMA: ranking of the geneset/pathway using addCLT
pBLMA.fdr: FDR-corrected p-values
Tin Nguyen and Sorin Draghici
[1] T. Nguyen, R. Tagett, M. Donato, C. Mitrea, and S. Draghici. A novel bi-level meta-analysis approach – applied to biological pathway analysis. Bioinformatics, 32(3):409-416, 2016.
[2] A. L. Tarca, S. Draghici, P. Khatri, S. S. Hassan, P. Mittal, J.-s. Kim, C. J. Kim, J. P. Kusanovic, and R. Romero. A novel signaling pathway impact analysis. Bioinformatics, 25(1):75-82, 2009.
[3] S. Draghici, P. Khatri, A. L. Tarca, K. Amin, A. Done, C. Voichita, C. Georgescu, and R. Romero. A systems biology approach for pathway level analysis. Genome Research, 17(10):1537-1545, 2007.
[4] R. A. Fisher. Statistical methods for research workers. Oliver & Boyd, Edinburgh, 1925.
[5] S. Stouffer, E. Suchman, L. DeVinney, S. Star, and J. Williams, RM. The American Soldier: Adjustment during army life, volume 1. Princeton University Press, Princeton, 1949.
[6] L. H. C. Tippett. The methods of statistics. The Methods of Statistics, 1931.
[7] B. Wilkinson. A statistical consideration in psychological research. Psychological Bulletin, 48(2):156, 1951.
bilevelAnalysisGeneset, pe, phyper
# load KEGG pathways x <- loadKEGGPathways() # load example data dataSets <- c("GSE17054", "GSE57194", "GSE33223", "GSE42140") data(list=dataSets, package="BLMA") names(dataSets) <- dataSets dataList <- lapply(dataSets, function(dataset) get(paste0("data_", dataset))) groupList <- lapply(dataSets, function(dataset) get(paste0("group_", dataset))) IAComb <- bilevelAnalysisPathway(x$kpg, x$kpn, dataList, groupList) head(IAComb[, c("Name", "pBLMA", "pBLMA.fdr", "rBLMA")])# load KEGG pathways x <- loadKEGGPathways() # load example data dataSets <- c("GSE17054", "GSE57194", "GSE33223", "GSE42140") data(list=dataSets, package="BLMA") names(dataSets) <- dataSets dataList <- lapply(dataSets, function(dataset) get(paste0("data_", dataset))) groupList <- lapply(dataSets, function(dataset) get(paste0("group_", dataset))) IAComb <- bilevelAnalysisPathway(x$kpg, x$kpn, dataList, groupList) head(IAComb[, c("Name", "pBLMA", "pBLMA.fdr", "rBLMA")])
Combine independent p-values using the minus log product
fisherMethod(x)fisherMethod(x)
x |
is an array of independent p-values |
Considering a set of m independent significance tests, the resulted p-values are independent and uniformly distributed between 0 and 1 under the null hypothesis. Fisher's method uses the minus log product of the p-values as the summary statistic, which follows a chi-square distribution with 2m degrees of freedom. This chi-square distribution is used to calculate the combined p-value.
combined p-value
Tin Nguyen and Sorin Draghici
[1] R. A. Fisher. Statistical methods for research workers. Oliver & Boyd, Edinburgh, 1925.
x <- rep(0,10) fisherMethod(x) x <- runif(10) fisherMethod(x)x <- rep(0,10) fisherMethod(x) x <- runif(10) fisherMethod(x)
Calculate genes summary statistic across multiple datasets
getStatistics(allGenes, dataList, groupList, ncores = 1, method = addCLT)getStatistics(allGenes, dataList, groupList, ncores = 1, method = addCLT)
allGenes |
Vector of all genes names for the analysis. |
dataList |
A list of expression matrices, in which rows are genes and columns are samples. |
groupList |
A list of vectors indicating sample group corresponding with expression matrices in dataList. |
ncores |
Number of core to use in prallel processing. |
method |
Function for combining p-values. It must accept one input which is a vector of p-values and return a combined p-value. Three methods are embeded in this package are addCLT, fisherMethod, and stoufferMethod. |
To estimate the effect sizes of genes across all studies, first standardized mean difference for each gene in individual studies is compute. Next, the overall efect size and standard error are estimated using the random-efects model. This overall efect size represents the gene's expression change under the efect of the condition. The, z-scores and p-values of observing such efect sizes are computed. The p-values is obtained from classical hypothesis testing. By default, linear model and empirical Bayesian testing \(limma\) are used to compute the p-values for diferential expression. The two-tailed p-values are converted to one-tailed p-values (lef- and right-tailed). For each gene, the one-tailed p-values across all datasets are then combined using the addCLT, stouffer or fisher method. These p-values represent how likely the diferential expression is observed by chance.
A data.frame of gene statistics with following columns:
Two-tailed p-values
Two-tailed p-values with false discovery rate correction
left-tailed p-values
left-tailed p-values with false discovery rate correction
right-tailed p-values with false discovery rate correction
right-tailed p-values
Effect size
Two-tailed p-values for effect size
Two-tailed p-values for effect size with false discovery rate correction
Left-tailed p-values for effect size
Left-tailed p-values for effect size with false discovery rate correction
Right-tailed p-values for effect size
Right-tailed p-values for effect size with false discovery rate correction
Tin Nguyen, Hung Nguyen, and Sorin Draghici
Nguyen, T., Shafi, A., Nguyen, T. M., Schissler, A. G., & Draghici, S. (2020). NBIA: a network-based integrative analysis framework-applied to pathway analysis. Scientific reports, 10(1), 1-11. Nguyen, T., Tagett, R., Donato, M., Mitrea, C., & Draghici, S. (2016). A novel bi-level meta-analysis approach: applied to biological pathway analysis. Bioinformatics, 32(3), 409-416. Smyth, G. K. (2005). Limma: linear models for microarray data. In Bioinformatics and computational biology solutions using R and Bioconductor (pp. 397-420). Springer, New York, NY.
datasets <- c("GSE17054", "GSE57194", "GSE33223", "GSE42140") data(list = datasets, package = "BLMA") dataList <- lapply(datasets, function(dataset) { get(paste0("data_", dataset)) }) groupList <- lapply(datasets, function(dataset) { get(paste0("group_", dataset)) }) names(dataList) <- datasets names(groupList) <- datasets allGenes <- Reduce(intersect, lapply(dataList, rownames)) geneStat <- getStatistics(allGenes, dataList, groupList) head(geneStat) # perform pathway analysis library(ROntoTools) # get gene network kpg <- loadKEGGPathways()$kpg # get gene network name kpn <- loadKEGGPathways()$kpn # get geneset gslist <- lapply(kpg,function(y) nodes(y)) # get differential expressed genes DEGenes.Left <- rownames(geneStat)[geneStat$pLeft < 0.05 & geneStat$ES.pLeft < 0.05] DEGenes.Right <- rownames(geneStat)[geneStat$pRight < 0.05 & geneStat$ES.pRight < 0.05] DEGenes <- union(DEGenes.Left, DEGenes.Right) # perform pathway analysis with ORA oraRes <- lapply(gslist, function(gs){ pORACalc(geneSet = gs, DEGenes = DEGenes, measuredGenes = rownames(geneStat)) }) oraRes <- data.frame(p.value = unlist(oraRes), pathway = names(oraRes)) rownames(oraRes) <- kpn[rownames(oraRes)] # print results print(head(oraRes)) # perfrom pathway analysis with Pathway-Express from ROntoTools ES <- geneStat[DEGenes, "ES"] names(ES) <- DEGenes peRes = pe(x = ES, graphs = kpg, ref = allGenes, nboot = 1000, seed=1) peRes.Summary <- Summary(peRes, comb.pv.func = fisherMethod) peRes.Summary[, ncol(peRes.Summary) + 1] <- rownames(peRes.Summary) rownames(peRes.Summary) <- kpn[rownames(peRes.Summary)] colnames(peRes.Summary)[ncol(peRes.Summary)] = "pathway" # print results print(head(peRes.Summary))datasets <- c("GSE17054", "GSE57194", "GSE33223", "GSE42140") data(list = datasets, package = "BLMA") dataList <- lapply(datasets, function(dataset) { get(paste0("data_", dataset)) }) groupList <- lapply(datasets, function(dataset) { get(paste0("group_", dataset)) }) names(dataList) <- datasets names(groupList) <- datasets allGenes <- Reduce(intersect, lapply(dataList, rownames)) geneStat <- getStatistics(allGenes, dataList, groupList) head(geneStat) # perform pathway analysis library(ROntoTools) # get gene network kpg <- loadKEGGPathways()$kpg # get gene network name kpn <- loadKEGGPathways()$kpn # get geneset gslist <- lapply(kpg,function(y) nodes(y)) # get differential expressed genes DEGenes.Left <- rownames(geneStat)[geneStat$pLeft < 0.05 & geneStat$ES.pLeft < 0.05] DEGenes.Right <- rownames(geneStat)[geneStat$pRight < 0.05 & geneStat$ES.pRight < 0.05] DEGenes <- union(DEGenes.Left, DEGenes.Right) # perform pathway analysis with ORA oraRes <- lapply(gslist, function(gs){ pORACalc(geneSet = gs, DEGenes = DEGenes, measuredGenes = rownames(geneStat)) }) oraRes <- data.frame(p.value = unlist(oraRes), pathway = names(oraRes)) rownames(oraRes) <- kpn[rownames(oraRes)] # print results print(head(oraRes)) # perfrom pathway analysis with Pathway-Express from ROntoTools ES <- geneStat[DEGenes, "ES"] names(ES) <- DEGenes peRes = pe(x = ES, graphs = kpg, ref = allGenes, nboot = 1000, seed=1) peRes.Summary <- Summary(peRes, comb.pv.func = fisherMethod) peRes.Summary[, ncol(peRes.Summary) + 1] <- rownames(peRes.Summary) rownames(peRes.Summary) <- kpn[rownames(peRes.Summary)] colnames(peRes.Summary)[ncol(peRes.Summary)] = "pathway" # print results print(head(peRes.Summary))
This dataset consists of 5 acute myeloid leukemia and 4 control samples. The data frame data_GSE17054 includes the expression data while the vector group_GSE17054 includes the grouping information.
data(GSE17054)data(GSE17054)
data_GSE17054 is a data frame with 4738 rows and 9 columns. The rows represent the genes and the columns represent the samples.
group_GSE17054 is a vector that represents the sample grouping for data_GSE17054. The elements of group_GSE17054 are either 'c' (control) or 'd' (disease).
Obtained from http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE17054
Majeti et al. Dysregulated gene expression networks in human acute myelogenous leukemia stem cells. Proceedings of the National Academy of Sciences, 106(9):3396-3401, 2009.
This dataset consists of 20 acute myeloid leukemia and 10 control samples. The data frame data_GSE33223 includes the expression data while the vector group_GSE33223 includes the grouping information.
data(GSE33223)data(GSE33223)
data_GSE33223 is a data frame with 4114 rows and 30 columns. The rows represent the genes and the columns represent the samples.
group_GSE33223 is a vector that represents the sample grouping for data_GSE33223. The elements of group_GSE33223 are either 'c' (control) or 'd' (disease).
Obtained from https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE33223
Bacher et al. Multilineage dysplasia does not influence prognosis in CEBPA-mutated AML, supporting the WHO proposal to classify these patients as a unique entity. Blood, 119(20):4719-22, 2012.
This dataset consists of 26 acute myeloid leukemia and 5 control samples. The data frame data_GSE42140 includes the expression data while the vector group_GSE42140 includes the grouping information.
data(GSE42140)data(GSE42140)
data_GSE42140 is a data frame with 4114 rows and 31 columns. The rows represent the genes and the columns represent the samples.
group_GSE42140 is a vector that represents the sample grouping for data_GSE42140. The elements of group_GSE42140 are either 'c' (control) or 'd' (disease).
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE42140
This dataset consists of 6 acute myeloid leukemia and 6 control samples. The data frame data_GSE57194 includes the expression data while the vector group_GSE57194 includes the grouping information.
data(GSE57194)data(GSE57194)
data_GSE57194 is a data frame with 4114 rows and 12 columns. The rows represent the genes and the columns represent the samples.
group_GSE57194 is a vector that represents the sample grouping for data_GSE57194. The elements of group_GSE57194 are either 'c' (control) or 'd' (disease).
Obtained from https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE57194
Abdul-Nabi et al. In vitro transformation of primary human CD34+ cells by AML fusion oncogenes: early gene expression profiling reveals possible drug target in AML. PLoS One, 5(8):e12464, 2010.
Perform an intra-experiment analysis in conjunction with any of the classical hypothesis testing methods, such as t-test, Wilcoxon test, etc.
intraAnalysisClassic(x, y = NULL, splitSize = 5, metaMethod = addCLT, func = t.test, p.value = "p.value", ...)intraAnalysisClassic(x, y = NULL, splitSize = 5, metaMethod = addCLT, func = t.test, p.value = "p.value", ...)
x |
a numeric vector of data values |
y |
an optional numeric vector of values |
splitSize |
the minimum number of size in each split sample. splitSize should be at least 3. By default, splitSize=5 |
metaMethod |
the method used to combine p-values. This should be one of addCLT (additive method [1]), fishersMethod (Fisher's method [5]), stoufferMethod (Stouffer's method [6]), max (maxP method [7]), or min (minP method [8]) |
func |
the name of the hypothesis test. By default func=t.test |
p.value |
the component that returns the p-value after performing the test provided by the func parameter. For example, the function t-test returns the class "htest" where the component "p.value" is the p-value of the test. By default, p.value="p.value" |
... |
additional parameters for func |
This function performs an intra-experiment analysis for the given sample(s) [1]. Given x as the numeric vector, this function first splits x into smaller samples with size splitSize, performs hypothesis testing using func, and then combines the p-values using metaMethod
intra-experiment p-value
Tin Nguyen and Sorin Draghici
[1] T. Nguyen, R. Tagett, M. Donato, C. Mitrea, and S. Draghici. A novel bi-level meta-analysis approach – applied to biological pathway analysis. Bioinformatics, 32(3):409-416, 2016.
bilevelAnalysisClassic, intraAnalysisGene, bilevelAnalysisGene
set.seed(1) x <- rnorm(10, mean = 0) # p-value obtained from a one-sample t-test t.test(x, mu=1, alternative = "less")$p.value # p-value obtained from an intra-experiment analysis intraAnalysisClassic(x, func=t.test, mu=1, alternative = "less") # p-value obtained from a one-sample wilcoxon test wilcox.test(x, mu=1, alternative = "less")$p.value # p-value obtained from an intra-experiment analysis intraAnalysisClassic(x, func=wilcox.test, mu=1, alternative = "less") set.seed(1) x <- rnorm(20, mean=0); y <- rnorm(20, mean=1) # p-value obtained from a two-sample t-test t.test(x,y,alternative="less")$p.value # p-value obtained from an intra-experiment analysis intraAnalysisClassic(x, y, func=t.test, alternative = "less") # p-value obtained from a two-sample wilcoxon test wilcox.test(x,y,alternative="less")$p.value # p-value obtained from an intra-experiment analysis intraAnalysisClassic(x, y, func=wilcox.test, alternative = "less")set.seed(1) x <- rnorm(10, mean = 0) # p-value obtained from a one-sample t-test t.test(x, mu=1, alternative = "less")$p.value # p-value obtained from an intra-experiment analysis intraAnalysisClassic(x, func=t.test, mu=1, alternative = "less") # p-value obtained from a one-sample wilcoxon test wilcox.test(x, mu=1, alternative = "less")$p.value # p-value obtained from an intra-experiment analysis intraAnalysisClassic(x, func=wilcox.test, mu=1, alternative = "less") set.seed(1) x <- rnorm(20, mean=0); y <- rnorm(20, mean=1) # p-value obtained from a two-sample t-test t.test(x,y,alternative="less")$p.value # p-value obtained from an intra-experiment analysis intraAnalysisClassic(x, y, func=t.test, alternative = "less") # p-value obtained from a two-sample wilcoxon test wilcox.test(x,y,alternative="less")$p.value # p-value obtained from an intra-experiment analysis intraAnalysisClassic(x, y, func=wilcox.test, alternative = "less")
perform an intra-experiment analysis in conjunction with the moderated t-test (limma package) for the purpose of differential expression analysis of a gene expression dataset
intraAnalysisGene(data, group, splitSize = 5, metaMethod = addCLT)intraAnalysisGene(data, group, splitSize = 5, metaMethod = addCLT)
data |
a data frame where the rows are the gene IDs and the columns are the samples |
group |
sample grouping. The elements of group are either 'c' (control) or 'd' (disease). names(group) should be identical to colnames(data) |
splitSize |
the minimum number of disease samples in each split dataset. splitSize should be at least 3. By default, splitSize=5 |
metaMethod |
the method used to combine p-values. This should be one of addCLT (additive method [1]), fishersMethod (Fisher's method [5]), stoufferMethod (Stouffer's method [6]), max (maxP method [7]), or min (minP method [8]) |
This function performs an intra-experiment analysis [1] for individual genes of the given dataset. The function first splits the dataset into smaller datasets, performs a moderated t-test (limma package) for the genes of the split datasets, and then combines the p-values for individual genes using metaMethod
A data frame (rownames are gene IDs) that consists of the following information:
logFC: log foldchange (diseases versus controls)
pLimma: p-value obtained from limma without spliting
pLimma.fdr: FDR-corrected p-values of pLimma
pIntra: p-value obtained from intra-experiment analysis
pIntra.fdr: FDR-corrected p-values of pIntra
Tin Nguyen and Sorin Draghici
[1] T. Nguyen, R. Tagett, M. Donato, C. Mitrea, and S. Draghici. A novel bi-level meta-analysis approach – applied to biological pathway analysis. Bioinformatics, 32(3):409-416, 2016.
bilevelAnalysisGene, intraAnalysisClassic, link{bilevelAnalysisClassic}
data(GSE33223) X <- intraAnalysisGene(data_GSE33223, group_GSE33223) head(X)data(GSE33223) X <- intraAnalysisGene(data_GSE33223, group_GSE33223) head(X)
Load KEGG pathways and names
loadKEGGPathways(organism = "hsa", updateCache = FALSE)loadKEGGPathways(organism = "hsa", updateCache = FALSE)
organism |
organism code. Default value is "hsa" (human) |
updateCache |
re-download KEGG pathways. Default value is FALSE |
A list of the following components
kpg a list of graphNEL objects
encoding the pathway information.
kpn a named vector of pathway tiles. The names of the vector are the pathway KEGG IDs.
Tin Nguyen and Sorin Draghici
keggPathwayGraphs, keggPathwayNames
x <- loadKEGGPathways()x <- loadKEGGPathways()
Calculate p-value for over-representation Analysis
pORACalc(geneSet, DEGenes, measuredGenes, minSize = 0)pORACalc(geneSet, DEGenes, measuredGenes, minSize = 0)
geneSet |
a vector of gene names belong to the geneset |
DEGenes |
a vector of differential expressed genes |
measuredGenes |
a vector of all genes in the analysis |
minSize |
the minimum number of DE genes in the geneSet |
p-value
Combine independent studies using the sum of p-values transformed into standard normal variables
stoufferMethod(x)stoufferMethod(x)
x |
is an array of independent p-values |
Considering a set of m independent significance tests, the resulted p-values are independent and uniformly distributed between 0 and 1 under the null hypothesis. Stouffer's method is similar to Fisher's method (fisherMethod), with the difference is that it uses the sum of p-values transformed into standard normal variables instead of the log product.
combined p-value
Tin Nguyen and Sorin Draghici
[1] S. Stouffer, E. Suchman, L. DeVinney, S. Star, and R. M. Williams. The American Soldier: Adjustment during army life, volume 1. Princeton University Press, Princeton, 1949.
x <- rep(0,10) stoufferMethod(x) x <- runif(10) stoufferMethod(x)x <- rep(0,10) stoufferMethod(x) x <- runif(10) stoufferMethod(x)