Comparisons of gene expression signatures provide a way to explore functional connections among biological events in global aspects of cell response. GeneExpressionSignature is an R package developed for the large-scale analysis of gene expression signatures. The package implements two rank-merging algorithms and two similarity-scoring algorithms. The functions of GeneExpressionSignature provide a flexible solution for gene expression signature-based studies and hold great potential in biomedical research applications, such as drug repurposing. GeneExpressionSignature is released under GPL v2 within the Bioconductor project and is freely available at http://www.bioconductor.org/packages/release/bioc/html/GeneExpressionSignature.html .