BACKGROUND: An integral component of cancer biology is the understanding of molecular properties uniquely distinguishing one cancer type from another. One class of such properties is histone post-translational modifications (PTMs). Many histone PTMs are linked to the same diverse nuclear functions implicated in cancer development, including transcriptional activation and epigenetic regulation, which are often indirectly assayed with standard genomic technologies. Thus, there is a need for a comprehensive and quantitative profiling of cancer lines focused on their chromatin modification states. RESULTS: To complement genomic expression profiles of cancer lines, we report the proteomic classification of 24 different lines, the majority of which are cancer cells, by quantifying the abundances of a large panel of single and combinatorial histone H3 and H4 PTMs, and histone variants. Concurrent to the proteomic analysis, we performed transcriptomic analysis on histone modifying enzyme abundances as a proxy for quantifying their activity levels. While the transcriptomic and proteomic results were generally consistent in terms of predicting histone PTM abundance from enzyme abundances, several PTMs were regulated independently of the modifying enzyme expression. In addition, combinatorial PTMs containing H3K27 methylation were especially enriched in breast cell lines. Knockdown of the predominant H3K27 methyltransferase, enhancer of zeste 2 (EZH2), in a mouse mammary xenograft model significantly reduced tumor burden in these animals and demonstrated the predictive utility of proteomic techniques. CONCLUSIONS: Our proteomic and genomic characterizations of the histone modification states provide a resource for future investigations of the epigenetic and non-epigenetic determinants for classifying and analyzing cancer cells.