Integrating cancer genomics and proteomics in the post-genome era

Abstract

The dawn of the post-genome era is leading to extraordinary opportunities in biomedicine. Our group has embarked on a major effort to integrate genomics, transcriptomics and proteomics for the profiling of tumor tissues, an approach we refer to as operomics. Our major goals are the molecular classification of tumors and the identification of markers for the early detection of cancer. Molecular analyses of tumors rely on microdissected tissues, which are simultaneously investigated for genomic, transcriptomic and proteomic changes. Genomic alterations in tumor cells being investigated include deletions, amplifications and methylation changes across the entire genome as well as point mutations in specific genes. Expression analysis at the RNA level is being undertaken using oligonucleotide and cDNA based microarrays. An important aspect of our approach is the large-scale identification and quantitative analysis of tumor proteins in whole cell lysates as well as in protein compartments. Protein separation strategies include two-dimensional polyacrylamide gel electrophoresis and liquid chromatography. Specific protein subsets, of interest include membrane proteins, secreted proteins and antigenic proteins as sources of biomarkers for early detection of cancer. Our current approach is illustrated with findings stemming from our studies of human gliomas.

Proteins and Protein Pattern Differences between Glioma Cell Lines and Glioblastoma Multiforme

“…Our study illustrates that over 60 proteins are lost during cell culture and over 160 proteins are gained, out of the f500 protein spots reliably identified on 2DGE over the pH range 4 to 7. It is likely that many more proteins are differentially expressed when an even larger fraction of the proteome is examined (20,21). These proteins may have many different influences on GBM growth, resistance to current therapies, and brain invasion and migration.…”

“…One advantage of the 2DGE method, compared with other protein-based techniques such as matrix-assisted laser desorption/ionization-MS or surface-enhanced laser desorption/ionization-MS, is that the protein can be spotted, excised from the gel, and sequenced directly (20,21). Whereas matrix-assisted laser desorption/ionization or surface-enhanced laser desorption/ ionization are more tractable for running large-scale experiments, selective tissue microdissection and 2DGE requires less in the way of instrumentation, cost, repairs, or personnel.…”

“…Several recent studies have highlighted the broad utility of microarrays in identifying transcriptional networks or in describing tumor subtypes, both in gliomas, in other tumors, and in normal cells such as fibroblasts in culture (30,31,(34)(35)(36)(37)(38)(39)(40)(41)(42)(43)(44)(45)(46). One disadvantage of microarrays is that they require significant investments in time and equipment, can be challenging technically, and require significant statistical expertise (20,34,44). Like our method, microarrays may require downstream confirmation and functional validation (20,34).…”

“…One disadvantage of microarrays is that they require significant investments in time and equipment, can be challenging technically, and require significant statistical expertise (20,34,44). Like our method, microarrays may require downstream confirmation and functional validation (20,34). It is also important to emphasize that with 2DGE methods such as this one, low-abundance proteins may be difficult to identify and special methods may be needed to isolate membrane-bound proteins or proteins that are very basic or acidic (the pH range in this study was 4-7, which is slightly basic; ref.…”

“…It is also important to emphasize that with 2DGE methods such as this one, low-abundance proteins may be difficult to identify and special methods may be needed to isolate membrane-bound proteins or proteins that are very basic or acidic (the pH range in this study was 4-7, which is slightly basic; ref. 20).…”