HSP90 as a marker of progression in melanoma.

BACKGROUND: HSP90 chaperones molecules critical for cell survival and malignant progression, including mutated B-raf. HSP90-targeting agents are in clinical trials. No large studies have been conducted on expression of HSP90 in melanomas. MATERIALS AND METHODS: Tissue microarrays containing 414 nevi, 198 primary and 270 metastatic melanomas were assessed using our automated quantitative analysis (AQUA) method of in situ protein measurement; we use S-100 to define pixels as melanocytes (tumor mask) within the array spot, and measure HSP90 expression within the mask using Cy5-conjugated antibodies. RESULTS: HSP90 expression was higher in melanomas than nevi (P < 0.0001) and higher in metastatic than primary specimens (P < 0.0001). No association was seen between high HSP90 expression and survival in the primary or metastatic patient subsets. In primary melanomas, high HSP90 expression was associated with higher Clark level (P = 0.0167) and increased Breslow depth (P < 0.0001). CONCLUSIONS: HSP90 expression was significantly higher in tumors than nevi and was associated with disease progression, indicating that it might be a valuable drug target in melanoma, as well as a useful diagnostic marker. Prospective studies are needed to confirm the diagnostic role of HSP90, as well as the predictive role of HSP90 expression in patients treated with HSP90 inhibitors.

The characterization of PPAR alpha ligand drug action in an in vivo model by comprehensive differential gene expression profiling.

Expression pharmacogenomics includes differential gene expression (DGE) profiling of drug responses in model systems to generate a set of differentially modulated drug-responsive genes which can serve as a surrogate measure for drug action. In this manner, expression pharmacogenomics bridges the fields of genomics and medicinal chemistry. Additionally, modulated genes can be organized into metabolic and signaling pathways that highlight the mechanism of drug activity in a selected tissue. Here, we describe the application of expression pharmacogenomics to characterize a drug response in the clinically relevant in vivo model, the Sprague-Dawley rat. Following oral dosing of rats with GW9578, a novel synthetic peroxisome proliferator activated receptor alpha (PPAR alpha) ligand indicated for lipid disorders, we applied GeneCalling, a differential mRNA transcript profiling technique, to rat liver cDNA. Following GW9578 treatment, 2.4% of the rat liver genes were differentially expressed. We confirmed the sequence identity of 50 distinctly modulated genes. DGE was observed among genes representative of at least six discrete metabolic pathways. Furthermore, we observed up-regulation of 20 genes involved in mitochondrial, peroxisomal and microsomal fatty acid oxidation, consistent with molecular biological and clinical data indicating PPAR alpha ligand principal efficacy to be through increasing fatty acid metabolism. Those pathways regulated in our study that are potentially contributory to target effect, non-target adverse effects, or of unknown consequence include xenobiotic detoxification and steroid modification. Finally, comprehensive drug response profiling can lead to the serendipitous discovery of novel disease indications. In this case, these results suggest a potential novel indication for GW9578 in the treatment of X-linked adrenoleukodystrophy. We have shown, therefore, that the organization of DGE results into metabolic and signaling pathways can elucidate mechanisms of pharmacologically desired (i.e., efficacious) and, where appropriate, undesired (i.e., potentially deleterious) effects.

Identification and comparative analysis of human colonocyte short-chain fatty acid response genes.

Short-chain fatty acids (SCFAs) butyrate, propionate, and acetate produced during fiber fermentation promote colonic differentiation and can reverse or suppress neoplastic progression. We sought to identify candidate genes responsible for SCFA activity on colonocytes and to compare the relative activities of independent SCFAs. cDNA was generated from polyA+ mRNA isolated from control Caco-2 cells and cells treated with equimolar butyrate, propionate, and acetate. GeneCalling, a restriction-based differential RNA expression platform linked to a DNA sequence database lookup, was applied. A total of 30,000 individual genetic sequences were analyzed for differential expression among the three SCFAs. Differentially expressed peaks corresponding to cancer-related genes were isolated, sequenced, and cross-referenced to the GenBank human database. Gene identities were independently confirmed by oligonucleotide poisoning. More than 1000 gene fragments were identified as being substantially modulated in expression by butyrate. Butyrate tended to have the most pronounced effects and acetate the least. Five fragments selected for further study were fully sequenced and proved 100% homologous with human sequences for clusterin, amyloid precursor-like protein 2, and caudal homeobox 2 protein, not previously known to be modulated by SCFAs. In each case, a similar order of potency for the three SCFAs studied was observed. The common SCFAs appear to exert different effects. This study suggests the diversity of the SCFA response at the molecular level and facilitates identifying genes important in the biologic activity of dietary fiber.