Abstract 4213: Gene expression of hsp27 in renal cell carcinoma and the correlation with tumor progression
Authors: Kokkat Theresa J. , Khoja Hamid , Lovecchio Lorraine , Krowczynska Anna , Patel Miral , Livolsi Virginia A. , McGarvey Diane 
Published: August 2012

Abstract
Background Overexpression of Hsp27 protein has been correlated with tumor progression in a variety of cancers including Renal Cell Carcinoma (RCC). Various phosphorylation patterns of Hsp27 correlate with the aggressiveness of tumor phenotype. Increased Hsp27 phosphorylation was reported to correlate with increased tumor progression in RCC. Since the phosphorylation status of Hsp27 is dependent on phosphorylation by kinases and dephosphorylation by phosphatases, a better understanding of these factors in the tumor microenvironment needed to be analyzed at the gene expression level. Gene expression studies of tumor tissues using microarray routinely fail to identify hsp27 because large amount of transcripts that microarrays analyze may mask the changes observed with hsp27 and its associated genes. Hence, we used quantitative RT-PCR to investigate the relative gene expressions of hsp27 in RCC tumor and normal adjacent tissues (NAT). We examined the gene expression levels of MAP kinases such as Mitogen Activated Protein Kinase 1 (mapk1) and MAPK-activated protein Kinase 5 (mk5) along with a major phosphatase, Protein Phosphatase 2 A (pp2a). Methods RCC tumor and normal control samples were frozen in liquid nitrogen within one hour post surgical excision. Frozen samples were pulverized in Covaris TissueTubes using the Covaris CryoPrep™ system. The pulverized tissue was homogenized for RNA extraction using the Covaris Adaptive Focused Acoustics (AFA) S220 system. Total RNA was extracted using a Qiagen RNeasy Plus kit. The gene expressions for Hsp27, MAPK1, MK5, and PP2A were analyzed by RT-PCR using validated TaqMan assays on an Applied Biosystems 7500 Fast Real-Time PCR System. A house keeping gene, beta actin, was measured in parallel to normalize the differences between samples. Results In RCC, in comparison to NAT, the relative gene expressions of hsp27 increased to 224.44% suggesting an upregulation of hsp27 in cancer tissue. Adding to this microenvironment, the phosphorylating enzymes were also upregulated in the RCC tissue. Expression levels of kinases, mapk1 and mk5 were increased to 22.11% and 37.18% respectively. Interestingly, the level of phosphatase (pp2a) was decreased 67.25 %. These data suggests that the combination of the upregulation of kinases along with down regulation of phosphatase may be a major factor creating the environment of enhanced phosphorylation in overexpressed hsp27. Conclusion Our data shows that the gene expression level increase of hsp27 in RCC is consistent with the predicted increase in protein level expression of Hsp27. However, the dramatic combined effect of the increase in the gene expression of the kinases, along with decrease in the gene expression of the phosphatase may result in a striking increase in phosphorylation. The enhanced phosphorylation will activate Hsp27 in the tumor microenvironment resulting increased tumor progression.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4213. doi:1538-7445.AM2012-4213