Identification of heat shock protein 90α as an IMH-2 epitope-associated protein and correlation of its mRNA overexpression with colorectal cancer metastasis and poor prognosis

We studied whether HSP90 alpha was associated with the special carbohydrate structures IMH-2 epitopes, and investigated its mRNA expression and clinical relevance in colorectal cancer (CRC) patients. The lysates and the culture media of colon cancer HCT-8 cells were immunoprecipitated with IMH-2 antibody, and the immunoprecipitates were subsequently analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry or by immunoblotting with anti-HSP90 alpha antibody. In vitro wound-healing assay was done to evaluate the role of IMH-2 epitope-associated HSP90 alpha in HCT-8 cell migration. Real-time RT-PCR was performed to detect the levels of HSP90 alpha mRNA expression in paired tumor and non-tumor tissues of 56 CRC patients. The correlation of tumor HSP90 alpha mRNA overexpression with CRC metastasis and poor survival outcome was determined by statistical analyses. HSP90 alpha was first identified as an IMH-2 epitope-associated protein by immunoprecipiation, mass spectrometry, and immunoblotting analysis. IMH-2 epitopes were detected in both cellular and secreted HSP90 alpha. HCT-8 cell migration induced by serum starvation-conditioned medium was blocked by anti-HSP90 alpha antibody or the HSP90 alpha inhibitor geldanamycin (GA) as efficient as by IMH-2 antibody, suggesting that IMH-2-associated HSP90 alpha was involved in serum starvation-induced CRC cell migration. On the other hand, HSP90 alpha mRNA expression was induced in HCT-8 cells after serum starvation. Clinically, 15 (26.8%) of 56 CRC patients exhibited tumor HSP90 alpha mRNA overexpression and had higher rates of metastatic occurrence (P = 0.003) and poor prognosis (P = 0.028). HSP90 alpha was an IMH-2 epitope-associated protein. Tumor HSP90 alpha overexpression was correlated with the metastasis and poor prognosis of CRC patients.