Our previous studies have shown that the treatment of human colorectal cancer cells HCT-116 with non-toxic concentrations of antineoplastic drugs such as paclitaxel (PAC) or doxorubicin (DOX) induces changes in the transcription of a wide variety of genes. The treatment was also able to make tumor cells more immunogenic and more sensitive to the activity of cytotoxic T cells generated in vitro. Among the genes, which transcription was increased by pre-treatment, we have found the heat shock proteins genes (HSP40 and HSP70). HSPs are chaperones that direct cross-presentation of antigenic peptides associated with class I molecules and can act as activators of immunocompetent cells, playing an important role in the activation of macrophages and lymphocytes, as well as the activation and maturation of dendritic cells (DCs). Therefore, in the present work we will evaluate whether changes of transcription of HSPs genes, previously identified by DNA microarray, translate into increased protein expression. Confirming such a phenomenon, we will test the hypothesis that the increased immunogenicity of colorectal cancer cells is due to increased expression of HSPs. For this purpose, HCT-116 and HT-29 tumor cells will be treated with minimum effective concentrations (cytostatic concentration) and non-toxic concentrations of 5-fluorouracil (5-FU) or PAC. Expression of HSP40 and 70 will be semi-quantitatively analysed by dot-blot method and qualitatively by immunocytochemical assay. Cells treated with these drugs will be used to sensitize human DCs generated from peripheral blood monocytes of normal donors to assess the effect of expression of HSPs on the DCs functions.
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