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Altered cell-cell adhesion in cisplatin-resistant human carcinoma cells: a link between b-catenin ratio and cisplatin resistance

Inroduction The majority of cancer patients are still treated with classical chemotherapy. In this respect, cisplatin is one of the most effective and commonly used agents for the treatment of a wide spectrum of solid tumors. Nevertheless, resistance to this drug is a major problem that limits the effectiveness of its use in cancer treatment. Greater insight into the molecular mechanisms regarding modulation of the cellular response to cisplatin should help to develop and optimize therapeutic strategies. We have developed cisplatin resistant human laryngeal carcinoma cells, which exhibited altered formation of cell-cell junctions comparing to their parental HEp2 cells. The aim of the present study was to examine molecular basis of this phenomenon. Materials and methods We analyzed and compared the basal and cisplatin-induced expression of cell-cell junctional proteins in parental laryngeal carcinoma HEp2 cells and its cisplatin resistant CA3ST and CK2 sublines:E-cadherin, N-cadherin, beta catenin, plakoglobin, p120 catenin and desmoglein. The expression of proteins was determined by Western blot assay and the expression of mRNA by semiquantitative RT-PCR. Immunocytochemical analysis was performed to estimate the subcellular distribution of selected junctional proteins. Coimmunoprecipitation method was used to examine cellular cadherin-catenin complex-interaction. Cell sensitivity to cisplatin was assayed by MTT spectrophotometric assay. Plasmid tranfection of CA3ST CELLS with plakoglobin gene was done by the standard procedure with lipofectamine transfection reagent. Results: In CA3ST and CK2 cells, N-cadherin, desmoglein and p120-catenin were expressed at the similar level, while none of the cell lines expressed E-cadherin. The expression of plakoglobin in cisplatin resistant cells was down-regulated (on both, mRNA and protein levels), and b-catenin up-regulated (only on protein level). Immunopre- cipitation of cadherin-catenin complex established that upregulation of b-catenin results from its stabilization through interaction with N-cadherin. While increase of b-catenin expression has been found in cisplatin resistant cell lines from different origin but not in vincristine resistant human laryngeal carcinoma cells, we speculate this could be a general phenomena accompanying cisplatin resistance. Transfection of plakoglobin-expressing plasmid vector in CA3ST cells reconstitutes beta-catenin/plakoglobin ratio of parental HEp-2 cells, but does not restore sensitivity to cisplatin. The shift in the composition of cadherin-catenin complexes was not induced by a single treatment with cisplatin. Conclusions It appears that β -catenin and plakoglobin are not involved in the resistance mechanism, implying that the observed alterations are an outcome of slowly generating process, which is presumably a secondary event of vital cellular response triggered by cisplatin toxicity.

cisplatin; drug-resistance; b-catenin; plakoglobin; cell adhesion

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