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De novo expression of integrin alpha v beta 3 in Cal27 cells results in pSrc (Y418) inhibition and resistance to antitumor drugs

Introduction: Integrins are cell-surface adhesion molecules that connect cells to components of the extracellular matrix. They play key roles in the regulation of tumor cell adhesion, migration, invasion and survival to antitumor drugs. In the present study we investigate the susceptibility of tongue squamous carcinoma cells Cal27 with de novo expression of integrin αvβ3 to antitumor drugs. Materials and methods: Mock transfected cell clone Cal27-Φ or cell clones Cal27-2B1 and Cal27-2B3 were obtained by stable transfection of Cal27 cells with empty plasmid pcDNA3 or a plasmid containing integrin β3 subunit gene, respectively. The sensitivity of cells to antitumor drugs was determined using MTT assay. Flow cytometry analysis was used to determine the expression of integrins αvβ3 and αvβ5 on the cell surface. Knockdown of integrin subunits αv and β5, Src and ILK was performed by transfection of specific siRNAs. The activity of Src kinase (pSrc(Y418)) was inhibited using dasatinib. The expressions of Src/pSrc(Y418) and ILK were measured by Western blot. Results and discussion: Cell clones Cal27-2B1 and Cal27-2B3 as compared to parental cell line Cal27 and control clone Cal27-Φ demonstrate: (i) de novo expression of integrin αvβ3, (ii) increased expression of integrin αvβ5, (iii) increased adhesion properties to fibronectin and vitronectin ; (iv) resistance to cisplatin, doxorubicin, mitomycin C and 5-fluorouracil ; (v) increased amount of ILK and (vi) decreased amount of Src and pSrc (Y418). ILK knockdown in Cal27-2B1 and Cal27-2B3 had different effects on the cell survival upon treatment with the aforementioned antitumor drugs ruling out its involvement in the resistance to these agents. The Src knockdown in Cal27 cells, as well as concomitant treatment with dasatinib, increased resistance to the four antitumor drugs to the level observed in cell clones Cal27-2B1 and Cal27-2B3. Conclusion: Since dasatinib treatment in Cal27 mimics the effect of a decreased amount of pSrc (Y418) observed in integrin αvβ3-expressing cell clones Cal27-2B1 and Cal27-2B3, we conclude that integrin αvβ3-induced downregulation of pSrc (Y418) is responsible for drug resistance. Given the strong attention that dasatinib receives as a potential drug for cancer therapy in combination with conventional chemotherapy, it is intriguing that decreased Src (Y418) activity confers a resistance to various antitumor drugs.

integrin αvβ3; integrin αvβ5; integrin mediated drug resistance; pSrc (Y418); ILK; migration

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