Focal adhesion alterations in G0-positive melanoma cells

Abstract

Background: Melanoma is a highly heterogeneous malignant tumor that exhibits various forms of drug resistance. Recently, reversal transition of cancer cells to the G₀ phase of the cell cycle under the influence of therapeutic drugs has been identified as an event associated with tumor dissemination. In the present study, we investigated the ability of chemotherapeutic agent dacarbazine to induce a transition of melanoma cells to the G₀ phase as a mechanism of chemoresistance.

Methods: We used the flow cytometry to analyze cell distribution within cell cycle phases after dacarbazine treatment as well as to identifyG₀ -positive cells population. Transcriptome profiling was provided to determine genes associated with dacarbazine resistance. We evaluated the activity of β-galactosidase in cells treated with dacarbazine by substrate hydrolysis. Cell adhesion strength was measured by centrifugal assay application with subsequent staining of adhesive cells with Ki-67 monoclonal antibodies. Ability of melanoma cells to metabolize dacarbazine was determined by expressional analysis of CYP1A1, CYP1A2, CYP2E1 followed by CYP1A1 protein level evaluation by the ELISA method.

Results: The present study determined that dacarbazine treatment of melanoma cells could induce an increase in the percentage of cells in G₀ phase without alterations of β-galactosidase positive cells which corresponded to the fraction of the senescent cells. Transcriptomic profiling of cells under dacarbazine induction of G₀ -positive cells percentage revealed that 'VEGFA-VEGFR2 signaling pathway' and 'Cell cycle' signaling were mostly enriched by dysregulated genes. 'Focal adhesion' signaling was also found to be triggered by dacarbazine. In melanoma cells treated with dacarbazine, an increase in G₀ -positive cells among adherent cells was found.

Conclusions: Dacarbazine induces the alteration in a percentage of melanoma cells residing in G₀ phase of a cell cycle. The altered adhesive phenotype of cancer cells under transition in the G₀ phase may refer to a specific intercellular communication pattern of quiescent/senescent cancer cells.

Keywords: G0; cell cycle; focal adhesion; melanoma; quiescence; senescence.