Intracellular targets of RGDS peptide in melanoma cells

In the present study we show for the first time that RGDS-peptide is internalized in melanoma cells in a time-dependent way and exerts strong anti-proliferative and pro-apoptotic effects independently from its extracellular anti-adhesive action. RGES control-peptide did not show biological effects, as expected; nevertheless it is internalized, although with slower kinetics. Survivin, a known cell-cycle and survival-regulator is highly expressed in melanoma cells. Co-immunoprecipitation assays in cell lysates and overlay assays with the purified proteins showed that RGDS interacts with survivin, as well as with procaspase-3, -8 and -9. RGDS-peptide binding to survivin was found to be specific, at high affinity (Kd 27.5 μM) and located at the survivin C-terminus. RGDS-survivin interaction appeared to play a key role, since RGDS lost its anti-mitogenic effect in survivin-deprived cells with a specific siRNA.RGDS inhibits melanoma growth with an adhesion-independent mechanism; it is internalized in melanoma cells and specifically interacts with survivin. The present data may indicate a novel role of RGDS-containing peptides physiologically released from the extracellular matrix and may suggest a possible novel anti-proliferation strategy in melanoma.RGD (Arg-Gly-Asp) motif is largely investigated as mediator of cell adhesion to extracellular matrix and to cells, via cell-surface receptors named integrins. These receptors belong to a large family of twenty-four heterodimeric members. Several integrins, including αvβ3, α5β1, αvβ5, αvβ6 and αIIbβ3, recognize the RGD motif present in various ECM proteins such as fibronectin, vitronectin, laminin, fibrinogen, von Willebrand factor, osteopontin, thrombospondin, and collagen [1] as well as in disintegrins; others including α1β1, α2β1, α10β1, and α11β1, interact with the matrix in a RGD-independent manner [2-4]. Integrins activation triggers different signals regulating cell adhesion, migration, survival, apoptosis [1,5,6] as w