Visit our Expo - Redox and Inflammation signaling 2012

Session III : Protein kinase cascades as therapeutic targets Poster III, 22
The possible role of PI3-kinase in melanoma cell migration.
Dorota Gil1, Joanna Duli"ska-Litewka1, Anna Pituch Noworolska2 and Piotr Laidler1
1Departament of Medical Biochemistry, Kopernika 7, 31-034 Kraków 2Department of
Clinical Immunology, Polish-American Children's Hospital, Jagielonian University
Medical College, e-mail: dgil@poczta.fm
Integrins are heterodimeric ("/# subunits) cell surface glycoproteins that bind to ECM
proteins and activate members of intracellular signalizing pathways cascades that can
stimulate or regulate motility and invasivnes, cell growth, and survival. Cancer cells very
often switch the types of extracellular matrix receptors (integrins) they express, favouring
ones that transmit progrowth signals. The expression of "3#1 integrin has been indicated as
possibly associated with invasive phenotype not only trough formation of a new cell-matrix
contact sites but also by regulating the expression of matrix-degrading enzymes, MMP-2 or
MMP-9 metalloproteinases. "3#1 integrin is a laminin (LN) receptor and recent studies
showed that laminin–5 and "5–containing laminins, such as laminin–10, and –11, are
preferred ligands for "3#1 integrin. Interaction of "3#1 integrin with LN–5 may provide
evidences to how it transduces signals that affect cell behavior.
METHODS: Studies were carried out on human melanoma metastatic cell lines: A 375 (from
solid tumor) and 1205Lu (from mouse lung). Cells were culture on plastic dishes alone or
covered with LN-5. Expression of integrins was studied by flow cytometry, secretion of
MMP-2 and MMP-9 by zymography, and the invasive potential by using conventional
Boyden chambers. Expression of cell signaling proteins was analyzed using Western Blot.
RESULTS: LN–5 stimulated up regulation of MMPs activity in both cell lines. Signaling
pathway necessary for migration of melanoma cells likely involved PI3-kinase activity as the
addition of the PI3-K specific inhibitor, LY294002, decreased the level of active MMP-2 in
A 375 and LU1205 cells. Western blot analyses of whole cell lysates confirmed the inhibition
of PI3-kinase activity after addition of LY294002, as determined with anti-phospho–AKT
(Ser 473) antibodies and observed increase in the level of phospho–AKT in the cells cultured
on LN–5.
CONCLUSION: Activation of PI3-kinase in response to ECM signals carried through
specific integrin receptor is likely an important event involved in metastasis of melanoma.
Understanding the intracellular signaling pathways in response to ECM proteins may be
useful in the development of tools for selective blocking of some signaling pathways.
This work is suported by the KBN: Jagiellonian University Medical College (W)/256/P/L)
and FP5 EU project “European Searchable Tumor Cell Lines Data Bank” – ESTDAB – NAS:
QLRI – CT-2001-01325.
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