Proteasenexpression einzelner disseminierter Tumorzellen aus Lymphknoten und Knochenmark von Patienten mit nichtkleinzelligem Bronchialkarzinom
Beschreibung
vor 19 Jahren
Non-small cell lung cancer (NSCLC) is characterised by early
dissemination of tumor cells resulting in a strikingly reduced
overall survival despite complete surgical removal of the primary
tumour. It has been suggested that proteases contribute to this
apparent aggressiveness of lung cancer because they are involved in
a variety of mechanisms associated with tumor progression such as
invasion, migration and angiogenesis. To investigate the
significance of protease expression and to identify potentially
co-regulated molecules during early dissemination and in minimal
residual disease, we performed cDNA array analysis of single
disseminated cancer cells or small cell clusters isolated from bone
marrow and lymph nodes of NSCLC patients. We obtained
macroscopically tumor free lymph nodes and bone marrow aspirates
from patients with operable NSCLC and enriched single disseminated
cancer cells and small cell clusters by density gradient
centrifugation. Subsequently, the freshly prepared cell suspensions
were stained with an antibody against the epithelial surface
molecule EpCAM and single positive tumour cells were isolated by
micromanipulation. After global amplification of the single cell
cDNA and non-radioactive labelling proteinase expression was
assessed using a cDNA array consisting of several matrix
metalloproteases, cathepsins, caspases, kallikreins and other
serine / cysteine proteases. Until now we could isolate 46 EpCAM+
cells from 72 lymph nodes and 19 EpCAM+ cells from 71 bone marrow
aspirates. For 30 cells the epithelial origin could be confirmed by
the co-expression of several epithelial markers or by expression of
the tumor specific MAGE antigens. Hybridisation of these cells on
our protease cDNA array revealed the expression of various
proteases in single cells and small cell clusters. Particularly,
serine proteases, cathepsins and other cysteine proteinases are
frequently expressed, while, contrary to our expectations, the
transcripts of matrix metalloproteases (MMP) were rarely detected
in the analysed cells.
dissemination of tumor cells resulting in a strikingly reduced
overall survival despite complete surgical removal of the primary
tumour. It has been suggested that proteases contribute to this
apparent aggressiveness of lung cancer because they are involved in
a variety of mechanisms associated with tumor progression such as
invasion, migration and angiogenesis. To investigate the
significance of protease expression and to identify potentially
co-regulated molecules during early dissemination and in minimal
residual disease, we performed cDNA array analysis of single
disseminated cancer cells or small cell clusters isolated from bone
marrow and lymph nodes of NSCLC patients. We obtained
macroscopically tumor free lymph nodes and bone marrow aspirates
from patients with operable NSCLC and enriched single disseminated
cancer cells and small cell clusters by density gradient
centrifugation. Subsequently, the freshly prepared cell suspensions
were stained with an antibody against the epithelial surface
molecule EpCAM and single positive tumour cells were isolated by
micromanipulation. After global amplification of the single cell
cDNA and non-radioactive labelling proteinase expression was
assessed using a cDNA array consisting of several matrix
metalloproteases, cathepsins, caspases, kallikreins and other
serine / cysteine proteases. Until now we could isolate 46 EpCAM+
cells from 72 lymph nodes and 19 EpCAM+ cells from 71 bone marrow
aspirates. For 30 cells the epithelial origin could be confirmed by
the co-expression of several epithelial markers or by expression of
the tumor specific MAGE antigens. Hybridisation of these cells on
our protease cDNA array revealed the expression of various
proteases in single cells and small cell clusters. Particularly,
serine proteases, cathepsins and other cysteine proteinases are
frequently expressed, while, contrary to our expectations, the
transcripts of matrix metalloproteases (MMP) were rarely detected
in the analysed cells.
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