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Effect of polyamine depletion on cancer stem cells

Background

Although breast cancer cell lines are more homogenous in cellular architecture than are breast tumours, different sub-populations based on the expression of CD44, CD24 and other markers of breast cancer stem cells (CSC) can be found. However, when cancer cell lines are grown as multicellular spheroids (MCSs), it appears that the proportion of CSC increases. Also, growth of cells in a three-dimensional MCS more resembles tumour growth in the body than growth of cells as adherent two-dimensional monolayers. Since a number of compounds that deplete polyamines are used in various clinical trials, it is of importance to investigate how the sub-population of CSC is affected.

Experimental system

We are using a number of cancer cell lines of breast cancer origin (MCF-7, SK-BR-3, JIMT1, MDA-MB-231, HCC1937 and L56Br-C1) grown as MCSs. We also grow the normal MCF-10A epithelial breast cancer cell line as MCSs. The MCS are grown in conditions of normoxia and hypoxia and treated with various compounds that lower the cellular polyamine pools.

Effects on various aspects of CSC by polyamine analogue treatment

Polyamine analogues are recognized by the cell as polyamines and are efficiently taken up by the polyamine transport system. They accumulate to high levels in the cell and this results in the down-regulation of polyamine biosynthesis. As too high polyamine levels are detrimental to the cell, polyamine catabolism is also activated. However, the polyamine catabolic pathway only catabolizes the natural polyamines while the polyamine analogues are spared. Thus, the cell will be depleted of the natural polyamines while accumulating the analogue. The analogues cannot take over the biological function of the polyamines and thus cell proliferation ceases and apoptosis may be induced. We are using immunofluorescence microscopy and flow cytometry to investigate how different cellular sub-populations in MCS are affected by polyamine analogue treatment. We are studying effects on cell cycle kinetics and apoptosis in the different sub-populations.

Multicellular spheroids of MCF-10A normal breast epithelial cells grown in normoxia or hypoxia,
Multicellular spheroids of MCF-10A normal breast epithelial cells grown in normoxia, A, or hypoxia, B. Green fluorescence: CD44. Red fluorescence: CD24. Blue fluorescence: bisbenzimide-stained nuclei. The bar represents 10 micrometer.
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