3D multicellular models reflect the efficiency of MSC-directed enzyme/prodrug treatment.

Mesenchymal stromal cells (MSC) exhibit beneficial properties to serve as cellular vehicles for enzyme/prodrug cancer gene therapy approaches. We have previously shown that engineered human adipose tissue-derived MSC in combination with non-toxic prodrug mediated substantial cytotoxic and antitumor effect. The aim of this study was to develop advanced 3D cultivation method to serve for modelling of the therapeutic outcome in vitro. We have used engineered MSC expressing fusion transgene cytosine deaminase::uracilphosphoribosyltransferase (CD-MSC) in combination with prodrug 5-fluorocytosine (5FC). This therapeutic regimen designated CD-MSC/5FC was combined with the human melanoma cells A375 or EGFP-A375 in both standard monolayer culture and 3-dimensional (3D) multicellular nodules. The extent of cytotoxicity was evaluated by standard viability assay MTS, ATP-based luminescence assay, fluorimetric test, measurement of Caspase-3/7 activation and DNA laddering. The data have shown that the extent of cytotoxic bystander effect mediated by CD-MSC/5FC is significantly lower in 3D culture conditions. However, these data better recapitulate the therapeutic efficiency as observed previously in vivo. We suggest here to use the 3D multicellular culture conditions for better prediction of the therapeutic outcome in mouse xenograft models.

Long-term efficiency of mesenchymal stromal cell-mediated CD-MSC/5FC therapy in human melanoma xenograft model.

Mesenchymal stromal cells (MSC) can be exploited as cellular delivery vehicles for the enzymes converting non-toxic prodrugs to toxic substances. Because of their inherent chemoresistance, they exert potent bystander and antitumor effect. Here we show that the human adipose tissue-derived MSC expressing fusion yeast cytosine deaminase::uracil phosphoribosyltransferase (CD-MSC) in combination with 5-fluorocytosine (5FC) mediated a long-term tumor-free survival in the 83.3% of tumor-bearing animals. CD-MSC/5FC treatment induced cytotoxicity against model human melanoma cells EGFP-A375. Only 4% of the therapeutic CD-MSC cells eliminated >98.5% of the tumor cells in vitro. Long-term tumor-free survival was confirmed in 15 out of the 18 animals. However, repeatedly used CD-MSC/5FC therapeutic regimen generated more aggressive and metastatic variant of the melanoma cells EGFP-A375/Rel3. These cells derived from the refractory xenotransplants exhibited increased resistance to the CD-MSC/5FC treatment, altered cell adhesion, migration, tumorigenic and metastatic properties. However, long-term curative effect was achieved by the augmentation of the CD-MSC/5FC regimen along with the inhibition of c-Met/hepatocyte growth factor signaling axis in this aggressive melanoma derivative. In summary, the CD-MSC/5FC regimen can be regarded as a very effective antitumor approach to achieve long-term tumor-free survival as demonstrated on a mouse model of aggressive human melanoma xenografts.

Tumor microenvironment and the role of mesenchymal stromal cells.

UNLABELLED: Solid tumors are generally composed of two major components: heterogeneous malignant cells and non-malignant stromal part. The latter comprises several types of non-malignant cells of mesenchymal, endothelial and immune origin and together with the extracellular matrix significantly affects the biological properties of the tumor. This minireview is focused on recent advances in the understanding the role of tumor stromal component and its particular cell types in the tumor behavior. It summarizes the impact of mesenchymal stromal cells and the ways of their potential contribution to the tumor biology. As their role in the tumor development and the effects on the tumor cells remain controversial, we review the recent experimental evidence regarding the crucial molecular factors which determine their role in the tumors. KEYWORDS: tumor microenvironment, human mesenchymal stromal cells, paracrine interaction, malignant cells.