Background: Uncontrolled tumor cell division and proliferation cause a rapid increase in cell number and oxygen consumption, leading to a hypoxic condition in the tumor microenvironment (TME) of the most developing tumors (Pathologic Hypoxia). Similarly, the adipose tissue-derived mesenchymal stem cells (hASCs) reside in a healthy hypoxic niche (Physiologic Hypoxia). Studies have shown mesenchymal stem cells (MSCs) are inherently prone to migrate toward solid or developing tumors. Therefore, hypoxia, as a common feature of hASCs niche and TME, and tumor tropism could be used in cancer therapy that would introduce hypoxia-regulated-gene-engineered hASCs as a potential carrier to transfer and release the anti-tumor agents, specifically at the TME. Materials and Methods: We investigated the effect of TME-mimicked hypoxia on a therapeutic transgene expression level in hASCs. For this purpose, a hypoxia-regulated therapeutic gene was constructed which contained the hypoxia-responsive elements (HRE) adjacent to the full cytomegalovirus (CMV) promoter, and cytosine deaminase::uracil phosphoribosyltransferase fusion gene (CD::UPRT), separated with an IRES2 from the enhanced green fluorescent protein gene (EGFP). Following the transfection of this recombinant construct into hASCs via lipofectamine, the cells were incubated in normoxia (21% O2) and hypoxia (2-5% O2) conditions. The expression level of the transgene was evaluated using fluorescence microscopy and flow cytometry. Results: Although the transgene expression in hASCs was transient and had a significant toxicity effect on the cell population, the microscopic evaluation shows the transient expression of transgene (12h post-transfection) was significantly increased under hypoxic conditions while the low expression in normoxia. Conclusion: This study indicates that the HRE results in higher transgene expression under hypoxia in hASCs. This result is optimistic for using engineered hASCs as a carrier for the antitumor therapeutic genes to exert their potential therapeutic effects after migration to the hypoxic TME.

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