Worldwide, 890 000 new cases and approximately 450 000 deaths are caused by head and neck cancer annually, making this the seventh most common cancer type worldwide. To reduce side-effects and to save healthy tissue, it is important to establish more targeted and specific therapeutic strategies.
We aim to develop a dental pulp stem cell (DPSC)-based suicide gene therapy. Hence, we overexpress the herpes simplex virus type 1 - thymidine kinase (HSV1-TK) in a polycistronic construct also including firefly luciferase (Fluc) and a Flag tag using lentiviral vectors in DPSC. HSV1-TK is used in combination with its non-toxic prodrug ganciclovir (GCV). Upon phosphorylation by HSV1-TK, GCV turns into its toxic metabolite and induces cell death.
Because DPSC are isolated from third molars from different donors, we aim to assess the expression of HSV1-TK and Fluc in different DPSC lines (N=3) following lentiviral transduction. Therefore, we performed polymerase chain reaction (PCR), immunocytochemistry (ICC) and western blot (WB) for the HSV1-TK gene and Flag tag. Additionally, bioluminescence imaging is executed for Fluc visualisation. By this, we can compare the different expression levels of the transgenes in different DPSC lines.
Results show that all transduced DPSC of different donors are expressing the HSV1-TK gene significantly on DNA as well as protein levels as confirmed using western blot and PCR. This was not observed in the control cells. Moreover, bioluminescence imaging confirms the expression of Fluc in all transduced DPSC lines, but not in controls.
In conclusion, we successfully induced the expression of HSV1-TK and Fluc in different DPSC lines (N=3). This allows us the explore their use in future experiments using stem cell-based suicide gene therapy.