STAT3/NF-κB-regulated Lentiviral TK/GCV Suicide Gene Therapy for Breast Cancer Treatment

1381 Objectives Triple-negative breast cancer (TNBC), defined as negative on estrogen receptor (ER-), progesterone receptor (PR-) and HER2-, represents approximately 20% of all breast tumors and appears resistance to conventional chemotherapy regimens, demonstrating a particularly poor prognosis and a significantly worse clinical outcome than other tumors. Suicide gene therapy have been used for the in vivo treatment of various solid tumors in recent clinical trials. In tumor microenvironment, STAT3/NF-κB pathways is constitutively activation in stromal cells as well as in cancer stem cells (CSCs) Methods In this study, we have cloned a novel STAT3/NF-κB-based reporter system to drive the expression of herpes simplex virus thymidine kinase (HSV-TK) and the red fluorescence protein (DsRed) against breast cancer. Lentiviral (LV) vector expressing HSV-TK under the regulation of STAT3/NF-κB fused response element was developed. In this setting, we exploited the constitutive STAT3/NFkB activation in tumors as a way to achieve higher transgene expression than a vector in which HSV-TK expression was driven by a constitutively active CMV promotor in vivo. An orthotopic MDA-MB-231 triple negative breast cancer mouse model for evaluating the feasibility of STAT3-NF-κB-TK suicide gene therapy system was also established. Results 3H-FEAU uptake assay was performed to compare the basal promoter activities of LV-CMV-TK and LV-STAT3-NF-κB-TK (Fig. 1A) in MDA-MB-231 cells. The LV-CMV-TK showed ~5 fold higher 3H-FEAU uptake compared to LV-STAT3-NF-κB-TK (Fig. 1B). Immunofluorescence staining with antibodies against HSV1-TK showed the different expression level in cells (Fig. 1C). Western blot analysis indicated the increased level of HSC1-TK in TNF-α treated-LV-STAT3-NF-κB-TK expressing cells (Fig. 1D). In vitro effect of STAT3-NF-κB-induced transgene expression were also observed by TNF-α induction (Fig. 2A) and confirmed by DsRedm fluorescent microscopy (Fig. 2B). In clonogenic assay, cells expressing LV-CMV-TK were 2-fold more sensitive to GCV than LV-STAT3-NF-κB-TK transduced cells (Fig. 3). In vivo therapeutic evaluation in animal models indicated that LV-STAT3-NF-κB-TK showed more tumor growth inhibition than LV-CMV-TK when GCV was administered (Fig. 4). Moreover, the invasion and the expression of CSC markers were both decreased after STAT3/NF-κB-regulated HSV-TK/GCV therapy (Fig 5). Conclusions This study successfully established a theranositic approach to treatment triple negative breast cancer via STAT3-NF-κB responsing element-driven suicide gene therapy. This platform also may be an alternative strategy to handle with cancer stem cells.