ARE/SUZ12 dual specifically-regulated adenoviral TK/GCV system for CML blast crisis cells

Abstract

Background: Treatment of blast phase chronic myeloid leukemia (BP-CML) remains a challenge, and the median survival is less than 6 months. Because effective treatments are lacking, we studied tight targeting of blast crisis CML cells using adenoviral (Ad) vectors expressing a HSV-TK system under dual control of a specific SUZ12 promoter and an antioxidant response element (ARE). Methods: A potential SUZ12 promoter fragment was designed with bioinformatics databases and identified with a luciferase assay. Next, we cloned the ARE element of the NQO1 gene and developed Ad vectors expressing TK kinase or luciferase under the dual control of a specific SUZ12 promoter and an ARE element. An in vitro transfection assay with Ad-ARE/SUZ12-Luc was used to determine promoter activity of ARE/SUZ12 regulatory element in blast crisis CML cells. After incubating human BP-CML-derived cells with Ad-ARE/SUZ12-TK and ganciclovir, Western blot, CCK8, Immunofluorescent assays and Annexin V assays were conducted to assess the efficacy of an ARE/SUZ12 dual-specific TK/GCV system for BP-CML cell lines. Results: Here, luciferase data confirmed significantly higher and specificer promoter activity of the ARE/SUZ12 composite component in CML blast crisis-derived cell lines (K562, KCL22, and K562/G01) compared to HepG2 cells, and Ad-AS-TK/GCV system could exhibit enhanced apoptotic effects and decreased cell viability for BP-CML cell lines. Additionally, Ad-AS-TK/GCV system altered expression of cycle-related and apoptosis-related proteins in BP-CML cell lines. Conclusions: Thus, ARE/SUZ12 dual targeting TK/GCV system was effective in killing BP-CML cells. Moreover, efficacy and specificity of CML cell eradication were enhanced by synergistic effects of ARE/SUZ12 dual-specific regulation. We conclude that suicide gene-targeted therapy might hold promise for BP-CML treatment.