Study On Introduction Of Oncolytic Adenovirus Into Acute Myeloid Leukeima Stem Cell Through CD123

BACKGROUND: Leukemia is a heterogeneous disease at the molecular level resulting from a variety of alterations in numerous genes important for cell growth, differentiation, and cell death, is one of malignancies of hematopoietic systems. Chemotherapy and transplantation are the primary therapeutic means of leukemia, but most of patients could only obtain short-term remission and eventually died of relapse or drug resistance. Studies have shown that, there is a group of leukemia stem cells with very low percentage in leukemia patients, which plays an important role in the development and progression of leukemia, and is considered as the major source of leukemia relapse and drug resistance. It is expected to completely cure leukemia through the strategy targeting leukemia stem cell.DESIGN & OBJECTIVE: CD123, which is widely reported to be overexpressed on AML blasts, CD34+ leukemic progenitors, and AML-LSCs in comparison with normal HSCs, seems to be an excellent target for the therapy of leukemia. In this study, the oncolytic adenovirus Ad.sp-E1A-sCAR-IL-3 was constructed, in which E1A was driven by survivin promoter, and a bifunction-fused gene sCAR-IL-3 was introduced into the adenovirus. The adenovirus could not only specificly target to leukemia stem cell, but also solve the problem that the traditional adenovirus type 5 is difficult to infect LSC. This virus has two advantages: (1) With the characteristics of high-specific expression in leukemia stem cells of survivin, utilization of survivin promoter to control the E1A gene (necessary for viral replication and proliferation) can make adenovirus replicate in the survivin-positive leukemia stem cells and not in survivin-negative nomal cells. (2) The expression of bifuction-fused gene sCAR-IL-3 could provide a bridge for adenovirus knob and CD123 on leukemia stem cell surface, which could mediate adenovirus into leukemia stem cells effectively.METHODS: (1) Construction of the shuttle vector for adenovirus package; (2) The oncolytic adenovirus were packaged in HEK293 cells by using homologous recombinant method; (3) The tumor-killing effects and the safety evaluation were proved through the cell viability assay; (4) The tumor cell apoptosis was analyzed through morphological analysis, apoptosis staining assay, TEM.RESULTS: The targeting AML-LSC oncolytic adenovirus Ad.sp-E1A-sCAR-IL-3 was successfully constructed; the results showed that oncolytic adenovirus exhibited potent anti-leukemia stem cell activity in vitro.CONCLUSIONS & SIGNIFICANCE: In the study, oncolytic adenovirus was firstly introduced into the therapy of acute myeloid leukemia through targeting CD123. Our results suggest that a targeted oncolytic adenovirus might be a valuable agent for acute myeloid leukemia patients.