Persistent expression of human clotting factor IX from mouse liver after intravenous injection of adeno-associated virus vectors

We previously found that gene transduction by adeno-associated virus (AAV) vectors in cell culture can be stimulated over 100-fold by treatment of the target cells with agents that affect DNA metabolism, such as irradiation or topoisomerase inhibitors. Here we show that previous γ-irradiation increased the transduction rate in mouse liver by up to 900-fold, and the topoisomerase inhibitor etoposide increased transduction by about 20-fold. Similar rates of hepatic transduction were obtained by direct injection of the liver or by systemic delivery via tail vein injection. Hepatocytes were much more efficiently transduced than other cells after systemic delivery, and up to 3% of all hepatocytes could be transduced after one vector injection. The presence of wild-type AAV, which contaminates many AAV vector preparations, was required to observe a full response to γ-irradiation. Injection of mice with AAV vectors encoding human clotting factor IX after γ-irradiation resulted in synthesis of low levels of human clotting factor IX for the 5-month period of observation. These studies show the potential of targeted gene transduction of the liver by AAV vectors for treatment of various hematological or metabolic diseases.

我们此前已发现，在细胞培养体系中，通过辐射或拓扑异构酶抑制剂等影响DNA代谢的试剂处理靶细胞，可将腺相关病毒（adeno-associated virus, AAV）载体介导的基因转导效率提升100倍以上。本研究证实，预先进行γ射线辐照可使小鼠肝脏的基因转导效率提升最高达900倍，而拓扑异构酶抑制剂依托泊苷可使转导效率提升约20倍。无论是直接肝脏注射还是通过尾静脉注射进行全身性递送，均可获得相当水平的肝脏基因转导效率。全身性递送后，肝细胞相较于其他细胞的转导效率显著更高，单次载体注射后即可使最多3%的肝细胞完成转导。许多AAV载体制剂中污染的野生型腺相关病毒，是观测到γ射线辐照产生完整应答反应的必要条件。在γ射线辐照后向小鼠注射编码人凝血因子IX的AAV载体，可在为期5个月的观测周期内检测到低水平的人凝血因子IX合成。本研究证实了AAV载体靶向转导肝脏在治疗各类血液系统或代谢性疾病方面的应用潜力。