ADENOASSOCIATED VIRUS VECTOR GENE-EXPRESSION OCCURS IN NONDIVIDING CELLS IN THE ABSENCE OF VECTOR DNA INTEGRATION
ADENOASSOCIATED VIRUS VECTOR GENE-EXPRESSION OCCURS IN NONDIVIDING CELLS IN THE ABSENCE OF VECTOR DNA INTEGRATION
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DOI：
10.1165/ajrcmb.11.5.7946381
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发表时间：
1994-11-01
影响因子：
6.4
通讯作者：
ZEITLIN, PL
中科院分区：
文献类型：
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作者：
FLOTTE, TR;AFIONE, SA;ZEITLIN, PL
基于腺相关病毒2型（AAV2）的载体能够在气道上皮中稳定表达，可能对人类疾病（如囊性纤维化）的基因治疗有用。某些病毒载体，如逆转录病毒，需要活跃的细胞分裂来整合和表达，但这在AAV2的情况下尚未得到正式评估。囊性纤维化支气管上皮细胞系IB3-1可以被AAV2载体转导，当生长到合流时，DNA合成下降到不可检测的水平。在小于0.1%细胞分裂的培养中，AAV-lacZ或AAV-neo载体仍然可以有效地转导，具有线性剂量反应，高达91%，每个细胞有3000个载体颗粒的多重性。载体DNA在非分裂靶细胞中的命运通过低分子量非整合DNA和高分子量基因组DNA片段的Southern印迹法进行研究。载体DNA的可检测水平仅在非整合状态下可见。这些结果表明，与逆转录病毒载体不同，基于aav2的载体不需要活跃的细胞分裂或整合来进行表达，因此具有独特的生物学特性。
Adeno-associated virus type 2 (AAV2)-based vectors are capable of stable expression in the airway epithelium and may be useful for gene therapy for human diseases, such as cystic fibrosis. Certain virus vectors, such as retroviruses, require active cell division for integration and expression, but this has not been formally evaluated in the case of AAV2. The cystic fibrosis bronchial epithelial cell line, IB3-1, which can be transduced by AAV2 vectors, was shown to undergo a decrease in DNA synthesis to undetectable levels when grown to confluence. Cultures in which < 0.1% of cells were dividing could still be efficiently transduced with AAV-lacZ or AAV-neo vectors, with a linear dose response, up to 91% with a multiplicity of 3,000 vector particles per cell. The fate of vector DNA in nondividing target cells was investigated by Southern blotting of both low molecular weight, nonintegrated DNA and high molecular weight, genomic DNA fractions. Detectable levels of vector DNA were only seen in the nonintegrated state. These results indicate that AAV2-based vectors, unlike retrovirus vectors, do not require active cell division or integration for expression to occur and thus possess a unique profile of biologic properties.