STABLE IN-VIVO EXPRESSION OF THE CYSTIC-FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR WITH AN ADENOASSOCIATED VIRUS VECTOR
STABLE IN-VIVO EXPRESSION OF THE CYSTIC-FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR WITH AN ADENOASSOCIATED VIRUS VECTOR
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DOI：
10.1073/pnas.90.22.10613
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发表时间：
1993-11-15
影响因子：
11.1
通讯作者：
CARTER, BJ
中科院分区：
文献类型：
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作者：
FLOTTE, TR;AFIONE, SA;CARTER, BJ
腺相关病毒（AAV）载体表达正常的囊性纤维化跨膜传导调节因子（CFTR）的cDNA补充囊性纤维化（CF）的缺陷在体外。与其他DNA病毒载体不同，AAV是一种稳定整合的病毒，它可以在体内长期补充气道上皮中的CF缺陷。我们报告了AAV-CFTR基因转移和表达后感染的原CF鼻息肉细胞和AAV-CFTR载体在体内交付后，通过纤维支气管镜兔肺的一个肺叶。在家兔中，给药后6个月内，可在感染的肺叶中检测到载体DNA。使用重组AAV-CFTR蛋白特有的26个氨基酸的多肽序列来产生寡核苷酸探针和多克隆抗体，其允许明确鉴定载体RNA和CFTR蛋白表达。使用这些试剂，在载体给药后长达6个月的感染肺叶的气道上皮中检测到CFTR RNA和蛋白质。因此，AAV载体确实有效地促进体内基因转移至气道上皮，其在数月内是稳定的。这些发现表明，AAV-CFTR载体可能是非常有用的基因治疗。
Adeno-associated virus (AAV) vectors expressing the normal cystic fibrosis transmembrane conductance regulator (CFTR) cDNA complement the cystic fibrosis (CF) defect in vitro. Unlike other DNA virus vectors, AAV is a stably integrating virus, which could make possible long-term in vivo complementation of the CF defect in the airway epithelium. We report AAV-CFTR gene transfer and expression after infection of primary CF nasal polyp cells and after in vivo delivery of AAV-CFTR vector to one lobe of the rabbit lung through a fiberoptic bronchoscope. In the rabbit, vector DNA could be detected in the infected lobe up to 6 months after administration. A 26-amino acid polypeptide sequence unique to the recombinant AAV-CFTR protein was used to generate both oligonucleotide probes and a polyclonal antibody which allowed the unambiguous identification of vector RNA and CFTR protein expression. With these reagents, CFTR RNA and protein were detected in the airway epithelium of the infected lobe for up to 6 months after vector administration. AAV vectors do, therefore, efficiently promote in vivo gene transfer to the airway epithelium which is stable over several months. These findings indicate that AAV-CFTR vectors could potentially be very useful for gene therapy.