Harvey B G, Hackett N R, Ely S, Crystal R G
Division of Pulmonary and Critical Care Medicine, Weill Medical College of Cornell University, New York, New York 10021, USA.
Mol Ther. 2001 Feb;3(2):206-15. doi: 10.1006/mthe.2000.0244.
Adenovirus (Ad)-mediated gene transfer to the respiratory epithelium of experimental animals and to nasal and airway epithelium of individuals with cystic fibrosis is followed by transient gene expression. Extensive studies in experimental animals are consistent with the concept that local cellular host anti-vector immune responses account for this short-term expression, and systemic and local [lung epithelial lining fluid (ELF)] anti-Ad neutralizing antibodies are generated following Ad vector administration to the respiratory epithelial surface. To determine if this paradigm holds in normal humans, a first-generation Ad vector (Ad(GV)CD.10, an E1(-)E3(-) Ad serotype 5-based vector coding for the Escherichia coli cytosine deaminase gene) was sprayed locally in escalating doses (8 x 10(8)-8 x 10(10) particle units (pu), n = 2/group) into the lung airway epithelium of six normal individuals. Serum, ELF, and endobronchial biopsies were obtained at baseline and at various time points following vector administration. In contrast to the observations in experimental animals in which lung administration of first-generation Ad vectors is followed by strong systemic and local host response, bronchial spray administration of the Ad vector to normal humans showed: (1) minimal inflammation in bronchial biopsies, bronchial brushing, and bronchoalveolar lavage fluid; (2) no blood lymphocyte proliferation in five of six individuals in response to in vitro stimulation with Ad antigens; and (3) no significant increase from baseline in blood or lung ELF anti-Ad neutralizing antibodies. Despite this minimal normal human anti-Ad host response, dose-dependent levels of vector DNA in the airway epithelium were transient. Vector DNA in the targeted airway epithelial cells peaked in a dose-dependent fashion at 0.007 to 1.1 copies/cell at day 7 and declined thereafter, reducing to <10% of peak levels by 2 weeks. These observations demonstrate both the strengths and the limits of using experimental animals to predict human responses to gene transfer vectors. While the transient nature of Ad vector persistence in the airway epithelium is predicted by most experimental animal studies, respiratory epithelial administration of first-generation Ad vectors at doses up to 10(10) pu to airway epithelium of healthy individuals elicits minimal to no detectable systemic and mucosal humoral and cellular immune responses, an observation diametrically opposed to the host responses measured in experimental animals. These findings suggest that, while adaptive anti-Ad immune responses likely play some role in the disappearance of the vector DNA following vector administration to the human lung, other mechanisms may also be involved in the response of humans to Ad gene transfer vectors.
腺病毒（Ad）介导的基因转移至实验动物的呼吸道上皮以及囊性纤维化患者的鼻腔和气道上皮后，会出现短暂的基因表达。在实验动物中的广泛研究与以下概念一致，即局部细胞宿主抗载体免疫反应导致了这种短期表达，并且在将Ad载体给予呼吸道上皮表面后会产生全身和局部[肺上皮衬液（ELF）]抗Ad中和抗体。为了确定这种模式在正常人类中是否成立，将第一代Ad载体（Ad(GV)CD.10，一种基于E1(-)E3(-)腺病毒血清型5的载体，编码大肠杆菌胞嘧啶脱氨酶基因）以递增剂量（8×10⁸ - 8×10¹⁰颗粒单位（pu），每组n = 2）局部喷入6名正常个体的肺气道上皮。在基线以及载体给药后的不同时间点采集血清、ELF和支气管活检样本。与在实验动物中的观察结果相反，在实验动物中向肺部给予第一代Ad载体后会引发强烈的全身和局部宿主反应，而向正常人类支气管喷雾给予Ad载体则显示：（1）支气管活检、支气管刷检和支气管肺泡灌洗液中的炎症轻微；（2）六名个体中有五名在受到Ad抗原体外刺激后血液淋巴细胞无增殖；（3）血液或肺ELF抗Ad中和抗体与基线相比无显著增加。尽管正常人类对Ad的宿主反应极小，但气道上皮中载体DNA的水平呈剂量依赖性且是短暂的。靶向气道上皮细胞中的载体DNA在第7天以剂量依赖性方式达到峰值，为0.007至1.1拷贝/细胞，此后下降，到2周时降至峰值水平的<10%。这些观察结果证明了使用实验动物预测人类对基因转移载体反应的优势和局限性。虽然大多数实验动物研究预测了Ad载体在气道上皮中持续存在的短暂性，但向健康个体的气道上皮给予高达10¹⁰ pu剂量的第一代Ad载体后，引发的全身和黏膜体液及细胞免疫反应极小甚至无法检测到，这一观察结果与在实验动物中测得的宿主反应截然相反。这些发现表明，虽然适应性抗Ad免疫反应可能在向人类肺部给予载体后载体DNA的消失中起一定作用，但其他机制也可能参与人类对Ad基因转移载体的反应。
