Recombinant Human Adenovirus: Targeting to the Human Transferrin Receptor Improves Gene Transfer to Brain Microcapillary Endothelium
Recombinant Human Adenovirus: Targeting to the Human Transferrin Receptor Improves Gene Transfer to Brain Microcapillary Endothelium
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DOI：
10.1128/jvi.74.23.11359-11366.2000
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发表时间：
2000-12
影响因子：
5.4
通讯作者：
Haibin Xia;Brian Anderson;Q. Mao;B. Davidson
中科院分区：
文献类型：
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作者：
Haibin Xia;Brian Anderson;Q. Mao;B. Davidson
由于可溶性溶酶体酶缺乏而导致的先天性代谢错误可导致全身性神经退行性疾病。代表性的例子包括婴儿和婴儿晚期的类蜡质脂fuscinosis（分别为CLN1或CLN2缺乏症）和粘多糖病VII型（MPS VII），一种β-葡萄糖醛酸酶缺乏症。治疗这些疾病的中枢神经系统部分将需要广泛的蛋白质或酶替代，要么通过传播蛋白质，要么通过传播编码它的基因。我们假设重组病毒载体将脑微血管内皮（BME）转导，并在基底侧分泌酶产物，可以使酶广泛传播。要做到这一点，应该修改病毒以瞄准BME。这需要(i)鉴定bme驻留的目标受体，（ii）鉴定靶向该分子的基序，（iii）构建修饰的病毒以允许与目标受体结合，以及（iv）证明表达受体的细胞的转导。为了证明主要实验，我们选择了人类转铁蛋白受体（hTfR），这是一种在人类BME上高密度发现的分子。筛选了一个能结合hTfR的非命名噬菌体展示文库。对43个克隆进行了测序，其中大部分克隆含有AKxxK/R、KxKxPK/R或KxK基序。在腺病毒5型纤维的HI环中克隆了10个具有代表性的肽基序。所有测试的基序都保留了三聚体化和结合转铁蛋白受体的能力，其中7个基序允许重组腺病毒的产生。重要的是，纤维修饰的病毒促进了基因向表达hTfR的细胞系和表达高水平内源性受体的人脑微血管内皮的转移（2- 34倍）。我们的数据表明腺病毒可以在HI环中进行修饰，以扩大向hTfR的趋向性。
ABSTRACT Some inborn errors of metabolism due to deficiencies of soluble lysosomal enzymes cause global neurodegenerative disease. Representative examples include the infantile and late infantile forms of the ceroid lipofuscinoses (CLN1 or CLN2 deficiency, respectively) and mucopolysaccharidoses type VII (MPS VII), a deficiency of β-glucuronidase. Treatment of the central nervous system component of these disorders will require widespread protein or enzyme replacement, either through dissemination of the protein or through dissemination of a gene encoding it. We hypothesize that transduction of brain microcapillary endothelium (BME) with recombinant viral vectors, with secretion of enzyme product basolaterally, could allow for widespread enzyme dissemination. To achieve this, viruses should be modified to target the BME. This requires (i) identification of a BME-resident target receptor, (ii) identification of motifs targeted to that molecule, (iii) the construction of modified viruses to allow for binding to the target receptor, and (iv) demonstrated transduction of receptor-expressing cells. In proof of principal experiments, we chose the human transferrin receptor (hTfR), a molecule found at high density on human BME. A nonamer phage display library was panned for motifs which could bind hTfR. Forty-three clones were sequenced, most of which contained an AKxxK/R, KxKxPK/R, or KxK motif. Ten peptides representative of the three motifs were cloned into the HI loop of adenovirus type 5 fiber. All motifs tested retained their ability to trimerize and bind transferrin receptor, and seven allowed for recombinant adenovirus production. Importantly, the fiber-modified viruses facilitated increased gene transfer (2- to 34-fold) to hTfR expressing cell lines and human brain microcapillary endothelia expressing high levels of endogenous receptor. Our data indicate that adenoviruses can be modified in the HI loop for expanded tropism to the hTfR.