Anti-htt single chain antibodies as intrabodies may be useful for gene-therapy in polyglutamine disease

亨廷顿病（HD）是一种神经退行性疾病，其特征是亨廷顿病基因外显子1 CAG重复扩增。扩展的pdyglutamine编码的CAG重复序列诱导与HD病理相关的蛋白-蛋白相互作用，神经细胞死亡可能是由亨廷顿蛋白（htt）或其n端部分形成的聚谷氨酰胺聚集体造成的。利用重组n端亨廷顿蛋白（htt， 16个残基）与麦糖结合蛋白（MBF）或谷胱甘肽s -转移酶（GST）融合，从约1×10^<12>抗体的人噬菌体展示文库中筛选出1056个单链Fv （scFv）抗体。抗体与HD基因外显子1第二蛋氨酸编码的MKAFESLKSF(Q)6肽结合检测。五种scFv抗体特异性结合到htt的n端，并在HD细胞模型中进行了测试。在293细胞中使用htt蛋白表达来检测与htt n端部分的结合，扩展的聚谷氨酰胺延伸融合到绿色荧光蛋白（EGFP）和体内。发现两种抗htt scFv抗体在细胞系293细胞中抑制聚谷氨酰胺聚集。这些结果表明，在治疗神经退行性疾病，如多谷氨酰胺病、阿尔茨海默病、帕金森病和朊病毒疾病的基因疗法中，体内体将被证明是有用的。

Huntington disease (HD) is a neurodegenerative disorder characterized by the expansion of CAG repeats in exon 1 of the HD gene. Expanded pdyglutamine-encoded CAG repeats induce protein-protein interactions related to the pathology of HD, and neuronal cell death may result from the formation of polyglutamine aggregates by huntingtin (htt) or its N-terminal portion. A total of 1056 single-chain Fv (scFv) antibodies were selected from a human phage display library of approximately 1×10^<12>antibodies using recombinant N-terminal huntingtin (htt, 16 residues) fused to maltose-binding protein (MBF) or glutathione S-transferase (GST). Antibodies were tested for binding with the MKAFESLKSF(Q)6 peptide encoded from the second methionine of HD gene exon1. Five scFv antibodies specifically bound to the N-terminal of htt and were tested in a cellular model of HD. Protein expression of htt was used in 293 cells to test binding to the N-terminal portion of htt, with expanded potyglutamine stretches fused to green fluorescent protein (EGFP) and intrabodies. Two anti-htt scFv antibodies were found to inhibit polyglutamine aggregates in cell line, 293 cells. These results suggest that intrabodies will prove useful in genetherapies against neurodegerative disorders such as polyglutamine disease, Alzheimer's disease, Parkinson's disease and prion diseases.