Jairath S, Vargas P B, Hamlin H A, Field A K, Kilkuskie R E
Hybridon, Inc., Worcester, MA 01605, USA.
Antiviral Res. 1997 Feb;33(3):201-13. doi: 10.1016/s0166-3542(96)01015-7.
Oligodeoxyribonucleotides targeted against respiratory syncytial virus (RSV) genomic RNA inhibited RSV replication in cell culture by an apparent antisense mechanism. HEp-2 cells were infected with RSV strain A2 and incubated in the presence of oligonucleotides. Virus replication was measured by enzyme-linked immunosorbent assay (ELISA), virus yield assay, or production of specific RSV mRNAs. Using ELISA, 50% effective concentration (EC50) values were about 0.5-1 microM for an antisense oligonucleotide targeted to the start of the NS2 gene. All oligonucleotides inhibited virus antigen production as measured by ELISA. In all assays, this antisense oligonucleotide was more potent than: (1) a control oligonucleotide containing the reverse sequence; (2) oligonucleotides targeted at RSV mRNA; (3) a random sequence oligonucleotide; and (4) ribavirin. Reverse transcriptase polymerase chain reaction (PT-PCR) showed sequence specific depletion of the genomic RNA target following treatment of cells with the antisense oligonucleotide. Specific cleavage of the genomic target RNA has been detected at the antisense oligonucleotide binding site, suggesting that cellular Rnase H participates in the reaction. These results indicate that antisense oligonucleotides targeted against RSV genomic RNA can effectively inhibit RSV replication and may have therapeutic value.
针对呼吸道合胞病毒（RSV）基因组RNA的寡脱氧核糖核苷酸通过一种明显的反义机制抑制了细胞培养中的RSV复制。用RSV A2株感染HEp-2细胞，并在寡核苷酸存在的情况下进行培养。通过酶联免疫吸附测定（ELISA）、病毒产量测定或特定RSV mRNA的产生来测量病毒复制。使用ELISA，针对NS2基因起始部位的反义寡核苷酸的50%有效浓度（EC50）值约为0.5 - 1微摩尔。通过ELISA测量，所有寡核苷酸均抑制病毒抗原的产生。在所有测定中，这种反义寡核苷酸比以下几种更有效：（1）含有反向序列的对照寡核苷酸；（2）针对RSV mRNA的寡核苷酸；（3）随机序列寡核苷酸；以及（4）利巴韦林。逆转录聚合酶链反应（PT-PCR）显示，在用反义寡核苷酸处理细胞后，基因组RNA靶标出现序列特异性缺失。在反义寡核苷酸结合位点检测到基因组靶标RNA的特异性切割，表明细胞核糖核酸酶H参与了该反应。这些结果表明，针对RSV基因组RNA的反义寡核苷酸可以有效抑制RSV复制，可能具有治疗价值。
