INHIBITION OF PROTEIN-KINASE C-ALPHA EXPRESSION IN MICE AFTER SYSTEMIC ADMINISTRATION OF PHOSPHOROTHIOATE ANTISENSE OLIGODEOXYNUCLEOTIDES
INHIBITION OF PROTEIN-KINASE C-ALPHA EXPRESSION IN MICE AFTER SYSTEMIC ADMINISTRATION OF PHOSPHOROTHIOATE ANTISENSE OLIGODEOXYNUCLEOTIDES
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DOI：
10.1073/pnas.91.24.11762
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发表时间：
1994-11-22
影响因子：
11.1
通讯作者：
MCKAY, R
中科院分区：
文献类型：
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作者：
DEAN, NM;MCKAY, R
设计用于与编码鼠蛋白激酶 C-α (PKC-α) 的 mRNA 的 AUG 翻译起始密码子杂交的 20 聚体硫代磷酸寡脱氧核苷酸可在体外和体内抑制 PKC-α 的表达。在小鼠 C127 乳腺上皮细胞中，PKC-α mRNA 表达的减少具有剂量和时间依赖性。寡脱氧核苷酸的 IC50 值为 100-200 nM，并可降低 PKC-α mRNA 表达长达 48 小时。与其他 PKC 同工酶（δ、ε 和 zeta）相比，这种减少对于 PKC-α 是特异性的，并且完全依赖于寡脱氧核苷酸序列。当对小鼠进行腹腔注射时，相同的寡脱氧核苷酸引起肝脏中 PKC-α mRNA 的剂量依赖性、寡脱氧核苷酸序列依赖性减少，IC50 值为 30-50 mg/kg 体重。单次 50 mg/kg 剂量后，表达抑制率为 64 +/- 11%。 PKC-delta、epsilon 和 zeta mRNA 的表达不受此处理的影响。体内寡脱氧核苷酸活性不需要阳离子脂质体或任何其他递送系统的存在，尽管在体外，寡脱氧核苷酸需要阳离子脂质体来抑制PKC-α表达。本研究证明了硫代磷酸寡脱氧核苷酸在全身给药后作为体内基因表达特异性抑制剂的效用。
A 20-mer phosphorothioate oligodeoxynucleotide designed to hybridize to the AUG translation initiation codon of mRNA encoding murine protein kinase C-alpha (PKC-alpha) inhibits the expression of PKC-alpha both in vitro and in vivo. In mouse C127 mammary epithelial cells, the reduction in PKC-alpha mRNA expression was both dose and time dependent. The oligodeoxynucleotide exhibited an IC50 value of 100-200 nM and reduced PKC-alpha mRNA expression for up to 48 hr. This reduction was specific for PKC-alpha versus other PKC isozymes (delta, epsilon, and zeta) and completely dependent upon oligodeoxynucleotide sequence. When administered intraperitoneally in mice, the same oligodeoxynucleotide caused a dose-dependent, oligodeoxynucleotide sequence-dependent reduction of PKC-alpha mRNA in liver, with an IC50 value of 30-50 mg/kg of body weight. Inhibition of expression was 64 +/- 11% after a single 50-mg/kg dose. The expression of PKC-delta, epsilon, and zeta mRNA was unaffected by this treatment. The oligodeoxynucleotide activity in vivo did not require the presence of cationic liposomes or any other delivery systems, although in vitro, the oligodeoxynucleotide required cationic liposomes for inhibition of PKC-alpha expression. This study demonstrates the utility of phosphorothioate oligodeoxynucleotides as specific inhibitors of gene expression in vivo after systemic administration.