Paleyanda R K, Velander W H, Lee T K, Scandella D H, Gwazdauskas F C, Knight J W, Hoyer L W, Drohan W N, Lubon H
Plasma Derivatives Department Holland Laboratory, Rockville, MD 20855, USA.
Nat Biotechnol. 1997 Oct;15(10):971-5. doi: 10.1038/nbt1097-971.
Deficiency or abnormality of coagulation factor VIII (FVIII) causes a bleeding disorder called hemophilia A. Treatment involves FVIII concentrates prepared from pooled human plasma or recombinant FVIII (rFVIII) prepared from mammalian cell culture. The cost of highly purified FVIII or rFVIII is a major factor in hemophilia therapy and restricts prophylaxis. We have sought to generate a new source of rFVIII by targeting expression of the human FVIII cDNA to the mammary gland of transgenic pigs using the regulatory sequences of the mouse whey acidic protein gene. The identity of processed heterodimeric rFVIII was confirmed using specific antibodies, by thrombin digestion and activity assays. The secretion of as much as 2.7 micrograms/ml of rFVIII in milk was over tenfold higher than in normal plasma. Up to 0.62 U/ml of rFVIII was detected in an assay in which rFVIII restored normal clotting activity to FVIII-deficient human plasma.
凝血因子VIII（FVIII）的缺乏或异常会导致一种名为甲型血友病的出血性疾病。治疗方法包括使用从混合人血浆中制备的FVIII浓缩物或从哺乳动物细胞培养物中制备的重组FVIII（rFVIII）。高纯度FVIII或rFVIII的成本是血友病治疗的一个主要因素，并限制了预防措施。我们试图通过利用小鼠乳清酸性蛋白基因的调控序列将人FVIII cDNA的表达靶向转基因猪的乳腺，来产生一种新的rFVIII来源。使用特异性抗体、凝血酶消化和活性测定法确认了加工后的异二聚体rFVIII的身份。牛奶中rFVIII的分泌量高达2.7微克/毫升，比正常血浆高出十倍以上。在一项检测中，rFVIII恢复了FVIII缺陷型人血浆的正常凝血活性，检测到的rFVIII含量高达0.62 U/毫升。
